Next Generation Sequencing to understand the pathogenic role of synovial fibroblasts in experimental models of arthritis

ABSTRACT: The goal of this study is to define the pathophysiology of synovial fibroblasts in chroninc joint inflammation using NGS-derived synovial transcriptome profiling (RNA-seq) Methods: Synovial fibroblasts were isolated by FACs sorting [CD45-CD31-Popdoplanin+] from the synovium of healthy mice and mice undergoing experimental Collagen-Induced Arthritis (12 weeks old)(CIA). Methods: Whole Transcriptome Profiling of healthy and CIA synovial fibroblasts were generated by deep sequencing, in triplicate, using Illumina NextSeq™ 500 platform. Libraries were prepared using polyA selection (TruSeq stranded mRNA kit). Methods: The sequence reads that passed quality filters were aligned to mouse reference genome (GRCM38) using Hisat2 version 2.1.0. we mapped about 30 million sequence reads per sample (75 bp, paired-end) Methods: Featurecounts version 1.4.6 was used to quantify reads counts. Data quality control, non-expressed gene filtering, median ratio normalization (MRN) implemented in DESeq2 package and identification of differentially expressed (DE) genes were done using the R bioconductor project DEbrowser. Results: Results: We detected several differentially expressed (DE) genes in CIA synovial fibroblasts compared to naïve cells. These included 298 up-regulated genes and 88 down-regulated (>2 fold, adjp <0.05). DE genes reflected the reflecting the hyperplasia and activation observed during chronic joint disease. Pathways associated to DE genes were cell cycle and cell division and inflammatory response. Conclusions: our study shows pathophysiological changes associated to inflammatory synovial fibroblasts in a model of rheumatoid arthritis. Cells were directly isolated from fresh tissue, providing a valuable tool to study stromal inflammation.

ORGANISM(S): Mus musculus

PROVIDER: GSE162306 | GEO | 2020/11/30

REPOSITORIES: GEO