Biomedical Center and Institute of Medical Biochemistry, Jessenius Faculty of Medicine in Martin, Comenius University in Bratislava, SK‑03601 Martin, Slovak Republic.
Department of Histology and Embryology, Jessenius Faculty of Medicine in Martin, Comenius University in Bratislava, SK‑03601 Martin, Slovak Republic.
Int J Mol Med. 2019 Jun;43(6):2420-2428. doi: 10.3892/ijmm.2019.4168. Epub 2019 Apr 16.
Mitochondria are crucial for neuronal cell survival and death through their functions in ATP production and the intrinsic pathway of apoptosis. Mitochondrial dysfunction is considered to play a central role in several serious human diseases, including neurodegenerative diseases, such as Parkinson's and Alzheimer's disease and ischemic neurodegeneration. The aim of the present study was to investigate the impact of transient global brain ischemia on the expression of selected proteins involved in mitochondrial dynamics and mitochondria‑associated membranes. The main foci of interest were the proteins mitofusin 2 (Mfn2), dynamin‑related protein 1 (DRP1), voltage‑dependent anion‑selective channel 1 (VDAC1) and glucose‑regulated protein 75 (GRP75). Western blot analysis of total cell extracts and mitochondria isolated from either the cerebral cortex or hippocampus of experimental animals was performed. In addition, Mfn2 was localized intracellularly by laser scanning confocal microscopy. It was demonstrated that 15‑min ischemia, or 15‑min ischemia followed by 1, 3, 24 or 72 h of reperfusion, was associated with a marked decrease of the Mfn2 protein in mitochondria isolated from the cerebral cortex, but not in hippocampal mitochondria. Moreover, a translocation of the Mfn2 protein to the cytoplasm was documented immediately after global brain ischemia in the neurons of the cerebral cortex by laser scanning confocal microscopy. Mfn2 translocation was followed by decreased expression of Mfn2 during reperfusion. Markedly elevated levels of the VDAC1 protein were also documented in total cell extracts isolated from the hippocampus of rats after 15 min of global brain ischemia followed by 3 h of reperfusion, and from the cerebral cortex of rats after 15 min of global brain ischemia followed by 72 h of reperfusion. The mitochondrial Mfn2 release observed during the early stages of reperfusion may thus represent an important mechanism of mitochondrial dysfunction associated with neuronal dysfunction or death induced by global brain ischemia.
线粒体通过其在 ATP 生成和细胞凋亡的内在途径中的功能，对于神经元细胞的存活和死亡至关重要。线粒体功能障碍被认为在包括神经退行性疾病（如帕金森病和阿尔茨海默病以及缺血性神经退行性疾病）在内的几种严重人类疾病中发挥核心作用。本研究旨在研究短暂全脑缺血对参与线粒体动力学和线粒体相关膜的选定蛋白质表达的影响。主要关注点是蛋白质融和蛋白 2（Mfn2）、动力相关蛋白 1（DRP1）、电压依赖性阴离子选择通道 1（VDAC1）和葡萄糖调节蛋白 75（GRP75）。对来自实验动物的大脑皮质或海马体的总细胞提取物和分离的线粒体进行了 Western blot 分析。此外，通过激光扫描共聚焦显微镜对细胞内 Mfn2 进行了定位。结果表明，15 分钟缺血，或 15 分钟缺血后再灌注 1、3、24 或 72 小时，与从大脑皮质分离的线粒体中 Mfn2 蛋白的明显减少有关，但在海马体的线粒体中则不然。此外，激光扫描共聚焦显微镜显示，全脑缺血后即刻，Mfn2 蛋白向细胞质易位，大脑皮质神经元中发生了这种情况。易位后，在再灌注期间观察到 Mfn2 表达减少。还记录了在全脑缺血 15 分钟后再灌注 3 小时的大鼠海马体和全脑缺血 15 分钟后再灌注 72 小时的大鼠大脑皮质的总细胞提取物中 VDAC1 蛋白水平显著升高。在再灌注的早期阶段观察到的线粒体 Mfn2 释放可能是与全脑缺血引起的神经元功能障碍或死亡相关的线粒体功能障碍的重要机制。
