PROJECT SUMMARY Fibrosis is a common final outcome of most human chronic inflammatory diseases and has been estimated to contribute to almost half of all deaths in the world1. It can result from autoimmune diseases such as scleroderma [systemic sclerosis (SSc)] and after stem cell transplant in graft-vs-host disease. SSc and the sclerodermoid subtype of graft-vs-host disease (sGvHD) most commonly affect the skin and the degree of skin involvement is associated with higher mortality and internal organ dysfunction2,3, suggesting common underlying mechanisms. T cells are fundamental regulators of fibrosis pathogenesis in both diseases, but the mechanisms by which T cells drive fibrosis remain unclear. Mismatch of minor histocompatibility (mHA) alleles is required for the induction of sGvHD4, but the importance of antigen-independent T cell functions is poorly understood. This proposal aims to understand how antigen non-related CD4+ T cells contribute to the pathogenesis of SSc in response to pro-inflammatory cytokines. In this project, we examine how innate-like functions of CD4+ T lymphocytes regulate skin fibrosis. CD4+ T cells respond to a specific antigen and differentiate into distinct subsets of helper T cells, including Th1, Th2, and Th175. However, differentiated CD4+ T cells can respond to IL-1 family cytokines to generate a diverse cytokine milieu, including IFNγ, IL-13 and IL-17A6. Such bystander activated CD4+ T cells have been shown to drive autoimmunity in diseases such as multiple sclerosis7. SSc and sGvHD skin are characterized by aberrant expression of cytokines spanning type 1, 2 and 17 immune responses8-10. We therefore propose that bystander activation of CD4+ T cells occurs in SSc and sGvHD and to investigate the mechanisms by which it is generated. To accomplish these goals, we will use sclerodermoid GvHD mouse models, which are the primary models of SSc skin fibrosis dependent on adaptive immune cells4,11. In our first aim, we will characterize bystander activation of CD4+ T cells in sGvHD mice by modulating the number of antigen unrelated CD4+ T cells and measuring their accumulation and cytokine expression profile in relation to fibrosis severity. This will allow us to establish to what degree bystander activated CD4+ T cells affects skin fibrosis in vivo. In the second aim, we will examine the cytokine signals that activate CD4+ T cells in this model. We will assess protein levels of inflammatory cytokines including IL-1 family members in the skin and blood and perform in vitro validation for their ability to drive expression of effector cytokines and extracellular matrix genes by fibroblasts. Together, these aims will provide mechanistic insight into how T cells drive the pathogenesis of SSc skin fibrosis, which may identify new targetable signaling pathways for development of treatments that impact patients’ lives.