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009 Single cell RNA sequencing reveals specific subsets in Systemic Sclerosis fibroblast cultures

      Systemic sclerosis (SSc) is a chronic autoimmune disease causing vasculopathy and fibrosis in the skin and vital organs. Following an inflammatory reaction, fibroblasts in the connective tissues become activated myofibroblasts and secrete excessive amounts of ECM related proteins, such as collagen, creating fibrosis. Many studies have been carried out in fibroblasts in culture, however, the information obtained remains limited due to high potential heterogeneity of these fibroblasts. We therefore used single cell RNA sequencing to better understand fibroblast heterogeneity in SSc to search for potential new therapeutic targets and new biomarkers. We conducted single cell RNA-sequencing (scRNA-seq) analysis of human dermal fibroblasts from 4 healthy and 4 SSc donors. We identified several up- and downregulated genes in defined cell populations; we also confirmed significant upregulation of tetraspanin CD9 in all SSc fibroblast subsets when compared to controls. CD9 upregulation at the cell surface of SSc fibroblasts was confirmed by flow cytometry. Additionally, we identified strong upregulation of Four and a half LIM domains 1 (FHL1) in a small cluster of SSc fibroblasts at the RNA and protein level. These cells did not express α-SMA and thereby constitute a small subpopulation of activated fibroblasts. CD9 was found in exosomes of SSc fibroblasts and FHL1 has been implicated in myofibroblast differentiation; there is not much known about their role in fibrosis. We are therefore focusing on the direct impact of both CD9 and FHL1 using specific deletion experiments.