Cytokines produced by activated by T lymphocytes play a critical role in the pathophysiology of LPS-induced lung injury. Recently, it has become clear that stimulation of T cells via the T cell antigen receptor (TCR), under some circumstances, leads to cytokine production and other effector functions while under other circumstances leads to program cell death (also known as apoptosis). Apoptosis of T lymphocytes plays a key role in regulating T cell population size following initial clonal expansion to combat an inciting antigen. Apoptosis occurs also as a means to ensure immune privilege in various tissues. Interestingly, it appears that the mechanism by which T cells undergo apoptosis, either following initial immune expansion or when infiltrating immune privileged sites, involves an interaction between CD95 on the surface of T cells and CD95 ligand expressed on immune-privileged tissues or on T cells themselves. While numerous studies have been published investigating the mechanism by which CD95 signal transduction regulates T cell apoptosis, little is known about how CD95 ligand expression is regulated. Recent work from several investigators suggests that CD95 and CD95 ligand are not expressed exclusively on hematopoietic cells. Several studies indicated that both molecules are expressed in the pulmonary system. While the expression of these molecules appears critical for induction of tolerance in the lung to repeat administration of antigens, it is not yet clear whether CD95 and CD95 ligand expression play a role in cell death due to lung injury. Additionally, it is not yet known what signals are required to induce expression of CD95 ligand in the lung. The overall goal of this proposal, therefore, is to began an analysis of transcriptional regulation of CD95 ligand in T cells as well as non-lymphoid tissue. To achieve this goal, three specific aims are presented. The first is to investigate signaling events which are causally related to expression and regulation of CD95 ligand at the level of gene transcription in T cells. Experiments for this aim involve those addressing what members of the MAP kinase family of protein kinases are important for TCR induced expression of CD95 ligand and to ask whether other molecules known to impact TCR mediated activation events also are important for regulation of this protein. Additional experiments are presented examining the transcription factors which are induced following TCR engagement. The second specific aim describes experiments addressing what transcription factors may be important for the regulation of CD95 ligand in a TM4 Sertoli cell line, a model for an immune-privileged tissue.
The final aim for this proposal is the generation of a mouse transgenic for a reporter construct which will include elements for the CD95 ligand promoter. This mouse will be used to investigate regulation of CD95 ligand in vivo with a focus on its expression in """"""""classical"""""""" immune-privileged sites, and in the lung both in the basal state and following LPS administration. Collectively, it is hoped that these studies will provide insight into the regulation of expression of CD95 ligand and may help us learn how to modulate expression of this important molecule for potential therapeutic purposes.
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