Circadian Regulation of Myocardial Insulin Signaling

描述（由申请人提供）：尽管临床治疗有所改善，心血管疾病（CVD）仍然是美国死亡的主要原因。心血管疾病与多种常见疾病一样，是复杂的基因-环境相互作用的产物，其中遗传信息内在地影响个体对环境刺激/压力的反应。我们最近强调了心肌细胞生物钟作为一种细胞自主分子机制，促进心脏对各种刺激/压力（如肾上腺素、脂肪酸、促肥厚刺激）的临时适当反应。通过遗传（如时钟组成基因的多态性）或环境（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增加有关。最近，我们观察到在小鼠（CBK小鼠）心肌细胞限制性缺失昼夜节律时钟转录因子BMAL1后，扩张型心肌病（和寿命缩短）的发展。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直接结合Pik3r1 （PI3K的p85调控亚基）启动子中的多个E- box，导致控制心脏中这种胰岛素信号成分的mRNA和蛋白质水平的时间依赖性振荡，而不是CBK。我们的初步研究还表明心肌细胞生物钟紊乱后心肌胰岛素信号受损，并且在Zucker糖尿病脂肪大鼠心脏（肥胖和2型糖尿病模型）中观察到的生物钟功能障碍通过依赖于时间的限制性喂养部分正常化。总的来说，这些观察结果使我们假设心肌细胞生物钟以一种依赖于时间的方式调节心肌胰岛素敏感性（通过调节p85¿），并且饮食诱导的肥胖引起的生物钟功能障碍会破坏心肌胰岛素信号，从而导致收缩功能障碍。以下具体目标将检验这一假设：1)确定心肌细胞生物钟是否以时间依赖的方式调节心肌胰岛素信号传导和关键胰岛素介导的过程（如代谢、自噬）；2)通过验证功能障碍继发于p85¿降低的假设，确定BMAL1缺陷心脏的心肌病机制；3)确定心肌细胞生物钟的正常化是否会减轻胰岛素抵抗小鼠模型（即饮食性肥胖）的心肌收缩功能障碍。这些研究的成功完成可能会确定心肌细胞生物钟作为调节心肌胰岛素敏感性的一种新的内在机制，并为未来的转化研究提供基础

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.