Rusnak J M, Lazo J S
Department of Pharmacology, University of Pittsburgh School of Medicine, Pennsylvania 15261, USA.
Exp Cell Res. 1996 Apr 10;224(1):189-99. doi: 10.1006/excr.1996.0127.
Protein kinase C (PKC) has been implicated in propagating signals for apoptosis. We have investigated the effect of pharmacological modulation of PKC activity in DU-145 human androgen-independent prostatic carcinoma cells. The apoptotic death of these cells is characterized by the acquisition of classical apoptotic morphology and generation of > or = 1-mbp and 450- to 600-kbp DNA fragments in attached preapoptotic cell populations prior to cellular detachment and accrual of 30- to 50-kbp DNA fragments. We found that induction of apoptosis was arrested by downregulation of PKC activity and not by transient activation or inhibition of the enzyme. Concentrations and durations of exposure to phorbol esters that downregulated PKC activity correlated with inhibition of VP-16 or melphalan-induced morphological apoptosis and generation of the 30-to-50-kbp DNA fragments. Chronic exposure to phorbol-12,13-dibutyrate (PDBu) did not, however, suppress production of the > or = 1-mbp and 450- to 600-kbp DNA fragments found in preapoptotic cell populations, suggesting that PKC downregulation may interfere with the transition between a preapoptotic cell and an apoptotic cell. PKC isozyme analysis revealed that chronic PDBu treatment caused downregulation of PKC-alpha and -epsilon in DU-145 cells. Using concentrations of the PKC inhibitor UCN-01 that were consistent with PKC-alpha inhibition (but not PKC-epsilon inhibition), however, did not mimic the effects of chronic PDBu treatment, implying that downregulation of PKC-epsilon may be of particular importance. Together, these findings suggest that phorbol esters may act as tumor promoters by suppressing apoptosis.
蛋白激酶C（PKC）与细胞凋亡信号的传导有关。我们研究了PKC活性的药理调节对DU-145人雄激素非依赖性前列腺癌细胞的影响。这些细胞的凋亡性死亡表现为获得经典的凋亡形态，在细胞脱离之前，附着的凋亡前细胞群体中产生≥1兆碱基对和450至600千碱基对的DNA片段，以及积累30至50千碱基对的DNA片段。我们发现，PKC活性的下调可阻止细胞凋亡的诱导，而该酶的瞬时激活或抑制则不能。下调PKC活性的佛波酯的浓度和暴露持续时间与VP-16或美法仑诱导的形态学凋亡及30至50千碱基对DNA片段的产生受到抑制相关。然而，长期暴露于佛波醇-12,13-二丁酸酯（PDBu）并未抑制凋亡前细胞群体中发现的≥1兆碱基对和450至600千碱基对DNA片段的产生，这表明PKC下调可能会干扰凋亡前细胞与凋亡细胞之间的转变。PKC同工酶分析显示，长期PDBu处理导致DU-145细胞中PKC-α和-ε下调。然而，使用与抑制PKC-α（而非PKC-ε）一致的PKC抑制剂UCN-01浓度，并未模拟长期PDBu处理的效果，这意味着PKC-ε的下调可能尤为重要。这些发现共同表明，佛波酯可能通过抑制细胞凋亡而起到肿瘤促进剂的作用。
