梁巧琴,刘亚南,田振军.间歇运动降低炎症和蛋白泛素化降解改善心梗导致的大鼠骨骼肌萎缩[J].北京体育大学学报,2018,41(9):59-69+87.
间歇运动降低炎症和蛋白泛素化降解改善心梗导致的大鼠骨骼肌萎缩
Interval Exercise Reduces Inflammation and Protein Ubiquitination Degradation to Improve Skeletal Muscle Atrophy Caused by Myocardial Infarction in Rats
投稿时间:2018-05-13  
DOI:10.19582/j.cnki.11-3785/g8.2018.09.009
中文关键词:  关键词:间歇运动  心肌梗死  蛋白泛素化降解  骨骼肌萎缩  Etanercept
英文关键词:Keywords: interval exercise  myocardial infarction  protein ubiquitination degradation  skeletal muscle atrophy  Etanercept
基金项目:国家自然科学基金资助项目(编号:31371199;31671240)。通信作者:田振军。
作者单位
梁巧琴 陕西师范大学体育学院暨运动生物学研究所陕西 西安 710119 
刘亚南 陕西师范大学体育学院暨运动生物学研究所陕西 西安 7101192.深圳高级中学(集团)广东 深圳 518000 
田振军 陕西师范大学体育学院暨运动生物学研究所陕西 西安 710119 
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中文摘要:
      摘要:目的:探讨间歇运动及间歇运动联合TNF-α抑制剂Etanercept对心梗大鼠骨骼肌MuRF1和Smyd1表征的影响及炎症水平与骨骼肌功能改善的关系。方法:雄性3月龄SD大鼠70只,随机分为假心梗组(S)、心梗组(MI)、心梗+间歇运动组(ME)、心梗+ Etanercept组(MIT)、心梗+生理盐水组(MIS)、心梗+间歇运动+ Etanercept组(MET)、心梗+间歇运动+生理盐水组(MES),每组10只。采用左冠状动脉结扎法制备心梗模型,S组只在心脏相同位置穿线不结扎。ME组、MET组和MES组术后1周采用大鼠跑台进行间歇运动干预。其中MIT组和MET组于术前3 d腹腔注射Etanercept(1.2 μg/g)1次,术后i.p. 1次/3 d×4周,MIS组和MES组注射定量生理盐水作为参照。各组训练结束后次日腹腔麻醉,测定心脏功能,之后冰浴条件下快速剥离并切取胫骨前肌,入甲醛固定或液氮冷藏备用。胫骨前肌重量/胫骨长度比值测定骨骼肌质量变化,免疫荧光测定骨骼肌HSP70和骨架蛋白Myosin表达变化及细胞横截面积,RT-qPCR测定骨骼肌MuRF1和MAFbx mRNA基因表达变化,Western Blot测定骨骼肌MuRF1、MAFbx、Smyd1、HSP90、TNF-α、NF-κB(pp65)、IL-6、IL-10、HSP70、Myosin和α-actinin蛋白表达变化,糜蛋白酶活性试剂盒检测骨骼肌蛋白酶体活性变化。结果:心梗导致大鼠骨骼肌质量系数、细胞横截面积、细胞骨架蛋白Myosin和α-actinin及肌纤维生成关键因子Smyd1表达显著降低(P<0.01),MuRF1、MAFbx、TNF-α、IL-6、NF-κB(pp65)和伴侣分子HSP90表达显著升高(P<0.01),间歇运动或Etanercept干预以及二者联合干预均可显著提高心梗大鼠骨骼肌质量系数、细胞横截面积、IL-10、Myosin、α-actinin和Smyd1蛋白表达(P<0.05, P<0.01),降低MuRF1、MAFbx、TNF-α、IL-6、NF-κB(pp65)和HSP90水平(P<0.05, P<0.01);心梗大鼠骨骼肌的细胞浆和细胞核MuRF1蛋白表达显著升高(P<0.05, P<0.01),糜蛋白酶活性增加(P<0.01),间歇运动可进一步升高其胞浆和胞核HSP70表达(P<0.01),降低MuRF1蛋白表达、细胞浆/细胞核HSP70比值和糜蛋白酶活性(P<0.05, P<0.01),且机体炎症水平与细胞浆/细胞核HSP70比值呈显著正相关(P<0.01)。结论:间歇运动或Etanercept干预或二者的共同干预均可显著提高心梗大鼠骨骼肌质量、骨架蛋白和Smyd1的表达,增加细胞横截面积,降低MuRF1、MAFbx和HSP90的表达,改善骨骼肌功能。
英文摘要:
      Abstract: Objectives: This study aimed to determine the effects of interval exercise and interval exercise combined with TNF-α inhibitor Etanercept on the expression of skeletal muscle MuRF1 and Smyd1 in myocardial infarction(MI) rats and the relationship between inflammatory and skeletal muscle function. Methods: Totally 70 three-month-old male SD rats were randomly divided into 7 groups: sham-operated group (S), MI group (MI), MI with interval exercise group (ME), MI with Etanercept group (MIT), MI with normal saline group (MIS), ME with Etanercept group (MET), ME with normal saline group (MES), 10 rats in each group. The MI model was induced by ligation of the left anterior descending (LAD) coronary artery, and the S group was only threaded without ligation in the same position of the heart. Rats in ME, MET and MES groups finished 4-week treadmill interval exercise one week after myocardial infarction. Among them, rats in MIT and MET groups were intraperitoneal injected with Etanercept (1.2ug/g) at three days before operation, and 1 time/3d after operation, 4 week in total. MIS and MIES groups were injected with quantitative saline. After the last training, abdominal anesthesia was performed and the cardiac function was evaluated at the following day. The tibialis anterior was dissected rapidly under the condition of ice bath, and stored in formaldehyde fixation or liquid nitrogen. The change of skeletal muscle mass index was evaluated by the ratio of tibialis anterior/tibials length. The changes of HSP70 and Myosin skeleton protein expression and the cross section area of tibialis anterior were determined by immunofluorescence. The gene expressions of MuRF1 and MAFbx were detected by RT-qPCR. The protein expressions of MuRF1, MAFbx, Smyd1, HSP90, TNF-α, NF-κB(pp65), IL-6, IL-10 and HSP70 were measured by Western blotting. The change of skeletal muscle proteinase activity was evaluated by chymotrypsin kits. Results: In MI rats, the expressions of skeletal muscle mass index, cells cross-sectional area, Myosin, α-actinin and Smyd1 decresed (P < 0.01), and the expressions of MuRF1, MAFbx, TNF-α, IL-6, NF-κB(pp65)and HSP90 increased (P < 0.01). Interval exercise or/and the Etanercept intervention remarkably improved the skeletal muscle mass index, cells cross-sectional area, IL-10, Myosin, α-actinin and Smyd1 expression in MI rats (Ps < 0.05), and reduced the level of MuRF1, MAFbx, TNF-α, IL-6, NF-κB(pp65)and HSP90 (Ps < 0.05); the protein expression of cytoplasm and nucleus MuRF1 increased dramatically in MI rats (Ps < 0.05), the activity of chymotrypsin-like proteasome was significantly enhanced(P < 0.01). Interval exercise further increased the expression of cytoplasm and nucleus HSP70 (P < 0.01), and reduced the protein expression of MuRF1 and the activity of chymotrypsin-like proteasome (P < 0.01). And there was a significant positive correlation between the level of inflammation and the ratio of cytoplasm/cytoplasmic nucleus HSP70 (P < 0.01). Conclusions: Interval exercise or injected Etanercept or both of them can significantly improve the skeletal muscle mass of MI rats, improve the expression of skeleton protein and Smyd1, increase the cell cross-sectional area, and reduce the expression of MuRF1, MAFbx and HSP90 to improve the function of skeletal muscle.
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