Lucas M, Sánchez-Margalet V, Sanz A, Solano F
Departamento de Bioquímica Médica y Biología, Hospital Universitario Virgen Macarena, Facultad de Medicina, Sevilla, Spain.
Biochem Pharmacol. 1994 Feb 11;47(4):667-72. doi: 10.1016/0006-2952(94)90129-5.
The putative protein kinase C (PKC) inhibitors polymyxin B and staurosporine were used to test the influence of PKC activity on the viability of lymphocytes. The cytotoxic effect of polymyxin B was characterized and it was found to be both time and dose dependent, with an LD50 in micromolar range, and counteracted by phorbol myristate acetate (PMA). To explore further the possible mechanism of action involved in polymyxin B-induced cell death, PKC activity and intracellular calcium were measured in polymyxin B-challenged lymphocytes. Polymyxin B inhibited PKC activity in both resting (25% inhibition) and PMA-stimulated (50% inhibition) cells, and increased intracellular calcium without disruption of the plasma membrane, a signal which is known to trigger apopotosis. Additionally, a number of experiments were conducted to assess the effect of staurosporine on PKC activity, cell growth, cell death and survival of mature lymphocytes. Staurosporine inhibited PKC activity in a dose-dependent manner (Ki close to 1 microM) and this effect correlated to some extent with the inhibition of [3H]thymidine incorporation and the breakdown of DNA into oligonucleosome-sized fragments. These results support the hypothesis that PKC is involved in the survival of mature lymphocytes undergoing apoptosis.
使用推定的蛋白激酶C（PKC）抑制剂多粘菌素B和星形孢菌素，来测试PKC活性对淋巴细胞活力的影响。对多粘菌素B的细胞毒性作用进行了表征，发现其具有时间和剂量依赖性，半数致死剂量在微摩尔范围内，并且可被佛波酯（PMA）抵消。为了进一步探究多粘菌素B诱导细胞死亡可能涉及的作用机制，对受到多粘菌素B刺激的淋巴细胞中的PKC活性和细胞内钙进行了测量。多粘菌素B在静息细胞（抑制25%）和PMA刺激的细胞（抑制50%）中均抑制PKC活性，并增加细胞内钙含量，且不破坏质膜，这是一种已知可触发细胞凋亡的信号。此外，还进行了多项实验，以评估星形孢菌素对PKC活性、细胞生长、细胞死亡以及成熟淋巴细胞存活的影响。星形孢菌素以剂量依赖性方式抑制PKC活性（抑制常数接近1微摩尔），并且这种作用在一定程度上与抑制[3H]胸腺嘧啶核苷掺入以及DNA降解为寡核小体大小的片段相关。这些结果支持了PKC参与经历凋亡的成熟淋巴细胞存活的假说。
