Proteome Mapping of Heart Failure in Aging

老年心力衰竭的蛋白质组图谱

• 批准号: 6780070
• 负责人: MARVIN O BOLUYT
• 金额: $ 7.18万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-15 至 2006-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6780070
• 关键词: age difference; aging; aldosterone; animal old age; corticosteroid receptors; electrospray ionization mass spectrometry; fibrosis; heart failure; immunoprecipitation; laboratory rat; microarray technology; posttranslational modifications; protein quantitation /detection; protein structure function; proteomics; two dimensional gel electrophoresis; western blottings

Heart failure is a major health problem in the elderly. Evidence indicates that a cardiac aldosterone signaling system plays an important role in the fibrosis associated with heart failure. Spironolactone, an aldosterone receptor antagonist, counteracts some of the effects of aldosterone in culture and in vivo. Aldosterone exerts its effects in target cells by binding to the mineralocorticoid receptor (MR) and stimulating transcription of effector genes. According to this paradigm, increases in fibrosis can be explained by the action of the aldosterone-MR complex on cardiac fibroblast genes that encode extracellular matrix proteins. A number of observations are not explained by this paradigm, however, including our preliminary data suggesting that spironolactone treatment of old rats does not decrease fibrosis as predicted, but increases the collagen content of the heart. Another paradoxical finding is that conditonal suppression of MR expression in cardiac myocytes of transgenic mice leads to fibrosis and heart failure. Emerging evidence strongly suggests that rapid non-genomic effects of aldosterone may explain some of the confusion, but the mechanisms by which these non-genomic effects are enacted have been elusive. The specific aims of this proposal are to identify novel protein mediators of aldosterone action that may regulate the age-associated increases in fibrosis and to identify novel proteins that interact with aldosterone and/or the MR to account for the non-genomic effects of aldosterone. We will study existing heart tissue samples from young and old rats treated chronically with vehicle or spironolactone. We will also generate cardiac myocyte and cardiac fibroblast cell populations from isolated hearts of old rats perfused briefly with vehicle or aldosterone. Proteins from cytosolic and particulate fractions of each cell type will be prepared for proteomic analyses. 2-D gel electrophoresis will be employed to identify differentially expressed or posttranslationally modified proteins. We will target proteins involved in the aldosterone signaling system by immunoprecipitation of MR-associated proteins. These studies are expected to identify novel proteins that may mediate the effects of aldosterone in the aged failing heart. Identification of proteins involved in aldosterone signaling will lead to the development of new hypotheses for future grant proposals addressing their roles in the development of heart failure in the elderly.

心力衰竭是老年人的主要健康问题。证据表明，心脏醛固酮信号系统在与心力衰竭相关的纤维化中起着重要作用。醛固酮受体拮抗剂螺内酯会抵消醛固酮在培养物和体内的某些作用。醛固酮通过与盐皮质激素受体（MR）结合并刺激效应基因转录来施加其作用。根据该范式，纤维化的增加可以通过醛固酮-MR复合物对编码细胞外基质蛋白的心脏成纤维细胞基因的作用来解释。但是，此范式没有解释许多观察结果，包括我们的初步数据表明 螺内酯治疗老鼠不会像预测的那样降低纤维化，而是增加心脏的胶原蛋白含量。另一个自相矛盾的发现是，转基因小鼠心脏肌细胞中MR表达的孔子抑制会导致纤维化和心力衰竭。新兴的证据强烈表明，醛固酮的快速非基因组效应可能解释了某些混乱，但是这些非基因组效应被制定的机制是难以捉摸的。该提案的具体目的是确定醛固酮作用的新型蛋白质介体，这些促醛固酮作用可能调节纤维化的年龄相关增加，并鉴定与醛固酮和/或MR相互作用的新型蛋白质，以解释醛固酮的非基因组作用。我们将研究用载体或螺内酯长期治疗的年轻大鼠和老鼠的现有心脏组织样品。我们还将从孤立的老鼠的孤立心脏中产生心肌细胞和心脏成纤维细胞群，并用媒介物或醛固酮短暂灌注。每种细胞类型的胞质和颗粒分数的蛋白质将准备用于蛋白质组学分析。 2-D凝胶电泳将用于鉴定差异表达或翻译后修饰的蛋白质。我们将靶向蛋白质 通过免疫沉淀MR相关蛋白参与醛固酮信号系统。这些研究有望鉴定出新的蛋白质，这些蛋白质可能介导醛固酮在老年失败心脏中的作用。鉴定参与醛固酮信号传导的蛋白质将导致发展新的假设，以解决未来的赠款建议，以解决其在老年人心力衰竭发展中的作用。