Inhibition of inflammatory endothelial responses by a pathway involving caspase activation and p65 cleavage

Suppression of NF kappaB activation has been involved in the elimination of survival programs during endothelial cell (EC) apoptosis. We used alpha -t ocopheryl succinate (alpha -TOS) to trigger apoptosome formation and the su bsequent activation of executioner caspases. The level of bcl-2 was reduced by alpha -TOS, and its downregulation potentiated and its overexpression s uppressed pro-apoptotic effects of alpha -TOS, indicating a mitochondrial r ole in alpha -TOS-induced apoptosis in Ec. alpha -TOS treatment was associa ted with induction of TUNEL-positive apoptosis in EC with a high but not wi th a low proliferation index. The use of the pan-caspase inhibitor z-VAD.fm k suggested the involvement of caspases in cleavage of p65, and in inhibiti on of nuclear translocation of p65 and NF kappaB-dependent transactivation of a gene construct encoding the green fluorescence protein elicited by TNF alpha in contact-arrested EC. The suppression by alpha -TOS of inflammator y EC responses induced by TNF alpha such as VCAM-1 mRNA and surface protein expression and shear-resistant arrest of monocytic cells were also reverse d by z-VAD.fmk. NF kappaB-dependent transactivation was preserved in alpha -TOS-treated EC stably transfected with a caspase-noncleavable p65 mutant b ut not with its truncated form, thus establishing a direct link between alp ha -TOS-induced effects and p65 cleavage. Our data infer a pathway by which caspase activation in EC inhibits NF kappaB-dependent inflammatory activat ion and monocyte recruitment, and provide evidence for a relationship betwe en proapoptotic and anti-inflammatory pathways.