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低氧性微损伤时大鼠腓肠肌抗肌营养不良蛋白和结蛋白的变化

徐玉明, 李俊平, 王瑞元

School of Physical Education and Health, Hangzhou Normal University,Hangzhou 310036, China； Graduate School, Beijing Sport University, Beijing 100084, China

摘要

为了观察低氧性微损伤时大鼠腓肠肌抗肌营养不良蛋白(dystrophin)和结蛋白(desmin)的变化特征及其调节机制，本实验将雄性Sprague-Dawley 大鼠40 只随机分为常氧对照组和10% 氧浓度的低氧刺激1、2、4、7 d 组(n=8)，每组各取2 只大鼠进行伊文氏蓝(EBD)腹腔注射来检测肌纤维膜通透性，同时用异染ATP 酶法鉴别EBD 阳性肌纤维的类型；其余6 只采用Western blot、RT-PCR 和荧光法进行蛋白含量和基因表达等测定。结果显示，低氧第1 天即出现EBD 阳性纤维，主要为IIA和IIB 型。各低氧组的EBD 阳性细胞率和平均光密度均大于常氧对照组(P<0.05)。低氧第1 天dystrophin 和desmin 蛋白显著增加或有增加趋势，第2 天有所下降，第4 天又急剧增加，第7 天恢复至接近对照组水平，其变化呈波动性起伏；低氧组基因表达在第2 天开始大幅度增加，与蛋白含量变化趋势相对应。除了低氧第2 天外，其余各组钙激活中性蛋白酶(calpain)总活性均高于常氧对照组。热休克蛋白HSP70 和HSP90 分别在低氧第4 天和第7 天增加(P<0.05)。上述结果表明，机械应力并非细胞膜完整性破坏的唯一原因，但是仅靠低氧引起腓肠肌微损伤时的dystrophin 和desmin 变化与离心运动不一样，二者蛋白含量并没有出现明显的丢失。低氧性肌损伤的表现形式可能与肌纤维动员类型以及calpain 活性有密切关系。

关键词： 低氧; 大鼠; 腓肠肌; 膜完整性; 抗肌营养不良蛋白; 结蛋白; 肌损伤

分类号：G804.2；R363

[Changes of dystrophin and desmin in rat gastrocnemius under micro-damage induced by hypoxia.] [Ariticle in Chinese]

XU Yu-Ming, LI Jun-Ping, WANG Rui-Yuan

杭州师范大学体育与健康学院，杭州 310036； 北京体育大学研究生院，北京 100084

Abstract

To explore the changes and regulation mechanism of dystropin and desmin under muscle injury without mechanic stress, 40male Sprague-Dawley rats were randomly divided into 5 groups, which included normoxia control and hypoxia groups for 1, 2, 4 and7 d with 10% O2. Two rats from each group were examined for sarcolemma integrity using Evans blue dye (EBD) and EBD-positivefiber typing by metachromatic dye-ATPase method. The rest six rats from each group were analyzed for the changes of protein contentand gene expression using Western blot, RT-PCR and fluorescence assays. The results showed that the EBD-positive muscle fibers,mainly type IIA and type IIB, appeared at 1 d after hypoxia exposure. Both the ratio of EBD-positive cell and the mean fluorescencedensity were significantly higher in hypoxia groups than those in control group (P<0.05). The contents of dystrophin and desminfluctuated after hypoxia exposure, increased at 1 d, decreased at 2 d, increased dramatically again at 4 d, and returned to a normal levelat 7 d. Consistently, the gene expression began to increase significantly after 2 d. The total activity of calpain was significantly higherin hypoxia groups at 1, 4 and 7 d. Significantly higher levels of HSP70 and HSP90 were also observed at 4 and 7 d, respectively (P<0.05).These results suggest that the mechanical stress is not the only cause of damage of sarcolemma membrane integrity. In contrast toeccentric contraction, hypoxia-induced muscle damage is not accompanied by the loss of dystrophin and desmin. The types of musclefibers recruited by motor units and the activities of calpain may be important in hypoxia-induced damage of sarcolemma membraneintegrity.

Key words: Hypoxia; rat; gastrocnemius; sarcolemmal integrity; dystrophin; desmin; muscle damage

收稿日期：2010-03-15　　录用日期：2010-05-17

通讯作者：王瑞元　　E-mail: wangry@vip.sina.com

引用本文：

徐玉明, 李俊平, 王瑞元. 低氧性微损伤时大鼠腓肠肌抗肌营养不良蛋白和结蛋白的变化[J]. 生理学报 2010; 62 (4): 339-348.

XU Yu-Ming, LI Jun-Ping, WANG Rui-Yuan. [Changes of dystrophin and desmin in rat gastrocnemius under micro-damage induced by hypoxia.] [Ariticle in Chinese] . Acta Physiol Sin 2010; 62 (4): 339-348 (in Chinese with English abstract).