氢水对反复力竭运动大鼠海马组织氧化应激损伤及细胞自噬的影响

Effects of Hydrogen-rich Water on Rat Hippocampus' Oxidative Damage and Autophagy Induced by Repeated Exhaustive Exercise

目的:探讨补充氢水对反复力竭运动大鼠海马氧化应激损伤及细胞自噬的影响。方法:选取雄性SD大鼠48只,随机分为安静组(C),力竭运动组(EX),力竭运动+维生素E组(EX+E),力竭运动+氢水组(EX+H),每组12只。C组常规安静饲养。EX组、EX+E组和EX+H组均进行4周反复力竭运动,4周运动强度为(15 m/min×5 min→20 m/min×5 min→25 m/min×5 min→30 m/min×5 min→35 m/min,直至力竭,5 d/周),记录大鼠运动至力竭时间。EX组每次运动前30 min纯水灌胃4 mL,EX+E组维生素E溶液灌胃4 mL(20 mg),EX+H组氢水灌胃4 mL。训练结束后24 h取材。透射电子显微镜观察海马超微结构变化。海马组织抗氧化指标测定:硫代巴比妥(TBA)法测定丙二醛(MDA)含量,比色法测定总抗氧化能力(T-AOC)水平,黄嘌呤氧化酶法测定超氧化物歧化酶(SOD)活性;Western blot检测海马组织自噬相关蛋白,包括雷帕霉素靶蛋白(mTOR)、p-mTOR和微管相关蛋白1轻链3B(LC3B)表达。结果:EX+H组大鼠力竭时间较EX组显著延长(P<0.05);EX组海马神经元结构水肿严重,核内出现亮区,内质网、线粒体严重肿胀。EX+E组和EX+H组神经元结构水肿及内质网和线粒体肿胀程度明显减轻;EX组海马组织MDA、T-AOC水平及SOD活性均显著高于C组(P<0.01)。EX+H组MDA含量较EX组显著降低(P<0.05),T-AOC水平及SOD活性均显著升高(P<0.01,P<0.05)。EX+H组T-AOC水平较EX+E组显著升高(P<0.05);EX组海马组织mTOR、p-mTOR及LC3B蛋白表达均显著高于C组(P<0.01,P<0.05,P<0.01)。EX+H组mTOR、p-mTOR表达显著高于EX组(P<0.05)及EX+E组(P<0.01)。EX+H组LC3B表达显著低于EX组(P<0.05)。EX+H组LC3B表达与EX+E组比较无显著差异。结论:补充氢水可有效抑制反复力竭运动造成的海马组织超微结构、氧化应激损伤及细胞自噬现象,且补充氢水较传统抗氧化剂维生素E效果更显著。

Objectives： This study aims at detecting effects of hydrogen-rich water on rat hippocampus＇ oxidative damage and autophagy induced by repeated exhaustive exercise. Methods： Foru-eight male Sprague-Dawley rats were randomly divided into four groups （n = 12 ） ： control group （C）, exhaustive exercise group （Ex）, exercise ＋ vitamin E group （Ex ＋ E）, exercise ＋ hydrogen-rich water group （Ex ＋ H）. The rats in Ex, Ex ＋ E and Ex ＋ H groups finished 4 weeks repeated exhaustive treadmill exercise. The exercise program was increased gradually from light to exhausted, which starled at 15 m/min × 5 min, then increased to 20 m/min × 5 min, 25 m/min x 5 min, and ended at 35 m/min × exhausted, 5 days/week. The exhausted time was recorded. Ex group ratswere intragastrie administrations of pure water （4ml）, Ex ＋ E group rats were intragastrie administration of vitamin E （4ml, 20 mg）, and Ex ＋ H group rats were of hydrogen-rich water （4 ml）. Hippoeampus samples were ob- tained 24h after 4 weeks exercise. Ultrastruetural change of hippoeampus was observed by transmission electron microscope, content of MDA was tested by TBA, level of T-AOC was tested by eolorimetry, activity of SOD was measured by NBT, and protein expressions of mTOR, p-mTOR and LC3B were determined by western blotting. Results ： The exhausted time in Ex ＋ H group was longer than that in Ex group （P 〈 0.05）. The neurons in hippo- campus in Ex group had severe edema, the nuclei appeared bright area, and endoplasmie refieulum and mitoehon- dria had severe swelling. The swelling of endoplasmie retieulum and mitoehondria was less in Ex ＋ E and Ex ＋ H groups. MDA content, T-AOC level and SOD activity in hippoeampus in Ex group were higher than those in C group （P 〈 0.01）. In Ex ＋ H group, MDA content was lower than in Ex group （P 〈 0.05）, T-AOC level and SOD activity were higher than in Ex group （Ps 〈 0.05）. T-AOC level in Ex ＋ H group were higher than in Ex ＋ H group （P 〈0.05）. Protein expressions of mTOR, p-mTOR and LC3B in Ex group was higher than in C group （Ps 〈0.05）. Protein expressions of mTOR and p-mTOR in Ex ＋ H group was higher than in Ex and Ex ＋ E group groups （Ps 〈0.05）. LC3B＇s expression in Ex ＋ E group was lower than in Ex group （P 〈0.05）. There was no difference of LC3B＇s expression in Ex ＋ H group and in Ex ＋ E group. Conclusion： Consumption of hy- drogen-rich water can reduce rat hippoeampus＇ ultrastruetural damage, oxidative damage and autophagy induced by exhaustive exercise.