心肌肥厚预适应小鼠线粒体蛋白质组学分析

辛凯悦; 马雷雷; 董震; 马秀瑞; 孙爱军

doi:10.16098/j.issn.0529-1356.2020.03.011

PDF(3560 KB)

解剖学报 ›› 2020, Vol. 51 ›› Issue (3) : 378-384. DOI: 10.16098/j.issn.0529-1356.2020.03.011

心肌肥厚预适应小鼠线粒体蛋白质组学分析

辛凯悦^1,2马雷雷² 董震²马秀瑞² 孙爱军^2*

作者信息 +

Mitochondrial proteomics in myocardial hypertrophic preconditioning mice

XIN Kai-yue^1,2 MA Lei-lei² DONG Zhen² MA Xiu-rui² SUN Ai-jun^2*

Author information +

文章历史 +

摘要

目的使用线粒体蛋白质谱分析方法，探讨心肌肥厚预适应的分子保护机制。方法 14只雄性C57BL6/J小鼠随机分为假手术组（n=6）和心肌肥厚预适应组（n=8），建立小鼠心肌肥厚预适应模型（主动脉弓缩窄3 d、解除缩窄4 d），随机选假手术组3只、心肌肥厚预适应组4只小鼠做蛋白质组学分析，其余小鼠用于形态功能学实验。运用小鼠心脏超声及单个心肌细胞收缩检测技术检测心脏功能学变化，通过心脏常规病理切片及电子显微镜检测心肌组织形态学和线粒体超微结构变化，运用蛋白质组学技术及生物信息学方法，筛选出差异线粒体蛋白，并通过Western blotting验证关键蛋白表达水平变化。结果心肌肥厚预适应组小鼠与假手术组小鼠相比，心脏功能及心肌组织形态学改变无显著性，但透射电子显微镜检测提示，线粒体嵴密度增加，后续利用线粒体蛋白质组学方法共筛选到20个差异表达蛋白（5个上调，15个下调）；随后生物信息学分析显示，差异蛋白主要与线粒体核糖体蛋白有关；关键蛋白的Western blotting验证结果与组学分析结果相一致。结论心肌肥厚预适应激能够增强心肌线粒体功能，这可能与线粒体核糖体蛋白介导的线粒体氧化磷酸化复合体的转录加工、运输过程相关。

Abstract

Objective To investigate the molecular protective mechanisms of myocardial hypertrophic preconditioning by mitochondrial quantitative proteomics. Methods Fourteen C57BL6/J male mice were randomly divided into sham group(n=6) and cardiac hypertrophy preconditioning group(n=8). The murine model of cardiac hypertrophy preconditioning was established by imposing transverse aortic constriction for 3 days and debanding the aorta for 4 days. Three mice from sham group and four mice from cardiac hypertrophy preconditioning group were randomly selected for proteomic analysis, and the remaining mice were used for functional and morphological experiments. The cardiac function was detected by echocardiography, and mechanical properties of cardiomyocytes were assessed using a SoftEdge Myocam. Cardiac morphology and mitochondrial ultrastructure were detected by pathological sections and transmission electron microscopy. The most significant mitochondrial proteins were screened by label-free quantitative proteomics and analyzed by bioinformatics analysis. Western blotting was used to verify the expression changes. Results Compared with the sham group, there were no significant changes in cardiac function and myocardial tissue morphology in the cardiac hypertrophy preconditioning group. However, electron microscopy analyses showed that the density of mitochondrial cristae increased in cardiac hypertrophy preconditioning group. Proteomic analysis screened 20 differentially expressed mitochondrial proteins. Bioinformatics analysis revealed that differentially expressed proteins were mainly related to mitochondrial ribosomal proteins. Western blotting results of key proteins were consistent with proteomic analysis. Conclusion Myocardial hypertrophic preconditioning can promote the energy metabolism of myocardial mitochondria, which may be related to the transcription, processing and transportation of mitochondrial oxidative phosphorylation complex mediated by mitochondrial ribosomal proteins.

导出引用

辛凯悦马雷雷董震马秀瑞孙爱军. 心肌肥厚预适应小鼠线粒体蛋白质组学分析[J]. 解剖学报. 2020, 51(3): 378-384 https://doi.org/10.16098/j.issn.0529-1356.2020.03.011

XIN Kai-yue MA Lei-lei DONG Zhen MA Xiu-rui SUN Ai-jun. Mitochondrial proteomics in myocardial hypertrophic preconditioning mice[J]. Acta Anatomica Sinica. 2020, 51(3): 378-384 https://doi.org/10.16098/j.issn.0529-1356.2020.03.011

中图分类号： R541

参考文献

［1］ Guo ZhK, Wu JF, Shan WH. Expression of collagen Ⅰ and Ⅲ in the hypertrophic myocardial tissue of the rat heart treated by isoproterenol［J］. Acta Anatomica Sinica, 2012, 43(1): 93-97.(in Chinese)

郭志坤，武俊芳，单卫华. 肾上腺素诱导的大鼠肥厚心肌组织中Ⅰ、Ⅲ型胶原蛋白的表达［J］. 解剖学报，2012, 43(1): 93-97.

［2］ Maillet M, Vanberlo JH, Molkentin JD. Molecular basis of physiological heart growth: fundamental concepts and new players ［J］. Nat Rev Mol Cell Biol, 2013, 14(1): 38-48.

［3］ ShimizuⅠ, Minamino T. Physiological and pathological cardiac hypertrophy ［J］. J Mol Cell Cardiol, 2016, 97(2): 45-62.

［4］ Eghbail M, Wang Y, Toro L, et al. Heart hypertrophy during pregnancy: a better functioning heart［J］? Trends Cardiovasc Med, 2006, 16(8): 285-291.

［5］ Kim J, Wende AR, Sena S, et al. Insulin-like growth factor Ⅰ receptor signaling is required for exercise-induced cardiac hypertrophy ［J］. Mol Endocrinol, 2008, 22(11): 2531-2543.

［6］ Wei X, Wu B, Zhao J, et al. Myocardial hypertrophic preconditioning attenuates cardiomyocyte hypertrophy and slows progression to heart failure through upregulation of S100A8/A9 ［J］. Circulation, 2015, 131(17): 1506-1517.

［7］ Lee SH, Yang DK, Choi BY, et al. The transcription factor Eya2 prevents pressure overload-induced adverse cardiac remodeling ［J］. J Mol Cell Cardiol, 2009, 46(4): 596-605.

［8］ Yang DK, Choi BY, Lee YH, et al. Gene profiling during regression of pressure overload-induced cardiac hypertrophy ［J］. Physiol Genomics, 2007, 30(1): 1-7.

［9］ Gopisetty G, Thangarajan R. Mammalian mitochondrial ribosomal small subunit (MRPS) genes: A putative role in human disease ［J］. Gene, 2016, 589(1): 27-35.

［10］ Kolwicz SC, Purohit S, Tian R. Cardiac metabolism and its interactions with contraction, growth, and survival of cardiomyocytes ［J］. Circ Res, 2013, 113(5): 603-616.

［11］ Huang J, Wu J, Wang S, et al. Ultrasound biomicroscopy validation of a murine model of cardiac hypertrophic preconditioning: comparison with a hemodynamic assessment ［J］. Am J Physiol Heart Circ Physiol, 2017, 313(1): 138-148.

［12］ Shen C, Wang C, Fan F, et al. Acetaldehyde dehydrogenase 2 (ALDH2) deficiency exacerbates pressure overload-induced cardiac dysfunction by inhibiting Beclin-1 dependent autophagy pathway ［J］. Biochim Biophys Acta, 2015, 1852(2): 310-318.

［13］ Calvo SE, Mootha VK. The mitochondrial proteome and human disease ［J］. Annu Rev Genomics Hum Genet, 2010, 11: 25-44.

［14］ Mai N, Chrzanowska-Lightowlers ZM, Lightowlers RN. The process of mammalian mitochondrial protein synthesis ［J］. Cell Tissue Res, 2017, 367(1): 5-20.

［15］ Cavdar E, Burkhart W, Blackburn K, et al. The small subunit of the mammalian mitochondrial ribosome. Identification of the full complement of ribosomal proteins present ［J］. J Biol Chem, 2001, 276(22): 19363-19374.

［16］ Rackham O, Busch JD, Matic S, et al. Hierarchical RNA processing is required for mitochondrial ribosome assembly ［J］. Cell Rep, 2016, 16(7): 1874-1890.

［17］ Koc EC, Cimen H, Kumcuoglu B, et al. Identification and characterization of CHCHD1, AURKAIP1, and CRIF1 as new members of the mammalian mitochondrial ribosome ［J］. Front Physiol, 2013, (4): 183-188.

［18］ Kim SJ, Kwon MC, Ryu MJ, et al. CRIF1 is essential for the synthesis and insertion of oxidative phosphorylation polypeptides in the mammalian mitochondrial membrane ［J］. Cell Metab, 2012, 16(2): 274-283.

［19］ Shin J, Lee SH, Kwon MC, et al. Cardiomyocyte specific deletion of Crif1 causes mitochondrial cardiomyopathy in mice ［J］. PLoS One, 2013, 8(1): e53577.

［20］ Fiermonte G, Dolce V, Arrigoni R, et al. Organization and sequence of the gene for the human mitochondrial dicarboxylate carrier: evolution of the carrier family ［J］. Biochem J, 1999, 344(Pt 3): 953-960.

［21］ Yu H, Zhao Z, Yu X, et al. Bovine lipid metabolism related gene GPAM: Molecular characterization, function identification, and association analysis with fat deposition traits ［J］. Gene, 2017, 609: 9-18.