Inhibition of UVA-induced c-Jun N-terminal kinase activity results in caspase-dependent apoptosis in human keratinocytes
Abstract
Inhibition of c-Jun N-terminal kinase (JNK) with the pharmacologic inhibitor SP600125 in UVA-irradiated HaCaT cells and human primary keratinocytes resulted in dramatic phenotypic changes indicative of cell death. These phenotypic changes correlated with caspase 8, 9 and 3 activations as well as cleavage of the caspase substrate polyADP-ribose polymerase (PARP). Morphologic analysis and analysis of sub-G0 DNA content confirmed apoptotic cell death in these keratinocytes after combination treatment. Addition of the general caspase inhibitor zVAD-fmk to combination-treated HaCaT cells was able to completely block caspase activation, PARP cleavage, the increase in sub-G0 DNA content and the classic morphologic features of apoptosis, indicating that this combination treatment resulted in caspase-dependent apoptotic cell death. zVAD-fmk treatment of primary keratinocytes was able to completely inhibit caspase activation and PARP cleavage, reduce morphologic apoptosis at lower concentrations of SP600125 and decrease the sub-G(0) DNA content detected after UVA + SP600125 treatment. However, cell death and a significant amount of debris was still detected after caspase inhibitor treatment, particularly with 125 nM SP600125. At subconfl...Continue Reading
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