8周中等强度低负荷量运动对增龄大鼠骨骼肌蛋白质组差异表达的影响

Effects of Eight Weeks Low-loads of Medium Intensity Exercise on the Proteome Differential Expression of the Aged Rat's Skeletal Muscle

目的:运用比较蛋白质组学方法研究8周中等强度低负荷量运动对增龄大鼠腓肠肌蛋白质组表达图谱的影响,初步筛选出体育运动延缓骨骼肌衰老的差异蛋白。方法:将12只18月龄雌性SD大鼠随机分为实验组(n=6)和平行对照组(n=6)。实验组经中等强度(60%～75%VO2max)、低负荷量(每天每次15min,5天/周)跑台运动,与对照组一起提取腓肠肌全蛋白,进行蛋白质固相-pH梯度聚丙烯酰胺凝胶电泳(2-DE)分离,随后采用PDQuest软件进行图像分析和数据采集。选择运动后差异表达量大于2倍的备选目标蛋白点进行胶内原位酶解与质谱鉴定。结果:运动后腓肠肌重量增加30.0%(P<0.05),腓肠肌指数增加37.5%(P<0.01);差异表达量大于1倍的蛋白点共19个,取大于2倍的6个蛋白点经质谱鉴定为热应激同源蛋白70(HSC70)、26S蛋白酶体调节亚基6B分子(S6B)、肌动蛋白结合蛋白(Cofilin)、线粒体乙醛脱氢酶2(ALDH2)、α-酮戊二酸脱氢酶复合体(α-KG-DHC)、细胞膜修复相关蛋白(TRIM72)。结论:1)8周中等强度低负荷量运动后增龄大鼠腓肠肌蛋白质组发生显著性变化;2)成功筛选出6种目标差异蛋白,其表达量发生的适应性变化表明,8周中等强度低负荷量运动可能通过同时调控3个途径保护Sarcopenia:①调控衰老骨骼肌的线粒体代谢酶、肌动蛋白结合蛋白、热休克同源蛋白70的表达来抑制肌细胞凋亡;②下调26S蛋白水解酶表达抑制蛋白质分解,进而拮抗Sarcopenia的骨骼肌蛋白质丢失;③促进细胞膜修复蛋白表达改善Sarcopenia的肌细胞膜修复功能。

Objective：The differential proteome expression map of rats’skeletal muscle between aging and exercise group was established to screen significant target protein response to 8 weeks low-loads of medium intensity exercise by means of proteomics,and filtrating out the objective proteins which exercise postpone skeletal muscle aging.Methods：Dividing 12 male Aged（16-month-old） SD rats into exercise and parallel control groups at random（n=6）,the exercise group had a 8 weeks treadmill training with medium intensity（60%~75%VO2max）,low-loads（15 min/days,5 days/week）,together with the control group,preparing for whole proteins samples of gastrocnemius,and separating the protein samples by utilizing two dimensional gel electrophoresis（2-DG）,and analyzing image and collecting data by using PD Quest.The alternatively objective protein spots whose differential expression volume＆gt;2 folds were selected for elenzymologic extraction and MS/ MS identification.Result：There were 19 points expressed＆gt;1 folds.Taking out 6 whose differential expression volume＆gt;2 folds were selected for elenzymologic extraction and MS/MS identification,they were heat shock cognate 70 kDa protein（HSC70）,26S protease regulatory subunit 6B（S6B）,actin binding protein（Cofilin）,aldehyde dehydrogenase 2（ALDH2）,α-ketoglutarate dehydrogenase complex（α-KGDH）,membrane repair relative protein tripartite motif-containing protein 72（TRIM72）.Conclusion：1） Using the methods of proteoimcs,the change of gastrocnemius protein in exercise rat s were identified.2） We succeeded in filtrating 6 objective proteins which their expression volume had an adaptive changes,it indicated that 8 weeks of low-loads medium intensity exercise may protect sarcopenia by regulation three pathways：① it could bring a series of well changes such as improving the aging skeletal muscle muscle mitochondrial metabolizing enzymes,Cofilin,HSC70 inhibit muscle cell apoptosis;② it could downregulate S6B expression to inhibit protein degradation,which antagonizes the aged rat’s skeletal muscle protein loss;③it could upregulate TRIM72 expression to improve the sarcopenia myocyte plasma membrane repair dysfunction.