Davis M A, Chang S H, Trump B F
Department of Pathology, University of Maryland, School of Medicine, Baltimore 21201, USA.
Toxicol Appl Pharmacol. 1996 Nov;141(1):93-101. doi: 10.1006/taap.1996.0264.
H-ras oncogenes have been identified in greater than 50% of the most common forms of human neoplasia. Ras-related proteins have been postulated to mediated signal transduction pathways involving mitogen-activated protein (MAP) kinases and nuclear responses that may be involved in the induction of apoptosis. We examined whether expression of H-ras oncogene conferred resistance or susceptibility to the morphologic effects of the protein phosphatase inhibitor, okadaic acid, using a tumorigenic H-ras-transformed normal rat kidney epithelial cell line, NRK-H/6.1. We also examined whether okadaic acid induced apoptosis correlated with a differential effect on kinase activity in H-Ras-transformed cells as compared to the nontransformed NRK-52E cells. Treatment with various concentrations of okadaic acid produced rapid and extensive morphologic changes characteristic of apoptosis in both cell types. Equimolar okadaic acid concentrations for 2 or 4 hr resulted in cell detachment and loss of membrane integrity (as measured by propidium iodide uptake) in 74% (0.5 microM) and 78% (1.0 microM) of the H-Ras-transformed cells as compared to 8 and 25%, respectively, in the non-transformed cells. Furthermore, a higher basal level of kinase activity was observed in the H-Ras-transformed cells as compared to the nontransformed cells. Okadaic acid-induced apoptosis correlated with activation of members of the MAP kinase family, raf-1 and protein kinase C (PKC). These studies show that H-ras oncogene expression imparts selective susceptibility to cell death induced by phosphatase inhibition. The observed increase in susceptibility to okadaic acid-induced apoptosis appears to involve the modulation of raf-1, PKC, and MAP kinase activities. These findings may be significant in the elucidation of mechanisms for selective induction of cell death in tumor cells expressing H-ras oncogene.
在超过50%的最常见人类肿瘤类型中已鉴定出H-ras癌基因。据推测，Ras相关蛋白介导涉及丝裂原活化蛋白（MAP）激酶和可能参与细胞凋亡诱导的核反应的信号转导途径。我们使用致瘤性H-ras转化的正常大鼠肾上皮细胞系NRK-H/6.1，研究了H-ras癌基因的表达是否赋予对蛋白磷酸酶抑制剂冈田酸形态学效应的抗性或敏感性。我们还研究了冈田酸诱导的细胞凋亡是否与H-Ras转化细胞与未转化的NRK-52E细胞相比对激酶活性的差异效应相关。用不同浓度的冈田酸处理在两种细胞类型中均产生了快速且广泛的细胞凋亡特征性形态变化。与未转化细胞中分别为8%和25%相比，等摩尔浓度的冈田酸处理2或4小时导致74%（0.5微摩尔）和78%（1.0微摩尔）的H-Ras转化细胞发生细胞脱离和膜完整性丧失（通过碘化丙啶摄取测量）。此外，与未转化细胞相比，在H-Ras转化细胞中观察到更高的基础激酶活性水平。冈田酸诱导的细胞凋亡与MAP激酶家族成员raf-1和蛋白激酶C（PKC）的激活相关。这些研究表明，H-ras癌基因表达赋予对磷酸酶抑制诱导的细胞死亡的选择性敏感性。观察到的对冈田酸诱导的细胞凋亡敏感性增加似乎涉及raf-1、PKC和MAP激酶活性的调节。这些发现可能对阐明表达H-ras癌基因的肿瘤细胞中细胞死亡的选择性诱导机制具有重要意义。
