Ying Yang, Yun Ji, Guoyao Wu, Kaiji Sun, Zhaolai Dai, Zhenlong Wu
State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University, Beijing 100193, China.
State Key Laboratory of Animal Nutrition, College of Animal Science and Technology, China Agricultural University, Beijing 100193, China; Department of Animal Science, Texas A&M University, College Station, TX 77843, USA.
Exp Gerontol. 2015 May;65:35-41. doi: 10.1016/j.exger.2015.03.004. Epub 2015 Mar 10.
Dietary methionine restriction (MetR) has been reported to improve hepatocyte function in mammals. However, the underlying mechanisms remain largely unknown. This study was conducted with a swine model to test the hypothesis that MetR decreases generation of reactive oxygen species (ROS) and attenuates oxidative damage in hepatic mitochondria. Twenty-four 35-day old pigs were fed a control diet or a Met-restricted diet for two weeks. Liver mitochondria were isolated to determine: 8-oxodG in mitochondrial DNA, oxidative-derived proteins markers, including glutamic semialdehyde (GSA), aminoadipic semialdehydes (AASA), carboxyethyl-lysine (CEL), carboxymethyl-lysine (CML), and malondialdehyde lysine (MDAL), mitochondrial H2O2 generation rate; rates of oxygen consumption; free radical leak (FRL); anti-oxidative capacity, electron transport complex activity; and protein abundances of respiratory chain complex subunits (NDUFA9, SDHA, Core 2, and Cox 1), manganese superoxide dismutase (MnSOD), and apoptosis-inducing factor (AIF). Compared with the control, MetR decreased mitochondrial 8-oxodG content, H2O2 generation, FRL (P<0.05), and increased rates of oxygen consumption. Abundances of markers for protein oxidative damage, including GSA, AASA, CEL, and CML, were decreased (P<0.05) by 40%, 30%, 32%, and 28%, respectively, compared with the control. Western blot analysis revealed that MetR decreased (P<0.05) the protein abundances of complex subunits, NDUFA9 and AIF without affecting expression of SDHA, Core 2, Cox 1 or MnSOD. The complex I activity (P<0.05) were lowered in MetR group as compared with that of control. Collectively, our findings indicate that dietary MetR decreases mitochondrial ROS generation primarily via inhibiting complex I activity and ROS generation rather than augmenting anti-oxidative capacity, thereby ameliorating oxidative damage to hepatic mitochondrial DNA and proteins.
据报道，饮食中蛋氨酸限制（MetR）可改善哺乳动物的肝细胞功能。然而，其潜在机制在很大程度上仍不清楚。本研究采用猪模型来检验MetR可减少活性氧（ROS）生成并减轻肝线粒体氧化损伤这一假设。将24头35日龄仔猪分为两组，分别饲喂对照日粮或蛋氨酸限制日粮，为期两周。分离肝脏线粒体以测定：线粒体DNA中的8-氧代脱氧鸟苷（8-oxodG）、氧化衍生蛋白标志物，包括谷氨酸半醛（GSA）、氨基己二酸半醛（AASA）、羧乙基赖氨酸（CEL）、羧甲基赖氨酸（CML）和丙二醛赖氨酸（MDAL）、线粒体H2O2生成率、氧气消耗率、自由基泄漏（FRL）、抗氧化能力、电子传递复合体活性以及呼吸链复合体亚基（NDUFA9、SDHA、Core 2和Cox 1）、锰超氧化物歧化酶（MnSOD）和凋亡诱导因子（AIF）的蛋白质丰度。与对照组相比，MetR降低了线粒体8-oxodG含量、H2O2生成量、FRL（P<0.05），并提高了氧气消耗率。与对照组相比，蛋白质氧化损伤标志物GSA、AASA、CEL和CML的丰度分别降低了40%、30%、32%和28%（P<0.05）。蛋白质印迹分析显示，MetR降低了（P<0.05）复合体亚基NDUFA9和AIF的蛋白质丰度，而不影响SDHA、Core 2、Cox 1或MnSOD的表达。与对照组相比，MetR组的复合体I活性降低（P<0.05）。总的来说，我们的研究结果表明，饮食中的MetR主要通过抑制复合体I活性和ROS生成而非增强抗氧化能力来减少线粒体ROS生成，从而改善对肝线粒体DNA和蛋白质的氧化损伤。
