Role and Regulation of Skeletal Muscle Mitochondrial Dynamics in Type 2 Diabetes

 描述(申请人提供)：线粒体是孤立的、球形的、产生能量的细胞器的传统观点正在经历一场革命性的变革。新出现的数据表明，线粒体形成了一个动态的网络网络，受到裂变和融合周期的调节。一些调节这些活动的蛋白质的发现导致了 在认识人类疾病方面的重要进展。我们已经证明，动力蛋白相关蛋白1(Drp1)的激活，一种控制线粒体分裂的蛋白质，在糖尿病前期运动后减少，这种减少与胰岛素敏感性增加和脂肪氧化有关。我们现在建议在这项研究的基础上，测试线粒体动力学是2型糖尿病患者胰岛素抵抗的关键机制这一假设。我们的中心假设是，在糖尿病中，线粒体脂代谢升高导致DRP1的招募和激活-可能是通过增加活性氧，导致线粒体碎裂增加和线粒体通透性转换孔打开。在目标1a中，我们将对人体骨骼肌线粒体动力学进行体内和体外研究，研究范围从2型糖尿病患者到肥胖患者，再到瘦削的健康对照组。翻译首例人研究将使用急性脂类挑战(AIM) 1b)和运动训练(Aim 1c)，以探讨骨骼肌线粒体动力学改变对人类胰岛素敏感性的生理意义。胰岛素抵抗将使用正常血糖高胰岛素钳夹进行评估，体内底物代谢将使用间接量热法进行测量。将从肌肉活检组织和通透性肌肉纤维中评估线粒体的分裂/融合、断裂、功能、膜电位、线粒体活性氧物种和脂质中间体的积累。在目标2中，我们将使用抑制和表达克隆实验来直接检测在完整的体外培养的人骨骼肌细胞中操纵线粒体断裂的影响。这项研究将提供对骨骼肌线粒体动力学的全面和补充分析，并将产生关于运动和营养调节之间的联系的新数据，线粒体动力学和功能在2型糖尿病中的作用。这种实验方法利用创新的分子和细胞工具，与具有重要生理意义的人体研究相结合，以获得有关胰岛素抵抗的有意义的数据，并有可能产生洞察力，为子孙后代带来新的糖尿病治疗方法。

 DESCRIPTION (provided by applicant): The traditional view of mitochondria as isolated, spherical, energy producing organelles is undergoing a revolutionary transformation. Emerging data show that mitochondria form a dynamic networked reticulum that is regulated by cycles of fission and fusion. The discovery of a number of proteins that regulate these activities has led to important advances in understanding human disease. We have demonstrated that activation of dynamin related protein 1 (Drp1), a protein that controls mitochondrial fission, is reduced following exercise in prediabetes, and the decrease is linked to increased insulin sensitivity and fat oxidation. We now propose to build on this research and test the hypothesis that mitochondrial dynamics is a key mechanism of insulin resistance in type 2 diabetes. Our central hypothesis is that in diabetes elevated mitochondrial lipid metabolism causes recruitment and activation of Drp1 - likely through increased reactive oxygen species, leading to increased mitochondrial fragmentation and opening of the mitochondrial permeability transition pore. In Aim 1a we will perform in vivo and in vitro studies of human skeletal muscle mitochondrial dynamics across the metabolic phenotype ranging from patients with type 2 diabetes, to obese, to lean healthy controls. Translational first-in-man studies will use an acute lipid challenge (Aim 1b) and exercise training (Aim 1c) to investigate the physiological significance of altered skeleta muscle mitochondrial dynamics on insulin sensitivity in humans. Insulin resistance will be assessed using euglycemic hyperinsulinemic clamps, and in vivo substrate metabolism will be measured using indirect calorimetry. Mitochondrial fission/fusion, fragmentation, function, membrane potential, mitochondrial reactive oxygen species, and the accumulation of lipid intermediates will be assessed from muscle biopsy tissue and permeabilized muscle fibers. In Aim 2, we will use inhibition and expression cloning experiments to directly examine the impact of manipulating mitochondrial fragmentation in intact ex vivo cultured human skeletal muscle cells. This research will provide a comprehensive and complementary analysis of skeletal muscle mitochondrial dynamics, and will also generate novel data on the link between exercise and nutrient regulation of mitochondrial dynamics and function in type 2 diabetes. The experimental approach harnesses innovative molecular and cellular tools, interfaced with physiologically significant human studies to obtain meaningful data on insulin resistance, and has the potential to generate insights that will lead to new diabetes therapies for future generations.