CD8+ T cells drive myofibroblast activation and contraction via JAK/STAT3 and TGFβ signaling

Abstract

BackgroundFibrosis is a major cause of morbidity and mortality in rheumatic connective tissue diseases. In these conditions, fibrotic tissues are characterized by the infiltration of CD8+ T cells and pro-fibrotic myofibroblasts. However, the role of CD8+ T cells in driving myofibroblast activity remains unexplored. Our aim was to investigate this interaction between CD8+ T cells and skin myofibroblasts and to elucidate its underlying mechanisms. MethodsPrimary skin-derived myofibroblasts were co-cultured with peripheral blood mononuclear cells (PBMCs) or sorted T cells in a 3D collagen type I hydrogel. To model autoimmunity, allogeneic mismatching was applied. T cell activation and cytokine expression were assessed using flow cytometry and Luminex. Myofibroblast activation was analysed through immunohistochemistry (IHC) and quantitative polymerase chain reaction (qPCR), while activation-induced contraction was measured macroscopically. Intracellular signalling pathway activation in myofibroblasts was evaluated using luciferase reporter cell lines. ResultsCo-culture of myofibroblasts with PBMCs strongly induced hydrogel contraction and expression of activation-related markers such as podoplanin, fibroblast activation protein, pSTAT3 and IL-6 in myofibroblasts. CD4+ T cells and CD8+ T cells were also activated by co-culture as identified by increased CD25 and CD69 expression and elevated IL-2 and IFN-{gamma} production. Upon co-culture with either sorted CD4+ or CD8+ T cells, CD8+ T cells more strongly induced myofibroblast contraction and activation than CD4+ T cells. This was not associated with cytotoxicity but with increased IL-6 production by CD8+ T cells compared to CD4+ T cells and STAT3/TGF{beta}-induced signaling in myofibroblasts. Use of either the JAK/STAT3-inhibitor tofacitinib or the TGF{beta} receptor inhibitor SB-505124 blocked the activated myofibroblast phenotype, and combined use of both inhibitors had an additive effect on myofibroblast activation and contraction. ConclusionsCD8+ T cells drive primary skin derived myofibroblast contraction and activation not via cytotoxicity-related program but via cytokine release. This sheds light on novel mechanisms of immune cell mediated tissue fibrosis. Furthermore, our results suggest that combining JAK/STAT inhibition with an anti-fibrotic agent that blocks matrix synthesis might be promising in mitigating immune cell mediated tissue fibrosis in connective tissue rheumatic disorders and has added value over blocking these pathways individually.