Department of Molecular Pharmacology, Rappaport Family Institute for Research in the Medical Sciences, Technion-Israel Institute of Technology, P.O. Box 9649, Bat-Galim, 31096, Haifa, Israel.
Apoptosis. 2010 Jul;15(7):753-68. doi: 10.1007/s10495-010-0460-5.
Erucylphosphohomocholine (ErPC3, Erufosine) was reported previously to induce apoptosis in otherwise highly apoptosis-resistant malignant glioma cell lines while sparing their non-tumorigenic counterparts. We also previously found that the mitochondrial 18 kDa Translocator Protein (TSPO) is required for apoptosis induction by ErPC3. These previous studies also suggested involvement of reactive oxygen species (ROS). In the present study we further investigated the potential involvement of ROS generation, the participation of the mitochondrial respiration chain, and the role of the mitochondrial F(O)F(1)-ATP(synth)ase in the pro-apoptotic effects of ErPC3 on U87MG and U118MG human glioblastoma cell lines. For this purpose, cells were treated with the ROS chelator butylated hydroxyanisole (BHA), the mitochondrial respiration chain inhibitors rotenone, antimycin A, myxothiazol, and the uncoupler CCCP. Also oligomycin and piceatannol were studied as inhibitors of the F(O) and F(1) subunits of the mitochondrial F(O)F(1)-ATP(synth)ase, respectively. BHA was able to attenuate apoptosis induction by ErPC3, including mitochondrial ROS generation as determined with cardiolipin oxidation, as well as collapse of the mitochondrial membrane potential (Deltapsi(m)). Similarly, we found that oligomycin attenuated apoptosis and collapse of the Deltapsi(m), normally induced by ErPC3, including the accompanying reductions in cellular ATP levels. Other inhibitors of the mitochondrial respiration chain, as well as piceatannol, did not show such effects. Consequently, our findings strongly point to a role for the F(O) subunit of the mitochondrial F(O)F(1)-ATP(synth)ase in ErPC3-induced apoptosis and dissipation of Deltapsi(m) as well as ROS generation by ErPC3 and TSPO.
先前有报道称，Erucylphosphohomocholine（ErPC3，Erufosine）能够诱导原本对细胞凋亡具有高度抗性的恶性神经胶质瘤细胞系发生凋亡，而对其非致瘤性对应物没有影响。我们之前还发现，线粒体 18 kDa 转位蛋白（TSPO）是 ErPC3 诱导细胞凋亡所必需的。这些先前的研究还表明活性氧（ROS）的参与。在本研究中，我们进一步研究了 ROS 生成的潜在参与、线粒体呼吸链的参与以及线粒体 F（O）F（1）-ATP（synth）酶在 ErPC3 对 U87MG 和 U118MG 人神经胶质瘤细胞系促凋亡作用中的作用。为此，用 ROS 螯合剂丁基羟基茴香醚（BHA）、线粒体呼吸链抑制剂鱼藤酮、抗霉素 A、米酵菌酸和解偶联剂 CCCP 处理细胞。此外，还研究了寡霉素和白藜芦醇分别作为线粒体 F（O）和 F（1）亚基的抑制剂对线粒体 F（O）F（1）-ATP（synth）酶的作用。BHA 能够减弱 ErPC3 诱导的细胞凋亡，包括通过心磷脂氧化测定的线粒体 ROS 生成，以及线粒体膜电位（Deltapsi（m））的崩溃。同样，我们发现，寡霉素减弱了 ErPC3 诱导的细胞凋亡和 Deltapsi（m）的崩溃，包括伴随的细胞内 ATP 水平降低。线粒体呼吸链的其他抑制剂以及白藜芦醇没有表现出这种作用。因此，我们的研究结果强烈表明，线粒体 F（O）F（1）-ATP（synth）酶的 F（O）亚基在 ErPC3 诱导的细胞凋亡和 Deltapsi（m）耗散以及 ErPC3 和 TSPO 诱导的 ROS 生成中起作用。
