胰岛素信号通路与糖尿病性心肌病发病机理的研究

Insulin/PI 3-kinase/PKB (also known as Akt) signaling pathway plays important roles in regulating multiple cellular processes that are essential for maintaining cardiac function. Down-regulation of this signalling pathway due to insulin resistance contributes to the pathogenesis of diabetic cardiomyopathy. However, it is not well defined how PKB regulates cardiac function in adulthood and how insulin resistance causes diabetic cardiomyopathy. In order to address these questions, we took a proteomics approach and identified a number of potential PKB substrates from mouse heart in response to insulin. Some known PKB substrates such as AS160 and WNK1 were found in our study, which validated our approach. Importantly, we identified a few potential novel PKB substrates including Speg kinase that plays an important role in heart development during embryogenesis. Our unpublished data validated Speg as a novel PKB substrate and its phosphorylation was increased upon insulin stimulation. The expression level of Speg was significantly decreased in the heart of obese mice fed with a high fat diet. Furthermore, deletion of Speg specifically in the heart of adult mice caused dilated cardiomyopathy with impaired cardiac function and eventually led to heart failure. Based on these preliminary data, we put forward our working hypothesis as follows: Speg kinase mediates the regulation of cardiac function downstream of the insulin-PKB signalling pathway, and dysregulation of its expression or phosphorylation contributes to the pathogenesis of diabetic cardiomyopathy. In this proposal, we will address this hypothesis through various Speg mouse models in combination with cell-based systems. In the first part of this proposal, we will study the role of Speg in regulation of cardiac function in adult mouse heart, and investigate the contribution of its impaired expression to the pathogenesis of diabetic cardiomyopathy. In the second part of this proposal, we will elucidate the regulation of Speg by protein phosphorylation, and delineate the in vivo function of PKB-Speg signal nexus and its potential role in the pathogenesis of diabetic cardiomyopathy. This proposed work shall help to deepen our understanding of the molecular mechanisms underlying the pathogenesis of diabetic cardiomyopathy and to identify novel targets for discovery of drugs to combat this disease.

胰岛素对心脏功能的维持起重要作用，胰岛素抵抗可导致糖尿病性心肌病。蛋白激酶B(PKB，又叫Akt)介导胰岛素对心脏功能的调控，但到目前为止，心脏中PKB下游介导胰岛素对心脏功能调控的因子还不完全清楚。申请人团队通过蛋白质组学方法从小鼠心脏中分离了一组可能受PKB调控的蛋白，其中包括已知的PKB底物如AS160和WNK1；同时也发现了一些潜在的PKB新型底物，如Speg蛋白激酶。申请人团队的前期结果表明胰岛素－PKB通路调控Speg的磷酸化；在胰岛素抵抗的肥胖小鼠心脏中Speg表达显著下降，而在成年心脏中诱导性敲除Speg可导致小鼠出现扩心和心衰表型。基于这些前期工作，本申请提出如下假设：Speg蛋白激酶参与介导胰岛素－PKB信号通路对心脏功能的调控；2型糖尿病情况下心肌中Speg表达或调控异常可能是糖尿病性心肌病发病原因之一。本申请拟通过基因工程小鼠模型结合多种生物学手段深入研究这一假设。

糖尿病性心肌病，作为代谢类疾病中的一种，其发生发展与胰岛素抵抗发展密切相关。蛋白激酶B (PKB，又叫Akt) 介导了胰岛素对心脏功能的调控。本项目前期通过蛋白质组学方法从小鼠心脏中分离得到了受胰岛素-PKB调控的新型底物，SPEG蛋白激酶，并发现在胰岛素抵抗的小鼠心脏中Speg表达显著下降。本研究通过对SPEG3A突变小鼠模型和SPEG心脏特异性缺失小鼠模型的分析，结合细胞及蛋白质组学的研究手段，发现胰岛素-PKB可以磷酸化蛋白激酶SPEG，激活SPEG第二个激酶结构域（SK2）的活性。SPEG作为蛋白激酶进而磷酸化SERCA2a、促进SERCA2a的寡聚化，增强其钙转运能力，从而影响心肌细胞的钙稳态，最终调节心脏功能。本项目的实施充分阐述了胰岛素信号通路在糖尿病性心肌病发病过程中的重要分子调控机制，为糖尿病性心肌病的临床诊疗提供了新思路。