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目的:采用蛋白组学技术研究6周有氧跑台运动对C57BL/6小鼠骨骼肌蛋白表达的影响。通过筛选对比安静组与运动组小鼠骨骼肌差异表达蛋白,分析有氧运动引发的骨骼肌代谢相关信号通路变化,探讨运动增进健康、防治疾病的分子机理。方法:选用C57BL/6雄性小鼠40只,将其随机分为正常饮食组(NC,n=20)和正常饮食运动组(NE,n=20),均饲以基础饲料。NE组进行6周强度为75%VO2max的跑台运动。6周后提取小鼠股四头肌总蛋白,采用Bradford法检测蛋白浓度后进行蛋白质双向电泳(2-DE),采用ImageMaster 2D Platinum V5.0软件分析2-DE电泳图谱,并对差异表达蛋白点采用基质辅助激光解吸附离子化飞行时间质谱(MALDI-TOF-MS)和高效液相色谱-串联谱法(LC-MS/MS)进行分析、采用Mascot检索软件结合NCBI nr数据库鉴定差异表达蛋白,并运用相关分析软件对所得数据进行统计学处理。结果:经6周有氧跑台运动后,比较NC组与NE组小鼠骨骼肌蛋白表达图谱,以蛋白表达量变化1.5倍以上为显著性判定标准,共筛选出32个差异表达蛋白,排除高表达丰度蛋白背景信号后共选出6个差异表达蛋白。其中,与线粒体能量代谢相关的蛋白有3个,分别为呼吸链复合体Ⅰ(Nadh-Ubiquinone Oxidoreductase 18 Kda Ip Subunit),表达增加2.92倍,线粒体膜电压依赖阴离子通道1和2(Voltage-dependent Anion Channel,VDAC),表达上调1.82倍和3.51倍;与细胞膜修复相关的蛋白Trim72(Tripartite Motif-Containing Protein 72)表达增加2.10倍;糖代谢相关蛋白乳酸脱氢酶(L-Lactate Dehydrogenase B Chain,LDH-B)表达上调1.85倍;Ester Hydrolase C11Orf54 Homolog蛋白表达下降56%。结论:6周有氧运动引起小鼠骨骼肌32个蛋白表达显著改变,其中,与线粒体能量代谢相关蛋白、细胞膜修复相关蛋白、糖代谢相关蛋白等均受到影响。由于这些蛋白功能与骨骼肌能量代谢密切关系,提示有氧运动对促进骨骼肌能量代谢、预防代谢相关疾病具有重要作用。
OBJECTIVE: To study the effect of 6-week aerobic treadmill exercise on skeletal muscle protein expression in C57BL / 6 mice by proteomics. By screening and comparing the differentially expressed proteins in skeletal muscle of quiet mice and exercise mice, we analyzed the changes of skeletal muscle-related signal pathways induced by aerobic exercise and explored the molecular mechanism of exercise for improving health and preventing and curing diseases. Methods: Forty C57BL / 6 male mice were randomly divided into normal diet group (NC, n = 20) and normal diet group (NE, n = 20). NE group for 6 weeks treadmill exercise intensity of 75% VO2max. After 6 weeks, the total protein of quadriceps femoris was extracted. The protein concentration was determined by Bradford method and then analyzed by 2-DE. The 2-DE was analyzed by ImageMaster 2D Platinum V5.0 software. The differential expressed protein Point analysis was performed using matrix-assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF-MS) and high performance liquid chromatography-tandem spectroscopy (LC-MS / MS), using Mascot search software combined with the NCBI nr database to identify differentially expressed proteins , And the use of correlation analysis software for statistical analysis of the data obtained. RESULTS: After six weeks of aerobic treadmill exercise, the skeletal muscle protein expression profiles in NC group and NE group were compared. Fifty-five times of the protein expression level was used as the criterion of significance. 32 differentially expressed proteins were screened out A total of six differentially expressed proteins were selected after high expression of abundant protein background signal. Among them, there are three proteins related to mitochondrial energy metabolism, which are respectively 2.92 times higher than that of Nadh-Ubiquinone Oxidoreductase 18 Kda Ip Subunit. The mitochondrial membrane voltage is dependent on Voltage-dependent Anion (VDAC). The expression of Tripartite Motif-Containing Protein 72 (Trimite Motif-Containing Protein 72) increased by 1.82-fold and 3.51-fold respectively. The expression of L-lactate dehydrogenase B chain -B) was up-regulated by 1.85-fold; Ester Hydrolase C11Orf54 Homolog protein expression decreased by 56%. CONCLUSION: Aerobic exercise at 6 weeks significantly changes the expression of 32 proteins in skeletal muscle of mice, among which, the proteins related to mitochondrial energy metabolism, the proteins involved in membrane repair and the proteins related to glucose metabolism are affected. Because of the close relationship between these protein functions and skeletal muscle energy metabolism, it is suggested that aerobic exercise plays an important role in promoting energy metabolism of skeletal muscle and preventing metabolic-related diseases.