CCBIO seminar: Robert Lafyatis
Understanding cellular heterogeneity and fibrosis in systemic sclerosis skin using single cell RNAseq: SFRP2/DPP4 and FMO1/LSP1 define two major fibroblast populations in normal skin
Division of Rheumatology and Clinical Immunology, Department of Medicine, University of Pittsburgh Medical Center, PA, USA
Fibroblasts secrete extracellular matrix, mediating reparative and fibrotic processes. Fibroblasts play key roles in healing wounds but also as the mediators of fibrosis, responsible for end-stage organ damage in a wide range of diseases, including systemic sclerosis. In addition, they regulate inflammation, by anchoring leukocytes and regulating immune cell functions. In lymphoid tissues they contribute to secondary lymphoid organ structure and in non-lymphoid tissues to the development of tertiary lymphoid structures. In carcinogenesis fibroblasts have an emerging role, as cancer associated fibroblasts support and regulate tumor cell growth. Thus, fully understanding the complexity of normal fibroblasts is key to understanding their roles in a wide variety of pathological conditions.
Advances in scRNA-seq technology, allowing thousands of single cells to be analyzed in a single experiment, provide much more robust insight into this heterogeneity. Using scRNA-seq of digested human skin cells and known marker genes, we identified keratinocyte subtypes, endothelial cells, pericytes, arrector pili/smooth muscle cells, and fibroblasts. T-distributed stochastic neighbor embedding of scRNA-seq data from six biopsies showed that fibroblasts, expressing type I collagen and PDGFRA, form a discrete cell population. Within the grouping of fibroblasts, clustering revealed two major populations, defined by clusters of distinct genes, including SFRP2 and FMO1, genes expressed exclusively in the skin by these two major fibroblast populations. Clustering identified fibroblast subpopulations within each of the SFRP2 and FMO1 populations, as well as four minor fibroblast populations, each expressing discrete transcriptomes and markers genes. Immunofluorescent staining confirmed that SFRP2 and FMO1 expression define cell types of dramatically different morphology. SFRP2+ fibroblasts are small, elongated, and distributed between collagen bundles. FMO1+ fibroblasts are larger, and distributed in both interstitial and perivascular locations. High expression of COL1A1 by SFRP2+ and CXCL12 by FMO1+ fibroblasts suggests roles for these cell types in matrix deposition and inflammation, respectively. We have also begun to examine the alterations in dermal fibroblasts occurring in the fibrotic skin of patients with systemic sclerosis. We anticipate that these single cell analyses will help us to understand the origin and stimuli leading to the development of myofibroblasts and other profibrotic cells in the skin of these patients
Chairperson: Donald Gullberg <email@example.com>, CCBIO