Bcl-2 but not clusterin/apolipoprotein J protected human diploid fibroblasts and immortalized keratinocytes from ceramide-induced apoptosis: Role of p53 in the ceramide response

The role of clusterin/apolipoprotein J (Clu/ApoJ) and Bcl-2 on C2-ceramide-induced apoptosis of embryonic human diploid fibroblasts, MRC-5 and immortalized adult skin keratinocytes, HaCaT was investigated. C2-ceramide-induced apoptosis of HaCaT in a time- and dose-dependent manner, while in MRC-5 only at higher concentrations. There was a dose-dependent accumulation of Clu/ApoJ and downregulation of Bcl-2 which correlated with C2-ceramide-induced apoptosis of MRC-5. While overexpression of Bcl-2 suppressed C2-ceramide-mediated apoptosis in both cell types, Clu/ApoJ failed to do so, accessed by morphological changes, DNA fragmentation and PARP cleavage. There was no change in the expression of endogenous p53 or p21Waf1/Cip1 upon C2-ceramide treatment of MRC-5. However, mutant p53143ala increased the sensitivity of MRC-5 to C2-ceramide-induced apoptosis by markedly downregulating Bcl-2, pointing to a role for p53. These results suggested that whereas downregulation of Bcl-2 may be a crucial factor involved in C2-ceramide-induced apoptosis, accumulation of Clu/ApoJ may be a signal of stress response. Moreover, the ceramide-activated apoptotic pathway may be regulated by p53.