Induction of apoptosis in human oral squamous carcinoma cell lines by protein phosphatase inhibitors

To determine whether protein phosphorylation and dephosphorylation can affe ct apoptosis in oral epithelial cells we examined the effects of protein ph osphatase inhibitors, okadaic acid (OA) and calyculin A (CA), on cultured h uman oral squamous carcinoma (SCC) cell line, SCC-25 cells. After reaching subconfluence these cells were exposed to varying concentrations of the pro tein phosphatase inhibitors, OA and CA. Both OA and CA induced cell death i n SCC-25 cells in st dose-dependent fashion as determined by phase-contrast microscopy and WST-1 cell viability assay. By using the Hoechst 33342 stai ning, marked nuclear condensation and fragmentation of chromatin was observ ed. DNA ladder formation also was detected in SCC-25 cells by treatment wit h OA and CA. The induced nuclear fragmentation and DNA ladder formation wer e dose-dependent with maximal effect at concentrations of 20 nM OA and 2 nM CA, respectively. OA also induced DNA ladder formation in other human oral SCC cell lines, SCCKN and SCCTF. To further determine if new gene transcri ption and protein synthesis are required for OA-induced apoptosis in SCC-25 cells, the cells were treated for 48 h with varying concentrations of cycl oheximide in the presence of 20 nM OA. Cycloheximide did not protect the ce lls against OA-induced cytotoxicity and DNA ladder formation. Based on the known selectivity of OA and CA, the present results indicate that the pathw ay of the apoptosis in the cultured oral SCC cells is in part regulated by protein phosphatase type 1 and type 2A. Our results also indicate that new protein synthesis is not involved in OA-induced apoptosis in SCC-25 cells. (C) 1999 Elsevier Science Ltd. All rights reserved.