Circadian Regulation of Myocardial Insulin Signaling

描述(申请人提供)：尽管临床治疗有所改进，心血管疾病(CVD)仍然是美国人死亡的主要原因。与多种常见疾病一样，心血管疾病是复杂的基因-环境相互作用的产物，其中遗传信息本质上影响个体对环境刺激/应激的反应。我们最近强调，心肌细胞昼夜节律时钟是一种细胞自主的分子机制，它促进心脏对各种刺激/应激(如肾上腺素、脂肪酸、促肥大刺激)的时间性适当反应。通过遗传(例如，时钟成分基因的多态性)或环境(例如，轮班工作、睡眠和饮食行为调节)手段破坏生物钟机制，与人类心血管疾病风险的增加有关。最近，我们观察到在心肌细胞限制的昼夜节律时钟转录因子BMAL1(称为CBK小鼠)缺失后，小鼠发生扩张型心肌病(和寿命缩短)。转录组和生物信息学方法(在年轻小鼠中，在心脏病理之前)确定了9个假定的直接BMAL1靶基因。随后的验证研究证实，BMAL1直接与Pik3r1(PI3K的P85调节亚单位)启动子的多个E-box结合，导致对照心脏(而不是CBK心脏)这一胰岛素信号成分的mRNA和蛋白质水平随时间而波动。我们的初步研究还表明，心肌细胞生物钟中断后，心肌胰岛素信号转导受损，在Zucker糖尿病肥胖大鼠心脏(肥胖症和2型糖尿病模型)观察到的生物钟功能障碍通过依赖时间的限制喂养部分恢复正常。总之，这些观察结果使我们假设，心肌细胞的生物钟以一天中的时间依赖的方式调节心肌的胰岛素敏感性(通过调节P85？)，而饮食诱导肥胖后的时钟功能障碍扰乱了心肌胰岛素信号，从而导致收缩功能障碍。以下具体目标将检验这一假说：1)确定心肌细胞生物钟是否以一天中的时间依赖的方式调节心肌胰岛素信号和关键的胰岛素介导过程(例如，新陈代谢、自噬)；2)通过测试BMAL1缺陷心脏的功能障碍是继发于P85降低的假说，确定BMAL1缺陷心脏的心肌病机制；以及3)确定心肌细胞生物钟正常化是否将缓解胰岛素抵抗(即饮食诱导的肥胖)小鼠的心脏收缩功能障碍。这些研究的成功完成将可能确定心肌细胞生物钟是调节心肌胰岛素敏感性的一种新的内在机制，并为未来的翻译研究提供基础 以心肌细胞生物钟为靶点，预防和/或治疗肥胖/糖尿病心肌病。

DESCRIPTION (provided by applicant): Despite improvements in clinical treatments, cardiovascular disease (CVD) remains the primary cause of mortality in the United States. CVDs, as with multiple common diseases, are the product of a complex gene- environment interaction, wherein genetic information intrinsically influences the responsiveness of an individual to environmental stimuli/stresses. We have recently highlighted the cardiomyocyte circadian clock as a cell autonomous molecular mechanism that facilitates temporally-appropriate cardiac responses to various stimuli/stresses (e.g., epinephrine, fatty acids, pro-hypertrophic stimuli). Disruption of the circadian clock mechanism, through either genetic (e.g., polymorphisms in clock component genes) or environmental (e.g., shift work, sleep and eating behavior modulation) means, is associated with increased CVD risk in humans. Recently, we have observed development of dilated cardiomyopathy (and reduced lifespan) in mice following cardiomyocyte-restricted deletion of the circadian clock transcription factor BMAL1 (termed CBK mice). Transcriptome and bioinformatic approaches (in young mice, prior to cardiac pathology) identified 9 putative direct BMAL1 target genes. Subsequent validation studies confirmed that BMAL1 directly binds to multiple E- boxes in the Pik3r1 (p85¿ regulatory subunit of PI3K) promoter, resulting in time-of-day-dependent oscillations in mRNA and protein levels of this insulin signaling component in control, but not CBK, hearts. Our preliminary studies also suggest impaired myocardial insulin signaling following cardiomyocyte circadian clock disruption, and that circadian clock dysfunction observed in Zucker Diabetic Fatty rat hearts (an obesity and type 2 diabetes model) is partially normalized through time-of-day-dependent restricted feeding. Collectively, these observations have led us to hypothesize that the cardiomyocyte circadian clock modulates myocardial insulin sensitivity in a time-of-day-dependent manner (through regulation of p85¿), and that dysfunction of the clock following diet-induced obesity disrupts myocardial insulin signaling, thereby contributing to contractile dysfunction. The following specific aims will test this hypothesis: 1) Determine whether the cardiomyocyte circadian clock modulates myocardial insulin signaling and critical insulin- mediated processes (e.g., metabolism, autophagy) in a time-of-day-dependent manner; 2) Determine the mechanism for cardiomyopathy in BMAL1 deficient hearts by testing the hypothesis that dysfunction is secondary to decreased p85¿; and 3) Determine if normalization of the cardiomyocyte circadian clock will attenuate cardiac contractile dysfunction in a mouse model of insulin resistance (i.e., diet-induced obesity). Successful completion of the proposed studies will likely identify the cardiomyocyte circadian clock as a novel intrinsic mechanism that modulates myocardial insulin sensitivity, and provide a foundation for future translational studies targeting the cardiomyocyte circadian clock for obesity/diabetic cardiomyopathy prevention and/or treatment.