Adenoviral-mediated overexpression of I kappa B alpha in endothelial cells inhibits natural killer cell-mediated endothelial cell activation

Activated natural killer (NK) cells have been found in rejecting discordant xenografts and may contribute to endothelial cell (EC) activation and damage. The transcription of genes associated with EG activation, such as E-selectin and interleukin (IL)-8, is regulated by the transcription factor NF-kappa B. In resting EC, NF-kappa B is complexed within the cytoplasm to I kappa B alpha, and EG activation leads to dissociation of the I kappa B alpha-NF-kappa B complex and nuclear translocation of NF-kappa B. We investigated whether overexpression of I kappa B alpha in EC, using adenoviral gene transfer, could block NF-kappa B translocation, thereby inhibiting NK cell-mediated EG activation. Co-culture of human NK cells with porcine EG resulted in a threefold increase in E-selectin expression after 4 hr and secretion of greater than 650 pg/ml porcine IL-8 over 24 hr, Overexpression of I kappa B alpha inhibited the NK cell-mediated induction of E-selectin expression and IL-8 secretion, whereas overexpression of beta-galactosidase did not. The inhibition of EC activation was not due to variation in NK-EC adhesion, as the level of adhesion was similar between adenovirally infected and noninfected EG over 4 hr. The level of NK cell-mediated EG cytotoxicity was not significantly different after 4 hr of co-culture, but after 24 hr, cytotoxicity was increased in virally infected cells. Cytotoxicity was more marked in cells overexpressing I kappa B alpha than cells overexpressing beta-galactosidase. SLA class I and the induction of SLA class II antigen in response to interferon-gamma treatment were reduced in cells infected with adeno-I kappa B alpha and empty adenovirus, demonstrating that viral infection alone can influence EC biology. Overexpression of I kappa B alpha using adenovirus provides a novel approach to inhibiting NK cell-mediated EG activation, but additional strategies will be required to inhibit NK cell-mediated cytotoxicity.