MITOCHONDRIAL DYSFUNCTION IN DIABETIC CARDIOMYOPATHY

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 糖尿病性心肌病是糖尿病患者死亡的主要原因。从机制上讲，线粒体功能的丧失可能在疾病的进展中起作用，这是由于生物能缺陷和自由基产生增加。 糖尿病引起的代谢状况可进一步加剧氧化应激。 例如，升高的游离脂肪酸通过非脂肪酸碳源抑制糖酵解和线粒体氧化磷酸化（Randle循环）。在高血糖条件下，脂肪酸的恒定、严格利用可促进持续增加的自由基产生，因为脂肪酸氧化相对于其他可氧化底物产生更多的自由基。 重要的是，Randle循环可能会被cAMP依赖性蛋白激酶（PKA）的激活所破坏，这表明缓解代谢应激的内源性手段可能会受到损害。 使用1型糖尿病的遗传模型（OVE 26小鼠），我们已经开始研究该疾病对心脏线粒体和PKA信号传导的纵向影响。 我们的研究结果表明，从OVE 26小鼠的心脏线粒体对脂肪酸氧化的严格依赖，以支持能源的生产。重要的是，我们还发现OVE 26小鼠心脏中PKA蛋白水平和活性降低。基于这些观察，我们假设：在高血糖条件下，线粒体对脂肪酸氧化的依赖增加了氧化应激。 对脂肪酸作为能量来源的依赖由于氧化失活和PKA信号传导的损失而加剧。 长期的氧化应激，加上PKA活性的丧失，有助于糖尿病心肌病。 将使用1型糖尿病的遗传模型（OVE 26小鼠）、细胞培养和蛋白质生物化学通过以下特定目的来检验该假设。 目标1.机械地确定随着糖尿病性心肌病的进展而发生的线粒体功能的改变。 假设与高血糖症相关的氧化应激增加部分是脂肪酸的刚性利用以及伴随的呼吸活性降低的功能。 将使用MRI进行生化研究、蛋白质组学分析和心脏结构/功能分析，以检验这一假设。 目标2.定义高血糖导致PKA活性和含量不足的分子机制。我们推测，增加氧化应激促进PKA氧化，失活和降解。 PKA活性的这种丧失通过持续和严格利用脂肪酸氧化而导致进一步的氧化应激。目标3.本研究的目的是确定PKA信号传导减少、线粒体功能减弱、氧化应激增加和糖尿病心肌病之间的因果关系。 将进行研究以确定胰岛素补充或PKA活化如何恢复代谢柔韧性。 假设维持PKA信号传导将维持线粒体功能并减弱糖尿病心肌病的进展。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Diabetic cardiomyopathy is a leading cause of death among diabetics. Mechanistically, loss of mitochondrial function likely plays a role in the progression of the disease due to bioenergetic deficits and increased free radical production. Oxidative stress can be further exacerbated by metabolic conditions that result from diabetes. For example, elevated free fatty acids inhibit glycolysis and mitochondrial oxidative phosphorylation by non fatty acid carbon sources (Randle cycle). A constant, rigid, utilization of fatty acids may promote sustained increased free radical production during hyperglycemic conditions because fatty acid oxidation generates more free radicals relative to other oxidizable substrates. Importantly, the Randle cycle may be disrupted by activation of cAMP-dependent protein kinase (PKA), suggesting endogenous means of alleviating metabolic stress that are present may be compromised. Using a genetic model of type 1 diabetes (OVE26 mice), we have begun examining the longitudinal effects of the disease on cardiac mitochondria and PKA signaling. Our results indicate a rigid reliance of cardiac mitochondria from OVE26 mice on fatty acid oxidation for supporting energy production. Importantly, we have also found that PKA protein levels and activity are decreased in hearts of OVE26 mice. Based on these observations we hypothesize: Under hyperglycemic conditions, mitochondrial reliance on fatty acid oxidation increases oxidative stress. Reliance on fatty acids as an energy source is exacerbated by oxidative inactivation and loss of PKA signaling. Prolonged oxidative stress, coupled with loss of PKA activity, contributes to diabetic cardiomyopathy. This hypothesis will be tested using a genetic model of type 1 diabetes (OVE26 mice), cell culture, and protein biochemistry by the following specific aims. Aim 1. Mechanistically determine the alterations in mitochondrial function that occur with the progression of diabetic cardiomyopathy. The hypothesis is that increased oxidative stress associated with hyperglycemia is a function, in part, of rigid utilization of fatty acids with concomitant decreases respiratory activity. Biochemical studies, proteomic analysis, and cardiac structure/function analysis using MRI will be performed to test this hypothesis. Aim 2. Define the molecular mechanisms that lead to deficits in PKA activity and content as induced by hyperglycemia. We hypothesize that the increased oxidative stress promotes PKA oxidation, inactivation, and degradation. This loss of PKA activity then contributes to further oxidative stress via sustained and rigid utilization of fatty acid oxidation. Aim 3. The goal of this aim is to define the causal link between decreased PKA signaling, diminished mitochondrial function, increased oxidative stress, and diabetic cardiomyopathy. Studies will be performed to determine how insulin supplementation or activation of PKA restores metabolic pliability. It is hypothesized that sustaining PKA signaling will sustain mitochondrial function and attenuate the progression of diabetic cardiomyopathy.