高原训练中运动员骨骼肌萎缩及分子网络调控机制研究

It was shown that only 40% of altitude training improved the athletic performance. The decrease of strength in athletes occurred in altitude acclimatization was believed as one of the most important factors,which caused the failure of the altitude training. Theoretically, the strength decrease is related to the hypoxia induced muscle atrophy during altitude exposure. However, no direct research focused on this phenomenon has been performed on human body and the underling mechanisms are still unknown. During the altitude training, hypoxia induces the muscular protein degradation and the synthesis suppression; on the other side, exercise facilitates the muscular protein synthesis. Therefore, the molecular regulation behind the interaction of the hypoxia and exercise stimuli must be more complicated than any one of the factors acting alone during altitude training. It is necessary to map the molecular regulation network to investigate the principles and mechanisms of the skeletal muscle atrophy during altitude training. In the project, we will investigate the effects of 2300m altitude training on the strength and morphologic parameters of skeletal muscle in athletes, test the human whole genome mRNA and miRNA expression with the muscle biopsy sampling from vastus lateralis, construct the regulation network of mRNA and miRNA and explore the key regulator in the signaling cascade with bioinformatics analysis; at the same time, we are going to test the miRNA in circulating blood which related the specific changes in skeletal muscle miRNA to provide a convenient and reliable index to evaluate the muscle atrophy during altitude training in athletes.

资料表明，高原训练的成功率只有40%，原因之一与高原训练中运动员力量丢失有关。理论上认为，高原训练中运动员力量丢失与高原低氧诱导的骨骼肌萎缩有关，但至今没有人体实验予以证实，更不了解其发生机制。高原训练中不仅存在低氧诱导骨骼肌蛋白质分解加强、合成减弱，进而导致骨骼肌萎缩的作用，也存在运动对骨骼肌合成的促进作用，其调控机制远比单纯的高原暴露复杂，有必要以分子网络为突破口，研究其调控规律与作用。本项目拟观察海拔3700m高原抗阻训练对运动员骨骼肌形态及收缩力量的影响，用肌肉活检技术提取运动员骨骼肌样本进行全基因组mRNA表达谱扫描，用生物信息学手段筛选分子调控的关键节点，并用分子生物学实验方法予以证实，以阐明高原训练中运动员骨骼肌萎缩的分子机制。

本项目通过基因芯片技术筛选高原抗阻练习后骨骼肌差异表达基因，运用生物信息学手段筛选关键调控通路并构建分子调控网络，探寻高原抗阻练习抗骨骼肌萎缩的分子调控机制。9名男性受试者按照ACSM力量练习指南在平原进行隔天抗阻练习，共10天，以验证练习方法的有效性。12名男性受试者随机分为高原对照组 (C，n=6) 和训练组 (HR，n=6)，在3700米高原暴露10天。期间，C组日常生活，HR组进行与平原组相同的抗阻练习。在高原暴露前、后，使用双能X线吸收测量法测试骨骼肌和脂肪含量；磁共振扫描测量大腿肌肉横截面积(CSA)；等速力量系统测量下肢力量。肌肉活检取股外侧肌中部骨骼肌，进行全基因组mRNA表达谱基因芯片扫描。使用生物信息学方法对高原抗阻练习后的差异表达基因进行功能注释、关键调控通路分析和转录因子预测并构建分子调控网络。用RT-qPCR和WB法验证关键蛋白。. 本研究发现，10天高原暴露后，对照组体重、全身瘦体重、腿部瘦体重和CSA显著性下降，训练组无显著性改变，表明高原抗阻练习可有效防止瘦体重和骨骼肌体积的下降；通过1.5倍差异基因筛选，高原抗阻练习后有432个差异表达基因，192个上调，240个下调；通过KEGG分析，发现 FOXO、胰岛素和ErbB等信号通路影响骨骼肌体积；通过转录因子调控网络构建，发现Nkx2-5处于核心位置；通过蛋白互作网络构建，发现SMAD3、MAPK1、MYC和ERBB2起核心作用；通过基因共表达网络构建，发现MAPK13、MYOT、EIF4B、TMOD4、GLI3、ACAT1、TUSC2、PTP4A1和NDUFS1 等9个核心基因可能与抗肌萎缩相关。本研究为进一步解析高原抗阻训练防止骨骼肌萎缩的分子机制提供了研究基础。.