摘要
背景:前期研究表明亚健康疲劳状态时血清对体外培养骨骼肌细胞的生存活力、细胞超微结构、细胞线粒体膜细胞色素C氧化酶活性和线粒体能量负荷具有显著影响。目的:进一步运用二维凝胶电泳和质谱技术观察亚健康疲劳状态血时清对骨骼肌细胞蛋白质表达谱的影响。设计、时间及地点:单一样本观察,于2007-03/05在南方医科大学中医证候研究实验室和天津生物芯片技术有限责任公司蛋白质组学实验室完成。材料:亚健康疲劳状态时血清和正常状态时血清为自制;人骨骼肌细胞为Sciencell公司产品。方法:体外培养骨骼肌细胞,随机分为正常组和亚健康组,分别用含30%正常血清和30%亚健康疲劳者血清的DMEM/F12细胞培养液培养72h,收集细胞,提取细胞总蛋白。主要观察指标:通过PDQuest7.3.0软件分析二维凝胶电泳图谱,比较各组间蛋白表达量,然后利用质谱制作差异蛋白点的肽谱,确定差异蛋白,通过生物信息学分析所得蛋白的功能。结果:在2组凝胶中有15个蛋白质点显著差异表达,亚健康组中表达升高的蛋白质点有10个,下降有5个,其主要涉及能量代谢,核苷酸合成,细胞修复,应激等相关蛋白。结论:亚健康疲劳状态时血清可影响人骨骼肌细胞相关蛋白的表达。
BACKGROUND： Previous study showed sub-health fatigue serum has obvious effect on the viability and ultrastructure of human skeletal muscle cells （HSkMCs）, which also markedly affect the activity of cytochrome C oxidase （COX） and energy charge mitochondrial membrane of HSkMCs in vitro.
OBJECTIVE： To study the influence of sub-health fatigue serum on the protein expression of HSkMCs.
DESIGN, TIME AND SETTING： A single sample observation was performed at the Laboratory of syndrome manifestations of Chinese medicine, Southern Medical University, and Laboratory of Proteome, Tianjin Biochip Corporation from March to May 2007.
MATERIALS： The serum with healthy or sub-health fatigue was self-made. HSkMCs were provided by Sciencell Company.
METHODS： HSkMCs were cultured in vitro and randomly divided into the normal and sub-health groups, then co-cultured with DMEM/F12 （contains 30% healthy serum and 30% sub-health fatigue serum） for 72 hours. The total proteins were extracted after culture.
MAIN OUTCOME MEASURES： The two-dimensional electrophoresis images were analyzed with the PDQuest7.3.0 software package. The peptide mass fingerprinting （PMF） was made with MALDI-TOF-MS, then the identification and function of the protein was made via bioinformatics.
RESULTS： There were 15 protein dots differential expression in two groups. In the sub-health group, there were 10 proteins up-regulated and 5 proteins down-regulated. These proteins were mainly associated with cell cycle regulation, stress response proteins, energy metabolism, apoptosis associated proteins. CONCLUSION： Sub-health fatigue serum can interfere with expression of proteins of HSkMCs.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2009年第11期2095-2100,共6页
Journal of Clinical Rehabilitative Tissue Engineering Research
基金
广东省自然科学基金资助项目(06105115)
广东省中医药管理局基金项目(3060009)~~
