{"database": "metadata", "table": "run_metadata", "rows": [[36065, "SRR33444777", "SRX28685517", "SRS24952420", "SRP583274", "PRJNA1259243", "Structure Dependent Developmental Toxicity of Alkyl Substituted Naphthalenes", "GSE296410", "Transcriptome Analysis", "Naphthalene and its alkyl substituted derivatives are among the most abundant polycyclic aromatic hydrocarbons PAHs in environmental and human exposure studies  yet their developmental toxicity and mode of action remain poorly understood due to challenges in testing of semi volatile compounds. This study developed a vial based high through put method to effectively assess the activity of naphthalenes and a set of 24 alkyl substituted naphthalenes. Early life stage zebrafish were exposed to a concentration series of each chemical 0 50 \u00b5M in rotating sealed glass vials to minimize volatilization. Benchmark concentration BMC50 values were calculated for morphological endpoints and lowest effect levels were determined for behavioral effects. The data were assessed for evidence of a narcotic mode of action using body burden measurements for select chemicals and logKow modeling. Targeted transcriptomics at a single concentration and timepoint as well as in silico molecular docking were conducted to generate mode of action hypothesis. The vial method enabled detection of highly variable developmental toxicity not previously observed using standard 96 well plate exposures. LogKow and body burden were poor predictors of toxicity  suggesting a non narcotic mode of action. Transcriptomic analysis revealed a limited but notable set of differentially expressed genes  with evidence for the disruption of glucocorticoid signaling pathways. Molecular docking identified potential protein targets e.g.  CYP1A2  NT5E  FOLR1 that may mediate observed effects. This study demonstrates the importance of appropriate exposure methods for semi volatile compounds  reveals structure dependent toxicity among alkyl substituted naphthalenes  and provides a foundation for further mechanistic studies and improved risk assessment of alkyl substituted PAHs. Overall design: Zebrafish were exposed to 25 chemicals  naphthalene and 24 alkyl sustituted naphthalenes  using a vial exposure method. Embryos were exposed to 20 uM of each chemical in groups of 8 in 2 mL of media in 2 mL glass vials from xxx hpf ttwo xxx hpf. RNA was collected at at 48 hpf. Exposures  collections and extractions were conducted over two days. Targeted transcriptomics was preformed using the TempO seq platform.", null, null, null, "1 ethylnaphthalene  C", "GSM8969924", null, "source name:whole animal|tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1|geo loc name:missing|collection date:missing", "1 ethylnaphthalene  C", "Demultiplexing and alignment was performed using the manufacture provided processing platform TempO SeqR BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Counts of genes with multiple probes were summed as instructed by the manufacturer resulting in 3112 unique genes Read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Principal component analysis PCA of normalized read counts was used to determine that a batch effect was present based on exposure and RNA extraction day nested by experimental design. At this time one control sample  control day1  B was identified as an outlier and removed from all future analysis. The batch effect was removed from unnormalized data using the ComBat seq function from the SVA R package Normalzied read counts were normalized by upper quartile normalization using the calcNormFactors function from the Bioconductor package in EdgeR using the 75th percentile of reads per samples as the scaling factor. Assembly: GRCz11 Supplementary files format and content: .csv file containing raw count data Supplementary files format and content: .csv file containing batch corrected and normalized data", "whole animal", "When the embryos were 6hpf they were bleached in groups of 1000 or less. Embryos were placed in a strainer for transfer between solutions in 500 mL containers. First embryos were placed in system fish water for 1 minute  then transferred to a 0.5% sodium chloride solution for 5 minutes  then transferred to a    % sodium hypochlorite solution for 1 minute and rinsed in EM for 5 minutes and transferred to a clean glass petri dish. Embryos were then loaded into 2mL glass exposure vials in groups of 8 using a glass pasture pipette. Excess EM was removed and 2mL of fresh EM was immediately added. Chemical stocks were diluted to 200 times the desired exposure concentration of 20 \u00b5M using DMSO. 10 \u00b5L of diluted chemical stock was added to each vial  vials were immediately capped and inverted 3 times to ensure mixing  no precipitate was observed for any chemicals at the concentrations tested. Vials were loaded into a custom 3D printed vial holder for slow end over end rotation in a dark 28\u00b0C incubator overnight. At 24 hours embryos were removed from exposure vials and plated into 96 well plates loaded with 100 \u00b5L clean EM. At 48 hpf embryos were collected for RNA extraction  embryos from two exposure vials were kept as holdback for morphological assessment. From the remaining wells  6 phenotypically normal embryos were collected and pooled in their vial groups into 1.5 mL Eppendorf tubes  placed on ice 30 sec  excess media removed  placed back on ice 30 sec  and 200 \u00b5L RNA shield added. Five pooled samples were collected from each treatment.", "1mL of 1\u00b5m beads and 500\u00b5L of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100\u00b5L of lysis buffer was added to tube containing pooled embryo samples and 200\u00b5L of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 \u00b5L of the supernatant was then transferred to two plates containing 275\u00b5L of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50\u00b5Lof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/\u00b5L. 1 \u00b5L per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer\u2019s instructions. PCR amplification products were pooled into a single sequencing library 5 \u00b5L each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.", null, "tissue:whole animal|genotype:5D WT|treatment:1 ethylnaphthalene|batch:1", "GSM8969924", "GSM8969924: 1 ethylnaphthalene  C; Danio rerio; OTHER", "GSM8969924 r1", "GSM8969924", "1", "1mL of 1\u00b5m beads and 500\u00b5L of lysis buffer were loaded into a round bottom 2mL 96 well plate. 100\u00b5L of lysis buffer was added to tube containing pooled embryo samples and 200\u00b5L of RNA shield  the full contents of the tube were then transferred to the homogenization plate. The plate was homogenized for 1.5 minutes and centrifuged for 5 minutes at 3000 rcf. 275 \u00b5L of the supernatant was then transferred to two plates containing 275\u00b5L of ethanol. RNA extraction was performed using the KingFisher Apex system. Briefly this process uses Mag beads to collect RNA from the sample  post series of washes the RNA is eluted in 50\u00b5Lof ultra pure water. RNA quality was assessed using the Agilent 4150 TapeStation System. All samples used had RIN scores >9.8. RNA concentration was determined using ThermoFisher Quant iT RNA Reagent and assay kit and ThermoFisher Varioskan ALF multimode plate reader in triplicate  samples with high variability between reads were repeated in triplicate. The average of the three quantification reads was used for RNA concentration normalization. Each treatment had 4 5 viable RNA extraction replicates  for treatments with 5 viable samples the 4 with the highest concentration and RIN score were used 3 control samples were used from each day group. All 94 samples were normalized to the lowest concentration sample 14.9ng/\u00b5L. 1 \u00b5L per sample of normalized RNA was used as input for library preparation following manufactures protocol for the 96 sample TempO Seq S1500 + Zebrafish Surrogate Assay panel BioSpyder Technologies  Inc.  Carlsbad  CA  United States. Briefly  samples were added to the assay plate with an equal volume of 2x enhanced lysis buffer and processed through annealing with detector oligos  digestion  and ligation. Proper amplification was verified using real time PCR green fluorescence. Library purification was performed using the Macherey Nagel NucleoSpin Gel and PCR Cleanup Kit following adjustments detailed in the manufacturer's instructions. PCR amplification products were pooled into a single sequencing library 5 \u00b5L each into a 1.5mL Eppendorf tube the pooled purified libraries were sequenced using one lane of the NovaseqX 10 B flow cell BioSpyder Technologies  Inc.  Carlsbad  CA  United States.", null, "OTHER", "TRANSCRIPTOMIC", "other", "SINGLE", "ILLUMINA", "Illumina NovaSeq X", null, "SRP583274", null, null, "8_C.fastq.gz", "fastq", 659636703.0, 12934053.0, "GSM8969924 r1", "0:51", "A:134295377;C:173327337;G:153006654;T:198857895;N:149440", 51, null, null, null, 134295377, 173327337, 153006654, 198857895, 149440, "SRX28685517", "SRS24952420", "SRA2124446", "SARL, Environmental and Molecular Toxicology, Oregon State University", "SARL, Environmental and Molecular Toxicology, Oregon State University", null, null, null, null, null, null, null, null, null, null, null, "B", null, "usable mapping rate", "illumina", "novaseq_era", "unknown", "other", "unknown", "bulk", "unknown", "unknown", null, "United States", "2025-05-06", "Multi-stage", "Embryo", "Whole Organism", "All anatomical structures"]], "columns": ["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"], "primary_keys": ["rowid"], "primary_key_values": ["36065"], "units": {}, "query_ms": 8.252268999058288}