Kubo T, Enokido Y, Yamada M, Oka T, Uchiyama Y, Hatanaka H
Division of Protein Biosynthesis, Osaka University, Japan.
Brain Res. 1996 Sep 16;733(2):175-83. doi: 10.1016/0006-8993(96)00542-2.
We previously reported that PC12h cells are killed by a high oxygen atmosphere. In this study, we further characterized this oxygen-induced cell death and found apoptotic features, as follows. Firstly, chromatin condensation was observed in cells cultured in a 50% O2 atmosphere. Secondly, cycloheximide and cordycepin, protein and RNA synthesis inhibitors, respectively, prevented the oxygen-induced cell death in PC12h cells, suggesting that it is mediated by an intracellular death program. Thirdly, NGF, CPT-cAMP and depolarization by high potassium medium also effectively inhibited this apoptotic cell death in PC12h cells. The effect of high K+ is thought to be mediated by the influx of Ca2+ into cells through voltage-dependent Ca2+ channels, because nifedipine, an L-type Ca2+ channel blocker, inhibited the effect of high K+. In addition, since the oxygen-induced apoptosis was blocked by the antioxidant vitamin E, this oxygen toxicity is suggested to be mediated by reactive oxygen species. To further characterize this oxygen-induced apoptosis at the molecular level, we used PC12 cells overexpressing the proto-oncogene bcl-2. Although a large number of PC12 cells transfected with the control vector died in a 50% O2 atmosphere within 6 days, bcl-2-transfected PC12 cells survived and proliferated. These findings suggested that our system using PC12 cells will be a useful model with which to analyze the molecular mechanisms of apoptosis induced by oxidative stress in neuronal cells.
我们之前报道过PC12h细胞会被高氧环境杀死。在本研究中，我们进一步对这种氧诱导的细胞死亡进行了表征，并发现了凋亡特征，如下所示。首先，在50%氧气环境中培养的细胞中观察到染色质浓缩。其次，蛋白质合成抑制剂环己酰亚胺和RNA合成抑制剂虫草素分别阻止了PC12h细胞中氧诱导的细胞死亡，这表明它是由细胞内死亡程序介导的。第三，神经生长因子（NGF）、环磷腺苷（CPT-cAMP）以及高钾培养基引起的去极化也有效抑制了PC12h细胞中的这种凋亡性细胞死亡。高钾的作用被认为是通过电压依赖性钙通道使钙离子流入细胞介导的，因为L型钙通道阻滞剂硝苯地平抑制了高钾的作用。此外，由于氧诱导的凋亡被抗氧化剂维生素E阻断，提示这种氧毒性是由活性氧介导的。为了在分子水平上进一步表征这种氧诱导的凋亡，我们使用了过表达原癌基因bcl-2的PC12细胞。尽管大量转染了对照载体的PC12细胞在50%氧气环境中6天内死亡，但转染了bcl-2的PC12细胞存活并增殖。这些发现表明，我们使用PC12细胞的系统将是一个有用的模型，可用于分析氧化应激诱导神经元细胞凋亡的分子机制。
