Title: Defining pathogenic fibroblast lineages in scleroderma using mouse models
Funder: Versus Arthritis

Abstract: Scleroderma (Systemic Scleosis) represents a spectrum of immune-mediated rheumatic diseases with differing clinical course and diverse manifestations. Patients are stratified into disease endotypes by clinical and biochemical features including extent of skin disease, internal organ involvement, biomarkers (mainly serology) and more recently molecular profiling. Despite advances in the treatment for scleroderma associated pulmonary arterial hypertension and immune-suppressive therapies for autoimmune and inflammatory manifestations, there is no treatment to tackle the main pathogenic process in scleroderma resulting from the tissue remodelling, scarring and fibrosis. Indeed there is a fundamental deficiency in our understanding of pathobiology of fibrosis in scleroderma and this major challenge has to be confronted before innovative treatments and effective cures will be developed. Fibroblasts and myofibroblasts are the predominant cell types responsible for maintaining connective tissues, yet little is known regarding the origins or heterogeneity of these cell types in scleroderma, nor their functional diversity in causing tissue scarring and fibrosis. We propose to identify, isolate and study fibroblast populations responsible for dermal fibrosis using a mouse model of scleroderma. This mouse model (TßRII?K;DNR) recapitulates many of the clinical, histological and biochemical features seen in scleroderma and exhibits enhanced expression and deposition of collagen type I in lesional tissues including the skin. By employing an inducible Collagen type I gene (Col1a2)-driven Cre-recombinase with a double fluorescent (Red/Green) targeting approach we will isolate and characterise fibroblast population(s) using fluorescence activated cell sorting based on the gene expression of collagen type I. We will use RNASeq to identify fibroblast specific gene expression and culture fibroblast populations in order to examine their secretomes and functional activities including migration, proliferation, contraction and extracellular matrix production. Flow cytometry and immunofluorescence will also be used to further characterise fibroblasts (and confirm pathway data from RNASeq), explore fibroblast transition into smooth muscle actin positive (aSMA+ve) myofibroblasts and assess the contribution of previously reported dermal fibroblast lineages in fibrosis with cell surface phenotypic markers (CD26, Lrig1, ITGA8). Once lineage markers have been established we will investigate relevance to organ-specific fibrosis by examining their potential role in pulmonary fibrosis within the TßRII?K;DNR mouse. Finally, we will investigate whether analogous lineages and pathways are present in scleroderma tissues using our established biobank of samples from across the disease spectrum. These studies will highlight the lineage and functional variation of fibroblasts in scleroderma moue models, provide new tools, key candidates for therapeutics and biomarkers and a clear rationale to further explore and refine our understanding of fibroblast origins and lineages in fibrosis in scleroderma.


Research activity codes: 2.1 BIOLOGICAL AND ENDOGENOUS FACTORS
Health categories: INFLAMMATORY AND IMMUNE SYSTEM