RNA-seq approaches to understand the biological effects of sialic acid on murine synovial fibroblasts

The aim of this study was to explore the role sialic acid in synovial fibroblasts using NGS-derived transcriptome analysis (RNA-seq). Methods: Synovial fibroblasts were extracted from the synovium of healthy mice and expanded ex vivo. RNA was extracted from sialidase buffer (NT) and Clostridium perfringens Neuraminidase (CP) treated synovial fibroblast. Methods: Whole Transcriptome Profiling was 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: we detected differentially expressed genes in CP treated synovial fibroblast compared to NT synovial fibroblast, which pass the threshold of >2 fold, adjp <0.01. DE genes reflect increased cellular inflammatory response after CP treatment Conclusions: These findings underline the ability of sialic acid to modulate homeostatic and inflammatory responses in non-immune synovial fibroblasts, suggesting that sialylation plays a key role in perpetuating local inflammation in the arthritic joint. Our results show the transcriptome profile of NT and CP treated synovial fibroblast expanded ex vivo