SH2B1在成纤维细胞重编程为心肌样细胞中的作用和机制研究

SH2B1 is widely distributed in multiple organs and is an important protein that regulates transcription and participates in energy metabolism. Recent studies have found that SH2B1 can enhance neurite outgrowth and accelerate the maturation of human induced neurons (iNs)under defined conditions. The effects of SH2B1 on cardiac fibroblasts (CFs) were poorly studied. Our previous studies confirm that SH2B1 participates in the process of myocardial hypertrophy during the development of cardiac hypertrophy. We also observed SH2B1 promotes myogenic determination (MyoD) induced CFs reprogramming into induced cardiomycytes (iCMs). We thus postulated that SH2B1 can promote the transformation of MyoD induced CFs reprogramming into iCMs, and explore its specific effects and possible mechanisms. The rat myocardial infarction (MI) model would be constructed by using SH2B1knock-out and overexpressed transgenic rats. The retrovirus containing Myod would be injected into the periphery of MI zone, changes of MI area and heart function would be detected and formation of iCMs would be monitored. SH2B1-Myod retrovirus woud be constructed and transfecting into CFs, the markers of CFs reprogramming into iCMs would be observed. This study will provide a new perspective for the treatment of heart failure by identifying the role and mechanism of SH2B1 in Myod-induced myocardial reprogramming.

SH2B1分布于多个脏器，是调节转录、参与能量代谢的重要蛋白质。近期研究发现，在MyoD诱导成纤维细胞转化为诱导神经元的过程中，SH2B1可以起到显著的促进作用。SH2B1对心脏成纤维细胞（CFs）的作用尚无研究。我们前期的研究证实在心肌肥厚过程中，SH2B1参与心肌细胞肥大的过程，并初步观察到其对MyoD诱导的CFs转化心肌样细胞（iCMs)有促进作用。我们认为SH2B1可以促进MyoD诱导的CFs转化为iCMs的过程，并探索其具体效应和可能机制。本研究选择SH2B1基因敲除和过表达的转基因大鼠构建心肌梗死（MI）模型，在梗死区周边注射载有MyoD的逆转录病毒，检测MI面积和心功能变化，监测iCMs的形成情况；通过构建SH2B1-MyoD逆转录病毒并转染CFs，检测CFs转化为iCMs的指标。本研究通过探索SH2B1在MyoD诱导心肌重编程中的作用及机制，为心力衰竭治疗提供新视角。

病理性心肌肥厚发生发展过程中表现为高能磷酸盐含量下降、脂肪酸氧化率的降低以及对葡萄糖依赖性的增加，这种现象又称为“能量代谢重编程”。长期的能量代谢重编程最终会导致心肌能量供应效率下降，进一步加重心肌病理性损害。研究表明，同源性2b衔接蛋白1（Src homology 2 binding protein 1, SH2B1）对控制能量平衡尤其是葡萄糖稳态至关重要，本研究通过神经体液因子血管紧张素Ⅱ（Angiotensin Ⅱ，AngⅡ）构建心肌细胞肥大模型，旨在探讨SH2B1通过改变糖酵解通量，加重AngⅡ诱导的病理性心肌肥厚的具体机制。同时在体内实验中探讨SH2B1在心肌肥厚过程中扮演的角色已经对糖代谢的影响。在细胞水平结果表明Ang Ⅱ诱导心肌细胞肥大后SH2B1蛋白相对表达量增高，且GLUT4、G6PD和磷酸化的AMPK蛋白水平上调；另外，与AngⅡ+siNeg组相比，AngⅡ + siSH2B1组心肌肥厚指标即心肌细胞面积、ANP、BNP、Myh7下降，Myh6升高，GLUT4、G6PD下调，且AMPK信号通路被抑制，而过表达SH2B1时，与Ang Ⅱ + AdNC组相比，AngⅡ+AdSH2B1组心肌肥厚指标增加，GLUT4、G6PD和AMPK激活更明显；同时，使用AMPK抑制剂后，糖代谢活动减弱即糖代谢相关蛋白GLUT4、G6PD表达下降。在进一步在动物水平验证结果表明在压力超负荷诱导的心肌肥厚过程中，SH2B1表达和心肌糖代谢均增加， 采用siRNA抑制SH2B1表达后，心肌肥厚程度、糖代谢和PI3K/AKT信号通路的激活均受到抑制。然而，在使用PI3K/AKT抑制剂后，可改善心肌肥厚并且糖代谢通量降低。以上结果证明抑制SH2B1可通过减弱心肌细胞糖代谢而改善心肌肥厚程度，其机制可能与AMPK和PI3K/AKT信号通路激活有关。通过以上研究，我们阐明了SH2B1介导的心肌肥厚分子调控机制，阐明了糖代谢与心肌肥厚之间的关系和可能的机制，从而为心肌肥厚的防治提供了新的实验依据及干预靶点。