细胞色素C和Caspase-3在过度训练致大鼠急性心肌损伤中的作用及旋覆花素的干预效应

发布时间：2018-10-25 11:31

【摘要】：目的：近年过度训练逐渐成为运动医学、临床医学的研究热点。过度训练致急性心肌损伤（overtraining-induced acute myocardial injury，OTIAMI）是运动员发生猝死的重要原因之一，目前尚缺乏有效的防治措施，因此对OTIAMI机制及防治措施的研究具有重要的临床意义。本研究采用游泳力竭法建立过度训练致大鼠急性心肌损伤模型，探讨细胞色素C及Caspase-3在过度训练致急性心肌损伤中的作用及旋覆花素的干预效应，从而为OTIAMI的防治提供新的实验依据和治疗靶点。 方法：健康雄性Wistar大鼠48只，体重200～220g，采用随机数字表法，将大鼠随机分为3组：安静对照组（control，C组，n=8）、力竭运动组（exhaustive swimming，ES组，n=24）、旋覆花素干预组（InulaBritannica，IB组，n=16）。C组不做任何处理；ES组及IB组采用游泳力竭法建立大鼠过度训练急性心肌损伤模型，IB组于力竭前24h及力竭前即刻分别口腔灌入旋覆花素25ml/kg后进行游泳直至力竭。各组大鼠于实验前一天适应性游泳30min，次日再次放于水中游泳直至力竭。力竭标准：①沉入水底超过10s；②游泳动作明显不协调；③捞出后没有逃避反应，且放在平面上无法完成翻正反射。 ES组于力竭后即刻（ESI）、力竭后6h（ES6h）和力竭后24h（ES24h）；IB组于力竭后6h（IB6h）、力竭后24h（IB24h）分别随机抽取8只大鼠，乙醚麻醉下采集下腔静脉血3ml，，3000转/min，离心15min，取上清液，-20℃保存，采血后摘取心脏，取心尖同一部位心肌组织置于4%多聚甲醛固定。光镜下观察大鼠心肌组织病理学变化，采用末端脱氧核苷酸转移酶介导的缺口末端标记法（TUNEL)观察心肌细胞凋亡情况，ELISA法检测血清心肌肌钙蛋白I（cTnI）浓度，免疫组化法检测心肌组织细胞色素C及Caspase-3表达变化。 统计学处理采用SPSS13.0统计学软件进行分析，计量资料以均数±标准差（x±s）表示，组间比较以及ES组各时间点的比较采用单因素方差分析，P＜0.05为差异有统计学意义。 结果：1.心肌组织病理学变化C组大鼠心肌细胞呈椭圆形或短梭形，心肌纤维互相连接排列紧密，可见清晰的横纹。ES组大鼠力竭即刻见部分心肌纤维肿胀肥大，偶见心肌间质血管及毛细血管扩张；力竭后6h时可见部分心肌细胞的细胞核深染，有固缩现象，心肌纤维肿胀肥大明显，排列紊乱，出现不规整断裂；力竭后24h时心肌纤维肿胀肥大有所好转，心肌细胞间散在大量炎症浸润灶。IB组各时点心肌组织病理学变化明显减轻。 2.各组大鼠不同时点血清cTnI的比较与C组比较，ES组及IB组过度训练后各时点大鼠血清cTnI浓度升高（P＜0.05）；与ES组比较，IB组过度训练后6h、24h时大鼠血清cTnI浓度下降（P＜0.05）；与力竭后即刻相比，力竭后6h时大鼠血清cTnI浓度明显升高（P 0.05），24h时大鼠血清cTnI浓度有所降低(P＜0.05）。 3.各组大鼠不同时点心肌细胞凋亡指数的比较与C组比较，ES组及IB组力竭后各时点大鼠心肌细胞凋亡指数增加（P＜0.05）；与ES组比较，IB组力竭后6h及24h时大鼠心肌细胞凋亡指数明显降低（P＜0.05）；与力竭后即刻比较，力竭后6h及24h时大鼠心肌细胞凋亡指数增加（P＜0.05），且力竭后6h时达高峰，24h时有所下降（P＜0.05）。 4.各组大鼠心肌组织不同时点细胞色素C及Caspase-3表达的比较各组大鼠心肌组织各时间点Caspase-3与细胞色素C变化趋势一致。与C组比较，ES组及IB组力竭后各时点大鼠心肌组织细胞色素C及Caspase-3表达上调（P＜0.05）；与ES组比较，IB组力竭后6h、24h时大鼠心肌组织细胞色素C及Caspase-3表达下调（P＜0.05）；ES组力竭后6h、24h时与力竭后即刻比较，大鼠心肌组织细胞色素C及Caspase-3表达上调（P＜0.05），且力竭后6h时表达最强，24h时有所下调（P＜0.05）。 结论：1.游泳所致力竭可引起心肌损伤，但病变较轻，受损心肌可在24h内部分修复。 2.游泳致力竭运动可引起大鼠心肌细胞凋亡，心肌细胞凋亡可能为过度训练致大鼠心肌损伤的病理生理学基础之一。 3.细胞色素C及Caspase-3通过线粒体信号转导通路介导了过度训练致大鼠急性心肌损伤时心肌细胞凋亡。 4.旋覆花素可减轻过度训练致大鼠急性心肌损伤，其机制可能与其抑制心肌细胞凋亡相关信号转导因子Caspase-3及细胞色素C表达，减少心肌细胞凋亡有关，及时采取药物干预可能为OTIAMI防治的关键。
[Abstract]:Objective: Over-training has become a hot topic in sports medicine and clinical medicine in recent years. Overtraining induced acute myocardial injury (OTIAMI) is one of the most important causes of sudden death in athletes. The effects of cytochrome C and Caspase-3 on acute myocardial injury induced by over-training induced by over-training were studied by means of swimming exhaustion method. The effect of cytochrome C and Caspase-3 on acute myocardial injury was studied. Methods: Forty-eight healthy male Wistar rats were randomly divided into three groups: control group (control group, group C, n = 8), exhaustive exercise group (ES group, n = 24). 6) No treatment was done in group C; ES group and IB group used swimming exhaustion method to establish the model of acute myocardial injury in rats. After exhaustive swimming, the rats were allowed to swim for 30minutes the day before the experiment, and then put it again in water the next day until they were allowed to swim. Exhaustion. Exhaustive standard: the water is sunk to the water bottom for more than 10s; the swimming action of the swimming pool is obviously not coordinated; after fishing, no escaping reaction is avoided, and the turning is not completed on the plane. The results were as follows: after exhaustive exhaustion (ESI), 6h (ES6h) and 24h (ES24h) after exhaustive exhaustion, 8 rats were randomly sampled from the group IB after exhaustive exhaustion (IB6h). After exhaustive exhaustion, 8 rats were sampled at random, 3ml of venous blood were collected under ether anesthesia, then centrifuged for 15min, supernatant was removed, and blood was collected at -20 鈩

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