Department of Medicine, Uniformed Services University of the Health Sciences, Bethesda, Maryland, United States of America.
PLoS One. 2018 Apr 24;13(4):e0196277. doi: 10.1371/journal.pone.0196277. eCollection 2018.
Increases of the activity of mitochondrial electron transport chain generally lead to increases of production of ATP and reactive oxygen species (ROS) as by-products. MnSOD is the first line of defense against the stress induced by mitochondrial ROS. Our previous studies demonstrated that EAE progression increased Na,K-ATPase activity in the mouse kidney cortex. Since mitochondria are the major source of ATP, our present studies were sought to determine whether EAE progression increased mitochondrial activity. We found that severe EAE increased mitochondrial complex II and IV activities without significantly affecting complex I activity with corresponding increases of ROS in the isolated mitochondria and native kidney cortex. Severe EAE augmented both cytosolic and mitochondrial MnSOD protein levels and activities and decreased the specific activity of mitochondrial MnSOD when the total mitochondrial MnSOD activity was normalized to the protein level. Using HEK293 cells as a model free of interference from immune reactions, we found that activation of Na,K-ATPase by monensin for 24 hours increased complex II activity, mitochondrial ROS and MnSOD protein abundance, and decreased the specific activity of the mitochondrial MnSOD. Inhibition of Na,K-ATPase by ouabain or catalase attenuated the effects of monensin on the mitochondrial complex II activity, ROS, MnSOD protein level and specific activity. Kockdown of MnSOD by RNAi reduced the mitochondrial ability to generate ATP. In conclusion, EAE increases mitochondrial activity possibly to meet the energy demand from increased Na,K-ATPase activity. EAE increases mitochondrial MnSOD protein abundance to compensate for the loss of the specific activity of the enzyme, thus minimizing the harmful effects of ROS.
线粒体电子传递链活性的增加通常会导致 ATP 和活性氧(ROS)作为副产物的产生增加。MnSOD 是对抗线粒体 ROS 诱导应激的第一道防线。我们之前的研究表明,EAE 进展会增加小鼠肾脏皮质中的 Na,K-ATPase 活性。由于线粒体是 ATP 的主要来源,我们目前的研究旨在确定 EAE 进展是否会增加线粒体活性。我们发现严重的 EAE 增加了线粒体复合物 II 和 IV 的活性,而对复合物 I 的活性没有显著影响,同时在分离的线粒体和天然肾脏皮质中增加了 ROS。严重的 EAE 增加了细胞质和线粒体 MnSOD 蛋白水平和活性,并在将线粒体 MnSOD 总活性归一化为蛋白水平时降低了线粒体 MnSOD 的比活性。使用 HEK293 细胞作为一种不受免疫反应干扰的模型,我们发现,莫能菌素激活 Na,K-ATPase 24 小时会增加复合物 II 的活性、线粒体 ROS 和 MnSOD 蛋白丰度,并降低线粒体 MnSOD 的比活性。哇巴因或 catalase 抑制 Na,K-ATPase 会减弱莫能菌素对线粒体复合物 II 活性、ROS、MnSOD 蛋白水平和比活性的影响。MnSOD 的 RNAi 敲低降低了线粒体产生 ATP 的能力。总之,EAE 增加线粒体活性可能是为了满足因 Na,K-ATPase 活性增加而产生的能量需求。EAE 增加线粒体 MnSOD 蛋白丰度以补偿酶比活性的丧失,从而最小化 ROS 的有害影响。