The underlying hypothesis for this project is that the endothelial lining of blood vessels can regulated both quantitative and qualitative aspects of helper T-cell lymphocyte responses to antigens. These regulatory functions could be due to selective co-stimulation and/or recruitment of subsets of T-cells with different effector functions. The proposed studies will focus on endothelial stimulation of T-cell cytokines IFN-gamma and IL-4, as well as the recruitment of T-cells that produce these cytokines, namely Th1 versus Th2 cells. The experimental approach will involve in vitro and in vivo studies, and both mouse and human cells. The unique advantages of TCR transgenic mice and adoptive transfer experiments will be emphasized.
The specific aims are: 1: Characterize endothelial induced signals that enhance helper T-cell effector cytokine production.
This aim will test the hypothesis that endothelial cells can prime T-cells for rapid effector cytokine responses to extravascular antigens. The studies will entail a molecular analysis of endothelial co-stimulation of T-cell IFN-gamma or IL-4 production. Cellular immunological assays will be followed up with analysis of signal transduction and transcriptional regulation, focusing on Jak-Stat pathways. 2: Elucidate the molecular mechanisms underlying differential adhesive interactions between helper T-cell subsets and endothelium in vitro. The focus will be on expression or modification of molecules that promote T- cell migration with inflammatory sites. Experiments will analyze the influence of Th1 or Th2 differentiation factors on expression of functional selectin ligands, high affinity integrins, and chemokine receptors. 3: Define how recruitment patterns of T-cell subsets are regulated in vivo. Adoptive transfer experiments in mice will be performed to study recruitment of circulating Th1 or Th2 cells into sites of immune-mediated inflammation. The accumulation of T-cells from TCR transgenic mice into such sites will be followed. Antibody blockade as well as mutant recipient mice will be utilized to elucidate the importance of endothelial adhesion molecules in subset-specific recruitment. Additional experiments will examine human Th1 and Th2 recruitment into skin grafts on SCID mice. This project will provide detailed answers to the question of how endothelial cells promote different types of helper T-cell responses. The importance of these answers lies in the fact that the effector cytokines produced by Th1 and Th2 lead, respectively, to profoundly different types of immune responses. All three aims of this project address the signaling and communication pathways by which endothelium and leukocytes interact to influence inflammatory responses. As such, they are relevant to the overall theme of the program. The information to be gained will help define new targets for pharmacological treatment of immune mediated inflammatory processes such as autoimmune diseases and transplant rejection.
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