In vivo evidence that capase-3 is required for Fas-mediate apoptosis of hepatocytes

Caspase-3 is essential for Fas-mediated apoptosis in vitro. We investigated the role of caspase-3 in Fas-mediated cell death in vivo by injecting casp ase-3-deficient mice with agonistic anti-Fas Ab, Wild-type controls died ra pidly of fulminant hepatitis, whereas the survival of caspase-3(-/-) mice w as increased due to a delay in hepatocyte cell death. Bcl-2 expression in t he liver was dramatically decreased in wild-type mice following anti-Fas in jection, but was unchanged in caspase-3(-/-) mice. Hepatocytes from anti-Fa s-injected wild-type, but not caspase-3(-/-), mice released cytochrome c in to the cytoplasm, Western blotting confirmed the lack of caspase-3-mediated cleavage of Bcl-2. Presumably the presence of intact Bcl-2 in caspase-3(-/ -) hepatocytes prevents the release of cytochrome c from the mitochondria, a required step for the mitochondrial death pathway, We also show by Wester n blot that Bcl-x(L), caspase-9, caspase-8, and Bid are processed by caspas e-3 in injected wild-type mice but that this processing does not occur in c aspase-3(-/-) mice. This study thus provides novel in vivo evidence that ca spase-3, Conventionally known for its downstream effector function in apopt osis, also modifies Bcl-2 and other upstream proteins involved in the regul ation of Fas-mediated apoptosis.