Transplantation arteriosclerosis (TA) has continued to limit survival in cardiac allograft recipients. Endothelial cells (ECs) can be primary stimulators and targets of immunologic injury, of which TA may be a chronic form. The long-term objectives of this work are to further define novel molecular interactions between human lymphocytes and allogeneic ECs, and ultimately apply this information to the understanding of immune-mediated vascular injury. T cells interact minimally with resting endothelium. Natural killer (NK)cells require little antigen-priming and do interact with resting ECs in vitro. The underlying hypothesis is three-fold; (1) ECs are antigen-presenting cells (APCs) but their ability to present class I HLA-associated peptides is intrinsically deficient, (2) this deficiency provides the """"""""substrate"""""""" for allorecognition by and sensitivity to nK lymphocytes, and (30 NK cell allorecognition of the endothelium, NK-EC adhesion, mutual cellular activation and/or EC lysis are all highly interdependent phenomena which may constitute the cellular triggers for alloimmune-mediated vascular injury. Based upon our prior observations of alloantigen-specific EC lysis by NK lymphocytes and the important role for the leukocyte integrin LFA-1 (alphaL beta2) in this phenomenon, specific proposals now include to (1) investigate EC contact-dependent NK activation, utilizing murine fibroblasts stably transfected with EC counter-receptor genes, alphaL beta2 cytoplasmic deletion mutants expressed in COS cells, and RT-PCR to evaluate contact-dependent NK gene activation; (2) dissect the molecular mechanisms involved in NK cell contact-dependent EC Ca2+ oscillations at the single cell level, utilizing an interactive laser cytometer; (3) evaluate the nature of class I HLA-associated peptides on the EC membrane, address how IFNgamma affects MHC-encoded gene products in ECs, and attempt to relate these findings to EC NK sensitivity; and (40 further define NK cell allospecific recognition of the endothelium by characterizing NK receptor and target molecules, utilizing a recently generated panel of mAbs and HLA-serotyping data. Information generated in these studies will not only enhance our understanding of NK and leukocyte integrin biology, but provide insight into their respective roles in vascular pathology as well.
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