心脏表达CORIN基因多态性T555I/Q568P的转基因小鼠心肌肥厚的分子机制研究

Cardiac hypertrophy is a compensatory response to biomechanical and/or pathological stresses such as hypertension, myocardial injury and neurohumoral activation. Its major features include increased cardiomyocyte size and protein synthesis. Pathologically, cardiac hypertrophy increases the risk of arrhythmia and often leads to heart failure and sudden death. Corin is a cardiac protease which converts pro-ANP/pro-BNP to biologically active ANP/BNP. ANP can regulate cardiomyocyte size by acting as an anti-hypertrophic factor. In mice, deficiency in either ANP or its receptor NPRA causes cardiac hypertrophy. Corin null mice also develop hypertension and cardiac hypertrophy. Recently, two non-synonymous single nucleotide polymorphisms (SNPs) (T555I and Q568P) in the human CORIN gene have been identified in epidemiological studies. The minor I555/P568 allele is associated with hypertension and cardiac hypertrophy. To functionally characterize the CORIN variant T555I/Q568P, we generated transgenic mice epressing wild-type (WT) or T555I/Q568P variant corin in the heart under the control of α-myosin heavy chain promoter. The mice were crossed into a corin knockout (KO) background to create KO/TgWT and KO/TgV mice that expressed WT or variant corin, respectively, in the heart. Functional studies showed that KO/TgV mice had significantly higher levels of pro-ANP the heart compared with that in control KO/TgWT mice, indicating that the corin variant was defective in processing natriuretic peptides in vivo. By radiotelemetry, corin KO/TgV mice were found to have hypertension that was sensitive to dietary salt loading. The mice also developed cardiac hypertrophy at 12 to 14 months of age when fed a normal salt diet or at a younger age when fed a high-salt diet. The phenotype of salt-sensitive hypertension and cardiac hypertrophy in KO/TgV mice closely resembles the pathological findings in patients who carry the corin variant. However, the molecular mechanism underlying cardiac hypertrophy in the KO/TgV mice remains to be determined. In this proposal, we will extend our in vivo studies and further explore the possible pressure-independent mechanism for cardiac hypertrophy in the CORIN transgenic mice.

心肌肥厚是心血管疾病发生及致死的独立预测因子，也是心力衰竭发生的重要中间环节，其发生机制尚未完全阐明。丝氨酸蛋白corin是生成活性心钠肽/脑钠肽的关键酶。目前研究显示心脏组织中corin-ANP/BNP系统功能缺陷可能介导心肌肥厚的发生。人群中存在的CORIN基因多态性T555I/Q568P与心肌肥厚发生具有相关性，但具体机制未明。在我们前期制备的心脏特异性表达突变型CORIN I555/P568的转基因小鼠中，发生了显著的心肌肥厚，同时伴有ANP/BNP活化障碍。据此，本研究将在大量前期工作基础上，通过主动脉弓结扎术造模及降压药物平衡血压影响，从整体水平探讨突变型CORIN I555/P568介导心肌肥厚发生的非血压依赖性机制。研究结果将可能为心衰的发生发展提供新的理论解释，并有望为临床治疗提供新的药物靶点。

心肌肥厚是心血管疾病发生及致死的独立预测因子，也是心力衰竭发生的重要中间环节，其发生机制尚未完全阐明。丝氨酸蛋白corin是生成活性心钠肽/脑钠肽的关键酶。目前研究显示心脏组织中corin-ANP/BNP系统功能缺陷可能介导心肌肥厚的发生。人群中存在的CORIN基因多态性T555I/Q568P与心肌肥厚发生具有相关性，但具体机制未明。本研究中，我们首先于体外培养的心肌细胞水平探讨CORIN I555/P568介导心肌肥厚发生的具体分子机制。研究发现与表达野生型CORIN T555/Q568相比，表达突变型CORIN I555/P568心肌细胞中 ,自噬相关蛋白LCII/I、PINK1、parkin和Bnip3蛋白表达水平降低，mTOR磷酸化水平升高，线粒体ATP合成活性降低及活性氧ROS生成增加，提示CORIN突变型可能影响心肌细胞线粒体自噬过程，从而介导心肌肥厚发生。在体动物实验研究发现，TAC心肌肥厚小鼠模型中，加入3-MA抑制心肌细胞线粒体自噬过程，Corin/ANP的mRNA表达水平及蛋白表达水平均明显增高，提示Corin-ANP通路可能影响心肌细胞线粒体自噬过程从而介导心肌肥厚发生。另外我们一部分与心肌肥厚机制相关的探索性实验发现，在TAC心肌肥厚小鼠模型心肌细胞中miR-296-5p表达明显升高，降低 miR-296-5p表达能够显著减轻TAC小鼠心肌肥厚的程度，研究还发现miR-296-5p可能通过靶向作用于CACNG6基因发挥其促心肌肥厚作用。我们目前的研究发现对于从不同角度理解心肌肥厚的发生机制具有积极科学意义，如前述心肌肥厚是心力衰竭发生的重要中间环节，心力衰竭作为多种心血管疾病发展的终末阶段，目前临床治疗仍然十分棘手，主要以对症治疗为主。深入理解心肌肥厚发生的相关分子机制，对于探寻药物靶点有重要意义，我们的研究具有潜在的临床价值及应用前景，有望以心肌细胞膜蛋白Corin及ANP/BNP通路为靶点，干预心肌肥厚及心衰发生发展过程。为临床治疗这一类疾病提供更多选择。