{"database": "metadata", "table": "run_metadata", "rows": [[30627, "SRR27907721", "SRX23567475", "SRS20412966", "SRP488787", "PRJNA1074408", "Ctla 4 deficiency induces an inflammatory bowel disease like phenotype in a zebrafish model [scRNA seq]", "GSE255303", "Transcriptome Analysis", "Inflammatory bowel disease IBD is a chronic and relapsing immune mediated disorder characterized by intestinal inflammation and epithelial injury. The underlying causes of IBD are not fully understood  but genetic factors have implicated in genome wide association studies  including CTLA 4  an essential negative regulator of T cell activation. However  establishing a direct link between CTLA 4 and IBD has been challenging due to the early lethality of CTLA 4 knockout mice. In this study  we identified zebrafish Ctla 4 ortholog and investigated its role in maintaining intestinal immune homeostasis by generating a Ctla 4 deficient ctla 4 /  zebrafish line. These mutant zebrafish exhibit reduced weight  along with impaired epithelial barrier integrity and lymphocytic infiltration in their intestines. Transcriptomics analysis revealed upregulation of inflammation related genes  disturbing immune system homeostasis. Moreover  single cell RNA sequencing analysis indicated increased Th2 cells and interleukin 13 expression  along with decreased innate lymphoid cells and upregulated proinflammatory cytokines. Additionally  Ctla 4 deficient zebrafish exhibited reduced diversity and an altered composition of the intestinal microbiota. All these phenotypes closely resemble those found in mammalian IBD. Lastly  supplementation with Ctla 4 Ig successfully alleviated intestinal inflammation in these mutants. Altogether  these findings offer substantial evidence linking CTLA 4 to IBD and establish a new model for investigating pathogenesis and potential treatments. Overall design: intestine leukocyte WT and KO Ctla 4 deficiency  of the AB zebrafish were isolated by Ficoll Hypaque 1.080 g/mL density gradient centrifugation and analyzed using scRNAseq.", null, "pubmed:40392591", null, "WT  scRNAseq", "GSM8068832", null, "source name:intestine|tissue:intestine|cell type:leukocyte|genotype:Wildtype|geo loc name:missing|collection date:missing", "WT  scRNAseq", "The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v2.1.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: GRCz11 Supplementary files format and content: Tab separated values files and matrix files", "intestine", null, "Leukocytes were collected from the intestine samples through Ficoll Hypaque 1.080 g/mL density gradient centrifugation. Briefly the intestine was carefully excised and transferred through a 40 \u03bcm stainless nylon mesh Greiner Bio OneGmbH  Germany. The cell suspension was suspended in Leibovitz's L 15 Medium L 15  Gibco supplemented with penicillin 100 U/ml  Sigma Aldrich  streptomycin 100 \u03bcg/ml  Sigma Aldrich and heparin sodium 10 U/ml  Sigma Aldrich. The cell suspension was slowly added into a Ficoll Hypaque 1.080 g/mL density gradient centrifugation  centrifuged at 1 200 g for 25 min  and the cell layer of the interface was carefully aspirated and then washed with ice cold PBS at 400 g for 10 min. Cell quantity and viability were determined using 0.4% trypan blue Sigma  St. Louis  MO  USA  which showed that more than 95% were living cells. The cells of each group were counted using a cell counting plate. Library was performed according to the manufacter\u2019s instructions single cell 3\u2019 v2 protocol  10x Genomics. Briefly  GCs were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead in emulsion GEM. The poly A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA  which contains an Ilumina R1 primer sequence  Unique Molecular Identifier UMI and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads  amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence  paired end constructs with P5 and P7 sequences and a sample index were added.", null, "tissue:intestine|cell type:leukocyte|genotype:Wildtype", "GSM8068832", "GSM8068832: WT  scRNAseq; Danio rerio; RNA Seq", "GSM8068832 r1", "GSM8068832", "1", "Leukocytes were collected from the intestine samples through Ficoll Hypaque 1.080 g/mL density gradient centrifugation. Briefly the intestine was carefully excised and transferred through a 40 \u03bcm stainless nylon mesh Greiner Bio OneGmbH  Germany. The cell suspension was suspended in Leibovitz's L 15 Medium L 15  Gibco supplemented with penicillin 100 U/ml  Sigma Aldrich  streptomycin 100 \u03bcg/ml  Sigma Aldrich and heparin sodium 10 U/ml  Sigma Aldrich. The cell suspension was slowly added into a Ficoll Hypaque 1.080 g/mL density gradient centrifugation  centrifuged at 1 200 g for 25 min  and the cell layer of the interface was carefully aspirated and then washed with ice cold PBS at 400 g for 10 min. Cell quantity and viability were determined using 0.4% trypan blue Sigma  St. Louis  MO  USA  which showed that more than 95% were living cells. The cells of each group were counted using a cell counting plate. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics. Briefly  GCs were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead in emulsion GEM. The poly A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA  which contains an Ilumina R1 primer sequence  Unique Molecular Identifier UMI and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads  amplified by PCR and the apropiated sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence  paired end constructs with P5 and P7 sequences and a sample index were added.", null, "RNA-Seq", "TRANSCRIPTOMIC SINGLE CELL", "cDNA", "PAIRED", "ILLUMINA", "NextSeq 500", null, "SRP488787", null, "loader:fastq load.py", "WT_S1_L001_R1_001.fastq.gz WT_S1_L001_R2_001.fastq.gz", "fastq fastq", 36724066995.0, 308605605.0, "GSM8068832 r1", "0:28 1:91", "A:10425178276;C:8399183780;G:8411777361;T:9487202777;N:724801", 28, 91, null, null, 10425178276, 8399183780, 8411777361, 9487202777, 724801, "SRX23567475", "SRS20412966", "SRA1799569", "zhejiang university", "zhejiang university", 2, 0.01141, 0.94944, 0.0039, 0.1092, 0.99324, 0.841, 0.43558, 0.73314, 28, 91, "T", "B", "sc-like readlen", "illumina", "nextseq", "unknown", "poly_a", "unknown", "sc", "single_cell_droplet", "10x", null, "China", "2024-02-07", "Undetermined", "Undetermined", "Gut", "Digestive System"]], "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": ["30627"], "units": {}, "query_ms": 11.700832998030819}