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Natural killer (NK) cell mediated cellular response plays an important role in xenograft rejection. NK mediated cytotoxicity can directly cause endothelial damage during xenograft rejection. This in part is due to the inability of xenogeneic MHC class I molecules to provide inhibitory signals to human NK cells. Recent studies have shown that two of the human non-classical HLA class I molecules, HLA-G and HLA-E, are able to regulate NK function by direct interaction with NK inhibitory receptors. Previous study in our laboratory has shown that HLA-G expression is able to protect porcine endothelial cells from freshly isolated human NK cell mediated lysis. In this study, we show that a second non-classical HLA class I molecule, HLA-E, expression on porcine endothelial cells also inhibits human NK cells. The full length genomic HLA-E DNA and full length human beta-2-microglobulin DNA was co-transfected into porcine aortic endothelial cells by the lipofection method. The expression of HLA-E was confirmed by both RT-PCR using HLA-E specific primer and FACS analysis using a human HLA class I specific monoclonal antibody PA2.6. The expression of HLA-E inhibited the lysis of porcine endothelial cells by both the freshly isolated NK cells (67.2 ±6.3 %) and the IL-2 activated NK cell lines (38.6 ± 7.4 %). This inhibition was blocked by the anti-HLA-E receptor CD94/NKG2 monoclonal antibodies, which indicated that the protection was mediated through HLA-E and CD94/NKG2 interaction. In conclusion, the expression of a non-classical HLA class I molecule, HLA-E, effectively blocks the porcine endothelial cells against human NK mediated lysis. This study indicates that the expression of human non-classical HLA class I molecules, i.e. HLA-G and HLA-E, can provide an effective strategy to protect xenografts from human NK mediated cytotoxicity.