rowid,run.accession,experiment.accession,sample.accession,study.accession,bioproject,study.title,study.alias,study.type,study.abstract,study.attributes,study.PMIDs,sample.description,sample.title,sample.alias,sample.centername,sample.attributes,GEOsample.title,GEOsample.dataprocessing,GEOsample.source,GEOsample.treatmentprotocol,GEOsample.extractprotocol,GEOsample.growthprotocol,GEOsample.characteristics,GEOsample.accession,experiment.title,experiment.alias,experiment.library_name,experiment.design_description,experiment.library_construction_protocol,experiment.attributes,experiment.library_strategy,experiment.library_source,experiment.library_selection,experiment.library_layout,experiment.platform,experiment.instrument_model,experiment.spot_descriptor,experiment.study_ref,run.title,run.attributes,run.filename,run.semantic_name,run.total_bases,run.total_spots,run.alias,run.read_lengths,run.base_counts,run.r1_length,run.r2_length,run.r3_length,run.r4_length,run.Acount,run.Ccount,run.Gcount,run.Tcount,run.Ncount,run.experiment,run.pool_member,submission.accession,submission.srasource,submission.bioprojectsource,seqdetective.n_mates,seqdetective.mapping_rate.mate1,seqdetective.mapping_rate.mate2,seqdetective.nofeature_rate.mate1,seqdetective.nofeature_rate.mate2,seqdetective.sparsity.mate1,seqdetective.sparsity.mate2,seqdetective.pos_strand_rate.mate1,seqdetective.pos_strand_rate.mate2,seqdetective.readlen.mate1,seqdetective.readlen.mate2,seqdetective.judgement.mate1,seqdetective.judgement.mate2,seqdetective.judgement.reason,platform_family,instrument_generation,read_bias,selection_class,prep_kit,sc_or_bulk,tech_class,technology,tech_variant,submission.bioprojectsource.country,earliest_date,devstage_curation,devstage_curation_coarse,tissue_curation,tissue_curation_coarse
9892,ERR4172795,ERX4136409,ERS4580819,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sibling 3,SAMEA6853229,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853229|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sibling 3|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:wild type genotype|sample name:E MTAB 9113:sibling 3|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sibling 3 p,sibling 3 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:wild type genotype,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-sibs-3_S6_R1.fastq.gz UCLGNS1141-gfp-sibs-3_S6_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp sibs 3 S6 R,0:81 1:81,A:1144578958;C:1113109875;G:1138809559;T:1117038434;N:213932,81,81,,,1144578958,1113109875,1138809559,1117038434,213932,ERX4136409,ERS4580819,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.96684,0.96492,0.03244,0.0317,0.71236,0.71514,0.45871,0.46189,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
9893,ERR4172794,ERX4136408,ERS4580818,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sibling 2,SAMEA6853228,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853228|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sibling 2|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:wild type genotype|sample name:E MTAB 9113:sibling 2|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sibling 2 p,sibling 2 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:wild type genotype,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-sibs-2_S4_R1.fastq.gz UCLGNS1141-gfp-sibs-2_S4_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp sibs 2 S4 R,0:81 1:81,A:1056280368;C:1036355469;G:1045429486;T:1035773622;N:189389,81,81,,,1056280368,1036355469,1045429486,1035773622,189389,ERX4136408,ERS4580818,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.95511,0.95696,0.02941,0.02903,0.71492,0.71628,0.45765,0.46537,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
9894,ERR4172793,ERX4136407,ERS4580817,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sibling 1,SAMEA6853227,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853227|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sibling 1|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:wild type genotype|sample name:E MTAB 9113:sibling 1|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sibling 1 p,sibling 1 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:wild type genotype,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-sibs-1_S2_R1.fastq.gz UCLGNS1141-gfp-sibs-1_S2_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp sibs 1 S2 R,0:81 1:81,A:1121176859;C:1093927654;G:1108608188;T:1101082277;N:209264,81,81,,,1121176859,1093927654,1108608188,1101082277,209264,ERX4136407,ERS4580817,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.95795,0.96004,0.02885,0.02854,0.71648,0.71756,0.44348,0.43956,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
9895,ERR4172792,ERX4136406,ERS4580816,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sfpq 3,SAMEA6853226,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853226|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sfpq 3|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:sfpq / |sample name:E MTAB 9113:sfpq 3|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sfpq 3 p,sfpq 3 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:sfpq / ,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-neg_3_S5_R1.fastq.gz UCLGNS1141-gfp-neg_3_S5_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp neg 3 S5 R,0:81 1:81,A:845849249;C:772302798;G:911660213;T:787409272;N:154042,81,81,,,845849249,772302798,911660213,787409272,154042,ERX4136406,ERS4580816,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.96298,0.95486,0.03415,0.03476,0.7167,0.73503,0.4665,0.45947,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
9896,ERR4172791,ERX4136405,ERS4580815,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sfpq 2,SAMEA6853225,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853225|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sfpq 2|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:sfpq / |sample name:E MTAB 9113:sfpq 2|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sfpq 2 p,sfpq 2 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:sfpq / ,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-neg-2_S3_R1.fastq.gz UCLGNS1141-gfp-neg-2_S3_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp neg 2 S3 R,0:81 1:81,A:961156886;C:917011292;G:942713581;T:936109877;N:157724,81,81,,,961156886,917011292,942713581,936109877,157724,ERX4136405,ERS4580815,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.96276,0.96147,0.03436,0.03382,0.71892,0.72021,0.4581,0.46402,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
9897,ERR4172790,ERX4136404,ERS4580814,ERP121885,PRJEB38455,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E-MTAB-9113,Transcriptome Analysis,RNA was extracted from whole zebrafish embryos at 24 hpf to examine changes in gene expression and splicing in sfpq null mutants,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 05 22,,Protocols: Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,sfpq 1,SAMEA6853224,Centre for Developmental Neurobiology King's College London,ENA FIRST PUBLIC:2020 12 01T04:08:08Z|ENA LAST UPDATE:2020 05 22T17:13:15Z|External Id:SAMEA6853224|INSDC center name:Centre for Developmental Neurobiology King's College London|INSDC first public:2020 12 01T04:08:08Z|INSDC last update:2020 05 22T17:13:15Z|INSDC status:public|Submitter Id:E MTAB 9113:sfpq 1|age:24|broker name:ArrayExpress|common name:zebrafish|developmental stage:embryo|genotype:sfpq / |sample name:E MTAB 9113:sfpq 1|scientific name:Danio rerio,,,,,,,,,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,E MTAB 9113:sfpq 1 p,sfpq 1 p,RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,Embryos were collected from in crosses of sfpq+/  adult zebrafish RNA was extracted using the RNEasy Mini Kit Qiagen Library constructed with ribodepletion,Experimental Factor: genotype:sfpq / ,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina HiSeq 2500,,ERP121885,Illumina HiSeq 2500 paired end sequencing; RNA seq of sfpq /  zebrafish embryos and siblings at 24 hpf,ENA FIRST PUBLIC:2021 01 27|ENA LAST UPDATE:2020 06 19,UCLGNS1141-gfp-neg-1_S1_R1.fastq.gz UCLGNS1141-gfp-neg-1_S1_R2.fastq.gz,fastq fastq,,,E MTAB 9113:UCLGNS1141 gfp neg 1 S1 R,0:81 1:81,A:1046317524;C:1014946275;G:1036910299;T:1023619857;N:185237,81,81,,,1046317524,1014946275,1036910299,1023619857,185237,ERX4136404,ERS4580814,ERA2625401,Centre for Developmental Neurobiology King,Centre for Developmental Neurobiology King,2,0.96015,0.96173,0.03114,0.03098,0.7175,0.71865,0.46666,0.46399,81,81,B,B,biological fallback assumption,illumina,hiseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,United Kingdom,2020-05-22,Multi-stage,Multi-stage,Embryo Imprecise,All anatomical structures
11821,ERR11834690,ERX11232843,ERS16254762,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 4,SAMEA114265804,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265804|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 4|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250|organism part:whole organism|rin:9.6|sample name:E MTAB 13263:Sample 4|scientific name:Danio rerio|sex:na|stimulus:immunosuppression|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 4 p,Sample 4 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2113_dre_C1_clobetasolpropionate_lib667285_10169_3_2.fastq.gz NG-32417_R2113_dre_C1_clobetasolpropionate_lib667285_10169_3_1.fastq.gz,fastq fastq,9764006696.0,32331148.0,E MTAB 13263:NG 32417 R2113 dre C1 clobetasolpropionate lib667285 10169 3 ,0:151 1:151,A:2627275848;C:2223323771;G:2323190525;T:2590103027;N:113525,151,151,,,2627275848,2223323771,2323190525,2590103027,113525,ERX11232843,ERS16254762,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95424,0.95502,0.12596,0.12401,0.65884,0.65825,0.48134,0.48327,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11822,ERR11834689,ERX11232842,ERS16254761,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 3,SAMEA114265803,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265803|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 3|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:0|organism part:whole organism|rin:9.4|sample name:E MTAB 13263:Sample 3|scientific name:Danio rerio|sex:na|stimulus:n1|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 3 p,Sample 3 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2112_dre_nc_clobetasolpropionate_lib673071_10180_1_2.fastq.gz NG-32417_R2112_dre_nc_clobetasolpropionate_lib673071_10180_1_1.fastq.gz,fastq fastq,13326663550.0,44128025.0,E MTAB 13263:NG 32417 R2112 dre nc clobetasolpropionate lib673071 10180 1 ,0:151 1:151,A:3641640344;C:3020489123;G:3072643196;T:3591659412;N:231475,151,151,,,3641640344,3020489123,3072643196,3591659412,231475,ERX11232842,ERS16254761,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95271,0.95293,0.11945,0.1151,0.65819,0.6579,0.47257,0.47803,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11823,ERR11834684,ERX11232837,ERS16254756,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 1,SAMEA114265798,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265798|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 1|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:0|organism part:whole organism|rin:9.3|sample name:E MTAB 13263:Sample 1|scientific name:Danio rerio|sex:na|stimulus:n1|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 1 p,Sample 1 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2110_dre_nc_clobetasolpropionate_lib667282_10169_3_1.fastq.gz NG-32417_R2110_dre_nc_clobetasolpropionate_lib667282_10169_3_2.fastq.gz,fastq fastq,10152616370.0,33617935.0,E MTAB 13263:NG 32417 R2110 dre nc clobetasolpropionate lib667282 10169 3 ,0:151 1:151,A:2743478864;C:2319134955;G:2378048842;T:2711839167;N:114542,151,151,,,2743478864,2319134955,2378048842,2711839167,114542,ERX11232837,ERS16254756,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95273,0.95457,0.13567,0.13174,0.65896,0.65841,0.47947,0.47815,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11824,ERR11834691,ERX11232844,ERS16254763,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 5,SAMEA114265805,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265805|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 5|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250|organism part:whole organism|rin:9.2|sample name:E MTAB 13263:Sample 5|scientific name:Danio rerio|sex:na|stimulus:immunosuppression|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 5 p,Sample 5 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2114_dre_C1_clobetasolpropionate_lib667286_10169_3_1.fastq.gz NG-32417_R2114_dre_C1_clobetasolpropionate_lib667286_10169_3_2.fastq.gz,fastq fastq,11690304334.0,38709617.0,E MTAB 13263:NG 32417 R2114 dre C1 clobetasolpropionate lib667286 10169 3 ,0:151 1:151,A:3142106866;C:2677986743;G:2767050957;T:3103028799;N:130969,151,151,,,3142106866,2677986743,2767050957,3103028799,130969,ERX11232844,ERS16254763,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95418,0.95523,0.12548,0.12397,0.66026,0.66016,0.46767,0.47253,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11825,ERR11834688,ERX11232841,ERS16254760,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 2,SAMEA114265802,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265802|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 2|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:0|organism part:whole organism|rin:9.6|sample name:E MTAB 13263:Sample 2|scientific name:Danio rerio|sex:na|stimulus:n1|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 2 p,Sample 2 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2111_dre_nc_clobetasolpropionate_lib673070_10180_1_1.fastq.gz NG-32417_R2111_dre_nc_clobetasolpropionate_lib673070_10180_1_2.fastq.gz,fastq fastq,11827163184.0,39162792.0,E MTAB 13263:NG 32417 R2111 dre nc clobetasolpropionate lib673070 10180 1 ,0:151 1:151,A:3256286612;C:2657918302;G:2701905502;T:3210846546;N:206222,151,151,,,3256286612,2657918302,2701905502,3210846546,206222,ERX11232841,ERS16254760,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95203,0.95292,0.12277,0.11772,0.66048,0.65837,0.47243,0.4729,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11826,ERR11834693,ERX11232846,ERS16254765,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 7,SAMEA114265807,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265807|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 7|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:4000|organism part:whole organism|rin:9.7|sample name:E MTAB 13263:Sample 7|scientific name:Danio rerio|sex:na|stimulus:immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 7 p,Sample 7 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2116_dre_nc_imq_clobetasolpropionate_lib667288_10169_3_1.fastq.gz NG-32417_R2116_dre_nc_imq_clobetasolpropionate_lib667288_10169_3_2.fastq.gz,fastq fastq,11114382784.0,36802592.0,E MTAB 13263:NG 32417 R2116 dre nc imq clobetasolpropionate lib667288 10169 3 ,0:151 1:151,A:2997376944;C:2547742011;G:2613273908;T:2955864850;N:125071,151,151,,,2997376944,2547742011,2613273908,2955864850,125071,ERX11232846,ERS16254765,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95392,0.9539,0.12951,0.12707,0.65628,0.65731,0.48132,0.48179,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11827,ERR11834695,ERX11232848,ERS16254767,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 9,SAMEA114265809,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265809|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 9|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:4000|organism part:whole organism|rin:9.9|sample name:E MTAB 13263:Sample 9|scientific name:Danio rerio|sex:na|stimulus:immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 9 p,Sample 9 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2118_dre_nc_imq_clobetasolpropionate_lib667290_10169_3_1.fastq.gz NG-32417_R2118_dre_nc_imq_clobetasolpropionate_lib667290_10169_3_2.fastq.gz,fastq fastq,9422294300.0,31199650.0,E MTAB 13263:NG 32417 R2118 dre nc imq clobetasolpropionate lib667290 10169 3 ,0:151 1:151,A:2541445212;C:2157798531;G:2214381344;T:2508560588;N:108625,151,151,,,2541445212,2157798531,2214381344,2508560588,108625,ERX11232848,ERS16254767,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95337,0.95433,0.13046,0.12718,0.65845,0.65815,0.48198,0.48414,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11828,ERR11834687,ERX11232840,ERS16254759,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 12,SAMEA114265801,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265801|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 12|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250 CP + 4000 IMQ|organism part:whole organism|rin:9.7|sample name:E MTAB 13263:Sample 12|scientific name:Danio rerio|sex:na|stimulus:immunosuppression and immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 12 p,Sample 12 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2121_dre_C1_imq_clobetasolpropionate_lib667293_10169_3_2.fastq.gz NG-32417_R2121_dre_C1_imq_clobetasolpropionate_lib667293_10169_3_1.fastq.gz,fastq fastq,9285032280.0,30745140.0,E MTAB 13263:NG 32417 R2121 dre C1 imq clobetasolpropionate lib667293 10169 3 ,0:151 1:151,A:2504765552;C:2110325477;G:2196928032;T:2472905759;N:107460,151,151,,,2504765552,2110325477,2196928032,2472905759,107460,ERX11232840,ERS16254759,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95411,0.95451,0.12502,0.12228,0.65829,0.65774,0.48259,0.47946,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11829,ERR11834694,ERX11232847,ERS16254766,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 8,SAMEA114265808,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265808|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 8|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:4000|organism part:whole organism|rin:9.5|sample name:E MTAB 13263:Sample 8|scientific name:Danio rerio|sex:na|stimulus:immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 8 p,Sample 8 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2117_dre_nc_imq_clobetasolpropionate_lib667289_10169_3_1.fastq.gz NG-32417_R2117_dre_nc_imq_clobetasolpropionate_lib667289_10169_3_2.fastq.gz,fastq fastq,10247527118.0,33932209.0,E MTAB 13263:NG 32417 R2117 dre nc imq clobetasolpropionate lib667289 10169 3 ,0:151 1:151,A:2766675319;C:2342307825;G:2405294253;T:2733131648;N:118073,151,151,,,2766675319,2342307825,2405294253,2733131648,118073,ERX11232847,ERS16254766,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.9541,0.95434,0.13364,0.13098,0.6576,0.6565,0.48068,0.48029,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11830,ERR11834692,ERX11232845,ERS16254764,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 6,SAMEA114265806,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265806|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 6|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250|organism part:whole organism|rin:10|sample name:E MTAB 13263:Sample 6|scientific name:Danio rerio|sex:na|stimulus:immunosuppression|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 6 p,Sample 6 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2115_dre_C1_clobetasolpropionate_lib667287_10169_3_1.fastq.gz NG-32417_R2115_dre_C1_clobetasolpropionate_lib667287_10169_3_2.fastq.gz,fastq fastq,10440673634.0,34571767.0,E MTAB 13263:NG 32417 R2115 dre C1 clobetasolpropionate lib667287 10169 3 ,0:151 1:151,A:2805459166;C:2397120759;G:2463616298;T:2774359989;N:117422,151,151,,,2805459166,2397120759,2463616298,2774359989,117422,ERX11232845,ERS16254764,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95568,0.9548,0.13056,0.12563,0.65882,0.66332,0.49162,0.48586,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11831,ERR11834685,ERX11232838,ERS16254757,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 10,SAMEA114265799,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265799|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 10|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250 CP + 4000 IMQ|organism part:whole organism|rin:9.4|sample name:E MTAB 13263:Sample 10|scientific name:Danio rerio|sex:na|stimulus:immunosuppression and immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 10 p,Sample 10 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2119_dre_C1_imq_clobetasolpropionate_lib667291_10169_3_2.fastq.gz NG-32417_R2119_dre_C1_imq_clobetasolpropionate_lib667291_10169_3_1.fastq.gz,fastq fastq,11267068246.0,37308173.0,E MTAB 13263:NG 32417 R2119 dre C1 imq clobetasolpropionate lib667291 10169 3 ,0:151 1:151,A:3042432765;C:2574394239;G:2642086000;T:3008026288;N:128954,151,151,,,3042432765,2574394239,2642086000,3008026288,128954,ERX11232838,ERS16254757,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.95345,0.95411,0.13324,0.13062,0.65796,0.65892,0.48176,0.48174,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
11832,ERR11834686,ERX11232839,ERS16254758,ERP150242,PRJEB65099,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E-MTAB-13263,Transcriptome Analysis,Within the regulatory framework on the approval of new substances  the assessment of immunotoxic modes of action MoA is currently not covered. This is not least due to the lack of standardized methods and reliable validated biomarkers for immunotoxic effects. The experimental set up was designed to analyze the compound specific response of zebrafish embryos to two immunomodulative reference substances  clobetasol propionate CP  CAS 25122 46 7 and imiquimod IMQ  CAS 99011 02 6  on the global level of gene expression. The obtained results were used to i identify potential biomarker candidates for the assessment of immunotoxic MoAs  ii identify compound specific molecular signatures  iii evaluate the reliability of data acquisition using OMICs coupled approaches and iv to assess the suitability of the zebrafish embryo as an alternative model for the human representative investigation of psoriatic effects. For this  the transcriptomic profiles of embryos were bioinformatically analysed subsequent to an CP induced immunosuppression in absence or presence of an IMQ induced immune challenge  i.e. the simulation of a resting and an activated immune system.   In a modified version of the zebrafish embryo toxicity test OECD 236  15 fertilized eggs were exposed to either CP 250 nM or to IMQ 4000 nM or to a combination of both under semi static conditions. Exposure to CP started at 2 hpf until 72 hpf. Exposure to IMQ started at 48 hpf until 72 hpf. Untreated embryos reared in medium without xxx nor IMQ dissolved was used as a negative control. All conditions comprised three biological replicates. Medium was exchanged on a daily basis by renewing half its volume. Occurrence of morphological changes and indicators of lethality as described in the OECD test guideline 236 were examined microscopically on a daily basis. At 72 hpf  all larvae were pooled for each sample for RNA extraction using a NucleoSpin RNA/Protein kit Macherey Nagel following the manufacturer's protocol. RNA quality was assessed with a 2100 Bioanalyzer system Agilent before coding RNA was purified PolyA selection with TruSeq RNA Library Prep Kit v2 and sequenced on an Illumina NovaSeq 6000 System Illumina in 150 bp paired end mode  producing a minimum of 30 million reads per sample. Adapter sequences were removed with trimmomatic and sequences were aligned to the Danio rerio reference genome GRCz11 with STAR. Counting of feature mapped reads was performed through featureCounts. Library gene count tables were then merged to a single count matrix as input for differential gene expression analysis with DESeq2.,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,,Protocols: At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos. For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”. Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use. Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol” Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions. The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment. In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution. RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice. High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,Sample 11,SAMEA114265800,"Department Ecotoxicogenomics, Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Schmallenberg, Germany",ENA FIRST PUBLIC:2023 12 01T00:42:51Z|ENA LAST UPDATE:2023 12 01T00:42:51Z|External Id:SAMEA114265800|INSDC center name:Department Ecotoxicogenomics  Fraunhofer Institute for Molecular Biology and Applied Ecology IME  Schmallenberg  Germany|INSDC first public:2023 12 01T00:42:51Z|INSDC last update:2023 12 01T00:42:51Z|INSDC status:public|Submitter Id:E MTAB 13263:Sample 11|age:3|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|genotype:AB|geographic location country and/or sea:not collected|isolate:not applicable|nominal concentration in medium:250 CP + 4000 IMQ|organism part:whole organism|rin:10|sample name:E MTAB 13263:Sample 11|scientific name:Danio rerio|sex:na|stimulus:immunosuppression and immunostimulation|strain:Wildtype AB,,,,,,,,,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,E MTAB 13263:Sample 11 p,Sample 11 p,Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,At 72 hpf  for each sample  all 15 zebrafish embryos were picked from the glass well and carefully transferred to a 1.5 ml low binding Eppendorf tube. Excessive liquid was removed and tubes were placed on ice to euthanize the embryos.   For detailed information about embryonic incubation conditions and husbandry of adult broodstock please refer to the “Growth protocol”. For detailed information about exposure duration and conditions please refer to the “Treatment protocol”.  Adult animal husbandry: All experiments were performed using zebrafish Danio rerio embryos of the wild type strain AB. Adult fish were originally obtained from West Aquarium GmbH  Germany  and were kept in 150 l tanks under flow through conditions at 26 ± 2 °C and a 12:12 light/dark cycle at the Fraunhofer Institute for molecular biology and applied ecology IME  Schmallenberg  Germany. TetraMin® Tetra Werke  Germany was used as daily main feed  regularly supplemented with nauplii of Artemia salina. Fish were constantly bred for several generations without xxx between tanks  making the embryos deriving from different spawning groups tanks independent biological replicates. For egg collection  glass spawning trays equipped with whirled cords of green glass pearls to stimulate spawning and covered with wired mesh to prevent cannibalism were put into the tanks the day before tests were started OECD  2013. The morning post  eggs were collected from the trays using a sieve  rinsed with pre tempered system water to remove remaining feces  gathered in glass jars filled with pre tempered aerated system water and stored at a controlled temperature of 26 ± 1 °C until further use.  Fish embryo incubation: Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. For details on the treatment see “treatment protocol”   Clobetasol propionate 21 Chlor 9 fluor 11β 17 dihydroxy 16β methylpregna 1 4 dien 3 20 dion 17 propionat  purity ≥ 98 %  CAS no. 25122 46 7 and Imiquimod 1 2 methylpropyl 1H imidazo[4 5 c]quinoline 4 amine  purity ≥ 95 %  CAS no. 9901 02 6 were purchased from BIOMOL GmbH  Hamburg  Germany. For IMQ  a stock solution in acetone with a nominal concentration of 0.1 mg/ml was prepared. For CP  a stock solution in dimethyl sulfoxide DMSO with a nominal concentration of 2.5 mM was prepared. To obtain a test solution with a nominal concentration of 4666 nM IMQ  2242.5 µl of the IMQ stock solution was added to a glass bottle. To equalize the amount of added acetone amongst all solutions  equal volume of pure acetone without xxx was added to glass bottles for all remaining test solutions. For all test solutions containing 250 nM CP  20 µl of the CP stock solution was added to the glass bottles post complete evaporation of the acetone and filled with copper reduced tap water in the following referred to as cu red water to a final volume of 200 ml. Test solutions without xxx were prepared alike adding 20 µl DMSO without xxx. All solutions were sonicated for 30 min at room temperature Sonorex RK 100H  Bandelin  Germany. Final solutions were thus cu red water with 0.01 % DMSO  4666 nM IMQ in cu red water with 0.01 % DMSO  250 nM CP in cu red water with 0.01 % DMSO and 4666 nM IMQ plus 250 nM CP in cu red water with 0.01 % DMSO. Due to a mandatory dilution during the methodological procedure a nominal working concentration of 4000 nM was achieved for IMQ in these solutions.  The test solutions were freshly prepared one day prior the start of the experiment and stored at room temperature in the dark. They were constantly aerated and used over the course of the 72 hours exposure experiment. The glass wells for the incubation of the embryos were pre saturated with the respective test solution overnight and renewed at the beginning of the experiment.  In order to minimize the differences between replicates and conditions in exposure durations  the selection of embryos was performed in two steps: at 2 hpf  approx. 50 embryos from spawning batches with a minimum fertilization rate ≥ 90 % were transferred to previously overnight saturated glass petri dishes 6 cm diameter refilled with 7 ml of the corresponding test solutions for each replicate and condition. Embryos of the control and the IMQ condition were thus transferred to medium without xxx compound  embryos of the CP and combined CP+IMQ condition were transferred to medium supplemented with 250 nM CP. Once in solution  15 healthy and fertilized embryos  indicated by a well shaped blastodisc between the 128 cell and 1k cell stage of development were selected per replicate and condition for experiments. Embryos were incubated at a controlled temperature of 26 ± 1°C and a 14:10 h light/dark cycle artificially created with tubular fluorescent lamps with the lights switched on at 8 am. Medium was refreshed at 24 hpf by renewing half its volume. Embryos were inspected microscopically on a daily basis and at the end of the experiment using an OZL 451 stereomicroscope KERN & Sohn  Germany in order to monitor the occurrence of indicators of lethality as defined by the OECD FET test guideline  namely coagulation  lack of somite formation  lack of a heartbeat  non detachment of the tail OECD  2013 as well as further indicators of malformations such as scoliosis  formation of edema or lack of pigmentation. At 47 hpf  embryos were manually dechorionated using two pointy forceps. At 48 hpf  the immune challenge was started by transferring the embryos of the IMQ condition and the combined CP+IMQ condition to the respective test solution supplemented with IMQ. For this and in order to equalized the subsequent dilution of media between replicates and conditions  embryos were gathered in 1 ml of the aged medium and transferred with this volume into newly saturated glass dishes  filled with 6 ml of the corresponding fresh medium. Due to this dilution  a nominal working concentration of 4000 nM IMQ was achieved for the immune challenge. Embryos of the control condition and the CP condition were handled alike but without xxx the type of test solution.  RNA was extracted using a NucleoSpin RNA/Protein kit Macherey Nagel  Düren  Germany. Briefly  frozen embryos were transferred to screw cap tubes filled with 350 µl kit specific lysis buffer RP1 buffer and Lysing Matrix D ceramic beads MP Biomedicals  Irvine  USA and homogenized using a FastPrep 24 MP Biomedicals  Irvine  USA device at 5 m/s for 1 min. RP1 buffer was freshly prepared with TCEP as reducing agent prior extraction. Further steps were performed following the manufacturer's extraction protocol. As quality measures  purity and concentration of extracted RNA were determined using a NanoDrop 2000 device Thermo Scientific  Waltham  USA and RNA integrity numbers RIN were determined using a Bioanalyzer 2100 device Agilent  Santa Clara  USA with samples prepared using a RNA Pico 6000 kit Agilent  Santa Clara  USA following the manufacturer's instructions. Only samples with a RIN > 9 were used for downstream analysis. Samples were stored at  80°C until they were send to the sequencing facility on dry ice.  High quality RNA samples with RIN > 9 were selected for RNA seq library preparation and sequencing  performed by Eurofins Genomics Germany GmbH  Ebersberg  Germany. Briefly  Poly A containing protein coding mRNA molecules were purified from total RNA  randomly fragmentated and subjected to random primed strand specific cDNA library preparation using the TruSeq RNA library Prep Kit v2 Illumina following the manufacturer's instructions.,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,ERP150242,Illumina NovaSeq 6000 paired end sequencing; Transcriptomic profiling of  TLR 7 mediated immune challenge in zebrafish embryos in the presence and absence of glucocorticoid induced immunosuppression,ENA FIRST PUBLIC:2023 12 01|ENA LAST UPDATE:2023 12 01,NG-32417_R2120_dre_C1_imq_clobetasolpropionate_lib667292_10169_3_2.fastq.gz NG-32417_R2120_dre_C1_imq_clobetasolpropionate_lib667292_10169_3_1.fastq.gz,fastq fastq,9922486330.0,32855915.0,E MTAB 13263:NG 32417 R2120 dre C1 imq clobetasolpropionate lib667292 10169 3 ,0:151 1:151,A:2673462020;C:2268840894;G:2345658284;T:2634411815;N:113317,151,151,,,2673462020,2268840894,2345658284,2634411815,113317,ERX11232839,ERS16254758,ERA26742386,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,2,0.9551,0.95621,0.1193,0.11675,0.65833,0.65827,0.48596,0.48547,151,151,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,trueseq,bulk,unknown,unknown,,United Kingdom,2023-12-01,Multi-stage,Multi-stage,Whole Organism,All anatomical structures
36227,SRR33613672,SRX28842027,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD 3,HFD 3,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD-3-1.fq.gz HFD-3-2.fq.gz,fastq fastq,6810755100.0,22702517.0,HFD 3 1.fq.gz,0:150 1:150,A:1804127778;C:1596175018;G:1609090701;T:1801361603;N:0,150,150,,,1804127778,1596175018,1609090701,1801361603,0,SRX28842027,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36228,SRR33613673,SRX28842026,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD 2,HFD 2,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD-2-1.fq.gz HFD-2-2.fq.gz,fastq fastq,6826358400.0,22754528.0,HFD 2 1.fq.gz,0:150 1:150,A:1826465185;C:1582322712;G:1596188986;T:1821381517;N:0,150,150,,,1826465185,1582322712,1596188986,1821381517,0,SRX28842026,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36229,SRR33613674,SRX28842025,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD 1,HFD 1,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD-1-1.fq.gz HFD-1-2.fq.gz,fastq fastq,6598410300.0,21994701.0,HFD 1 1.fq.gz,0:150 1:150,A:1765905584;C:1529581410;G:1542056821;T:1760866485;N:0,150,150,,,1765905584,1529581410,1542056821,1760866485,0,SRX28842025,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36230,SRR33613675,SRX28842024,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD10 3,HFD10 3,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD10-3-1.fq.gz HFD10-3-2.fq.gz,fastq fastq,6609153300.0,22030511.0,HFD10 3 1.fq.gz,0:150 1:150,A:1780787716;C:1521027085;G:1532800826;T:1774537673;N:0,150,150,,,1780787716,1521027085,1532800826,1774537673,0,SRX28842024,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36231,SRR33613676,SRX28842023,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD10 2,HFD10 2,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD10-2-1.fq.gz HFD10-2-2.fq.gz,fastq fastq,6605145900.0,22017153.0,HFD10 2 1.fq.gz,0:150 1:150,A:1765731508;C:1533254667;G:1545960098;T:1760199627;N:0,150,150,,,1765731508,1533254667,1545960098,1760199627,0,SRX28842023,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36232,SRR33613677,SRX28842022,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,HFD10 1,HFD10 1,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,HFD10-1-1.fq.gz HFD10-1-2.fq.gz,fastq fastq,6780056700.0,22600189.0,HFD10 1 1.fq.gz,0:150 1:150,A:1819443100;C:1566081115;G:1580525465;T:1814007020;N:0,150,150,,,1819443100,1566081115,1580525465,1814007020,0,SRX28842022,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36233,SRR33613678,SRX28842021,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,NFD 3,NFD 3,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,NFD-3-1.fq.gz NFD-3-2.fq.gz,fastq fastq,6677775900.0,22259253.0,NFD 3 1.fq.gz,0:150 1:150,A:1794821611;C:1541478212;G:1552820105;T:1788655972;N:0,150,150,,,1794821611,1541478212,1552820105,1788655972,0,SRX28842021,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36234,SRR33613679,SRX28842020,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,NFD 2,NFD 2,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,NFD-2-1.fq.gz NFD-2-2.fq.gz,fastq fastq,6680197200.0,22267324.0,NFD 2 1.fq.gz,0:150 1:150,A:1810744167;C:1526383805;G:1539949310;T:1803119918;N:0,150,150,,,1810744167,1526383805,1539949310,1803119918,0,SRX28842020,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
36235,SRR33613680,SRX28842019,SRS25072222,SRP586119,PRJNA1263917,Danio rerio Transcriptome or Gene expression,PRJNA1263917,Other,Analyze the effect of TB KW fermentation on gene expression in high lipid zebrafish.,,,,Danio rerio,Danio rerio,,strain:missing|isolate:missing|breed:missing|cultivar:missing|ecotype:missing|age:7 day|dev stage:Juvenile fish|collection date:2024 04 16|geo loc name:China: Hang Zhou|sex:missing|tissue:Whole body|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of Danio rerio,NFD 1,NFD 1,Total RNA was extracted  digested with DNase I  and mRNA enriched. post fragmentation  cDNA synthesis  and library construction  the final single stranded DNA nanoball library was generated for sequencing via rolling circle replication.,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,DNBSEQ,DNBSEQ-T7,,SRP586119,,,NFD-1-1.fq.gz NFD-1-2.fq.gz,fastq fastq,6587923800.0,21959746.0,NFD 1 1.fq.gz,0:150 1:150,A:1773646514;C:1516320091;G:1530275621;T:1767681574;N:0,150,150,,,1773646514,1516320091,1530275621,1767681574,0,SRX28842019,SRS25072222,SRA2131918,Yibin University|Faculty of Quality Management and Inspection & Qua,Yibin University,,,,,,,,,,,,B,B,biological fallback assumption,bgi,bgi,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2025-05-18,Multi-stage,Multi-stage,Trunk,Surface Structure
77261,SRR25434203,SRX21168298,SRS18429832,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,dmrt / 2,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 6|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,dmrt2,dmrt2,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,dmrt2_R1_.fastq.gz dmrt2_R2.fastq.gz,fastq fastq,5751045000.0,19170150.0,dmrt2 R1 .fastq.gz,0:150 1:150,A:1488645455;C:1371887358;G:1405356847;T:1485112991;N:42349,150,150,,,1488645455,1371887358,1405356847,1485112991,42349,SRX21168298,SRS18429832,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.91589,0.91745,0.01703,0.01705,0.7637,0.76402,0.47113,0.46945,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System
77262,SRR25434204,SRX21168297,SRS18429831,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,dmrt / 1,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 5|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,dmrt1,dmrt1,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,dmrt1_R2.fastq.gz dmrt1_R1.fastq.gz,fastq fastq,7183531200.0,23945104.0,dmrt1 R1.fastq.gz,0:150 1:150,A:1847909835;C:1724710101;G:1769658622;T:1841199904;N:52738,150,150,,,1847909835,1724710101,1769658622,1841199904,52738,SRX21168297,SRS18429831,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.92259,0.92339,0.01692,0.01663,0.76558,0.76589,0.46917,0.46678,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System
77263,SRR25434205,SRX21168296,SRS18429830,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,cyp17a1 / ;dmrt1 / 2,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 4|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,dko2,dko2,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,dko2_R2.fastq.gz dko2_R1.fastq.gz,fastq fastq,6765257700.0,22550859.0,dko2 R1.fastq.gz,0:150 1:150,A:1746873541;C:1617601783;G:1668365344;T:1732367309;N:49723,150,150,,,1746873541,1617601783,1668365344,1732367309,49723,SRX21168296,SRS18429830,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.90858,0.90858,0.02015,0.01977,0.7612,0.76213,0.46791,0.47242,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System
77264,SRR25434206,SRX21168295,SRS18429829,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,cyp17a1 / ;dmrt1 / 1,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 3|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,dko1,dko1,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,dko1_R1.fastq.gz dko1_R2.fastq.gz,fastq fastq,6638354100.0,22127847.0,dko1 R1.fastq.gz,0:150 1:150,A:1713258874;C:1593272953;G:1635070441;T:1696702837;N:48995,150,150,,,1713258874,1593272953,1635070441,1696702837,48995,SRX21168295,SRS18429829,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.9182,0.91984,0.022,0.02186,0.76118,0.76167,0.47656,0.48847,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System
77265,SRR25434207,SRX21168294,SRS18429828,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,Control2,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 2|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,conf2,conf2,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,conf2_R1.fastq.gz conf2_R2.fastq.gz,fastq fastq,6223727700.0,20745759.0,conf2 R1.fastq.gz,0:150 1:150,A:1590221936;C:1500196214;G:1554765877;T:1578496570;N:47103,150,150,,,1590221936,1500196214,1554765877,1578496570,47103,SRX21168294,SRS18429828,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.92354,0.92626,0.01495,0.01561,0.76398,0.76418,0.47103,0.46492,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System
77266,SRR25434208,SRX21168293,SRS18429827,SRP451673,PRJNA999043,The transcription analysis of gonad in zebrafish,PRJNA999043,Other,To screen out the female biased genes,,,,,Control1,,isolate:not applicable|age:80days|dev stage:Adult|collection date:2021|geo loc name:China:Wuhan|sex:female|tissue:Gonad|replicate:replicate = biological replicate 1|BioSampleModel:Model organism or animal,,,,,,,,,RNA seq of gonad,conf1,conf1,screen out the key genes,,,RNA-Seq,TRANSCRIPTOMIC,PolyA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP451673,,,conf1_R1.fastq.gz conf1_R2.fastq.gz,fastq fastq,6799767600.0,22665892.0,conf1 R1.fastq.gz,0:150 1:150,A:1740276620;C:1642790185;G:1671812697;T:1744859854;N:28244,150,150,,,1740276620,1642790185,1671812697,1744859854,28244,SRX21168293,SRS18429827,SRA1681175,Chinese Academy of Sciences|Institute of Hydrology,Chinese Academy of Sciences,2,0.93305,0.93328,0.01522,0.01517,0.76477,0.76627,0.47519,0.47422,150,150,B,B,biological fallback assumption,illumina,novaseq_era,unknown,poly_a,unknown,bulk,unknown,unknown,,China,2023-07-27,Multi-stage,Multi-stage,Gonad,Reproductive System