中等强度有氧运动预干预下调小鼠骨骼肌TNF-R1水平并抵抗阿霉素诱导的骨骼肌萎缩

Pre-moderate-intensity Aerobic Exercise Reduces the Level of TNF-R1 in Skeletal Muscle of Mice and Protects Against Doxorubicin-induced Skeletal Muscle Atrophy

目的探讨进行长期中等强度有氧运动能否通过下调骨骼肌中TNF-α及其受体TNF-R1水平抵抗阿霉素诱导的骨骼肌萎缩,为运动抵抗阿霉素副作用提供方法建议。方法选用36只3月龄雄性C57BL/6小鼠,适应性喂养1周后,随机分为对照组(SED)和运动组(EXE),运动组进行8周中等强度跑台运动,对照组正常饲养;最后一次训练后进行最大跑速测试,结束后从SED和EXE组中随机抽取6只小鼠取材;剩余小鼠,SED组平均分为生理盐水组(SED-SAL)和阿霉素给药组(SED-DOX),EXE组平均分为运动后生理盐水组(EXE-SAL)和运动后阿霉素给药组(EXE-DOX)。SED-DOX组、EXE-DOX组小鼠腹腔注射阿霉素;SED-SAL组、EXE-SAL组小鼠注射对应体积生理盐水。给药结束后4组小鼠同时取材,取双侧股四头肌,H&E染色统计肌纤维横截面积,ELISA检测血浆TNF-α水平,WB检测股四头肌TNF-α、TNF-R1、PGC-1α、MuRF-1、Caspase-8蛋白表达水平。结果(1)与注射生理盐水的小鼠相比,注射阿霉素的小鼠最大跑速显著下降,体重、股四头肌质量显著下降,Ⅰ型、ⅡA型肌纤维数量显著减少,肌纤维横截面积显著减小,阿霉素给药导致小鼠骨骼肌萎缩;而8周中等强度有氧运动预干预,能显著减缓阿霉素引起的一系列骨骼肌不良反应,改善阿霉素导致的骨骼肌萎缩。(2)与注射生理盐水的小鼠相比,注射阿霉素的小鼠血浆TNF-α水平、股四头肌TNF-α及TNF-R1水平显著升高,且股四头肌中MuRF-1、Caspase-8蛋白表达水平显著升高,而8周中等强度有氧运动能显著抑制股四头肌TNF-α、TNF-R1、MuRF-1、Caspase-8水平。(3)8周中等强度有氧运动提高小鼠最大跑速,激活PGC-1α,抑制股四头肌TNF-α及TNF-R1表达水平。结论8周中等强度有氧运动抑制股四头肌TNF-α、TNF-R1表达水平,抑制阿霉素给药后TNF-α/TNF-R1通路下游的蛋白质降解和细胞凋亡反应,这可能是运动抵抗阿霉素诱导小鼠股四头肌萎缩的原因之一。

Objective The purpose of this study was to explore whether long-term moderate intensity aerobic exercise before doxorubicin(DOX)administration can reduce TNF-αand its receptor TNF-R1 levels in skeletal muscle to resist DOXinduced skeletal muscle atrophy,to provide a certain theoretical basis for exercise to avoid chemotherapy side effects.Methods Thirty-six 3-month-old male C57BL/6 mice were randomly separated into sedentary group(SED)and exercise group(EXE)after they adapted to the environment.The EXE mice were given moderate intensity treadmill running for 8 weeks,and the SED mice were fed normally.After the last exercise,6 mice randomly selected from both SED group and EXE group and then sacrificed.The remaining mice,SED group were grouped into normal saline group(SED-SAL)and DOX chemotherapy group(SED-DOX),and EXE mice were grouped into normal saline group after exercise(EXE-SAL)and DOX chemotherapy group after exercise(EXE-DOX).The SED-DOX group and EXE-DOX group were treated with DOX chemotherapy intraperitoneally for two weeks.And the SED-SAL group and EXE-SAL group were injected with normal saline of corresponding volume.At the end of chemotherapy,all four groups sacrificed.Bilateral quadriceps femoris muscle were removed and used H&E staining to calculate the cross-sectional area(CSA)of muscle fibers,used ELISA to detect TNF-αlevels of plasma,and used WB to detect the protein expression levels of TNF-α,TNF-R1,PGC-1α,MuRF-1,Caspase-8.Results(1)Compared with the mice injected with normal saline,the body weight,quadriceps femoris muscle mass,maximum running speed,the number of typeⅠmuscle fiber and muscle fiber CSA of the mice injected with DOX were significantly decreased,indicating that DOX administration caused skeletal muscle atrophy.However,8-week moderate-intensity aerobic exercise intervention before DOX administration can reduce a series of skeletal muscle adverse reactions caused by DOX.(2)Compared with the mice injected with normal saline,the level of TNF-αin plasma,the levels of TNF-αand TNF-R1 in quadriceps femoris of the mice injected with DOX were significantly increased,and the protein expression levels of MuRF-1 and Caspase-8 in quadriceps femoris were also significantly increased.While 8-week moderate intensity aerobic exercise before chemotherapy can significantly inhibit the expression of TNF-α,TNF-R1,MuRF-1 and Caspase-8 in quadriceps.(3)8-week moderate-intensity aerobic exercise increased the maximum running speed,activated PGC-1α,improved the activity of mitochondria-related proteins,and inhibited the expression of TNF-αand TNF-R1 in quadriceps femoris of mice.Conclusion 8-week moderate intensity aerobic exercise intervention before DOX administration can inhibit TNF-αand TNF-R1 expression level in quadriceps femoris and inhibit the protein degradation and apoptosis reaction which may be the downstream of TNF-α/TNF-R1 pathway.This might be one of the potential pathways for exercise to resistance to doxorubicin-induced quadriceps femoris atrophy in mice.