Skeletal Muscle Glucose Transport: Exercise and Insulin

描述(由申请人提供)：运动后(PEX)胰岛素敏感性增强，于1982年首次证实，是与改善健康有关的最好的运动益处之一，但其机制一直难以确定。瘦削的正常人的运动对近端的胰岛素信号几乎没有影响，这表明运动改变了远端的位置。研究进展停滞不前，直到在已知对葡萄糖转运(GT)至关重要的最远端信号步骤中发现PEX持续增加：160 kDa的Akt底物(AS160)的磷酸化。在瘦身大鼠中，磷酸化AS160(PAS160)的PEX持续升高与胰岛素诱导的GT升高密切相关，瘦人PEX的肌肉中也发现了持续的PAS160。PAS160升高已成为解释胰岛素敏感性改善PEX的一个有吸引力的候选因素。广泛的、长期的目标是充分阐明增强胰岛素敏感性PEX的机制。1)明确运动后瘦身大鼠骨骼肌AS160磷酸化水平持续升高的机制。2)通过在大鼠骨骼肌中表达防止关键部位磷酸化的AS160，确定AS160‘S在运动诱导的瘦身大鼠GT变化中的作用。3)确定瘦大鼠和肥胖大鼠在各自运动诱导的胰岛素刺激GT改善的具体机制上的相似和不同之处。有证据表明，PAS160中PEX的持续增加并不是由于持续激活的激酶所致。一个有待检验的新想法是，PAS160的持续升高是由于丝氨酸/苏氨酸蛋白磷酸酶减弱了AS160的去磷酸化。由于初步数据表明蛋白磷酸酶2A(PP2A)与AS160去磷酸化有关，因此AIM 1将使用多种方法来探讨PP2A对PAS160的调控。目的2将使用一种创新的方法来测量单个肌肉纤维的GT，这些纤维类型匹配，但表达不同的野生型AS160或AS160基因修饰，以防止关键部位的磷酸化，以了解改善的PEX胰岛素介导的GT是否可归因于更多的pAS160。目标3将确定众所周知的运动对瘦大鼠和肥胖大鼠胰岛素敏感性的益处是否通过不同的机制积累。肥胖大鼠的运动预计会降低肌肉中高水平的胰岛素抵抗介质(诱导型一氧化氮合酶；蛋白质S-亚硝化；脂代谢产物；JNK、IKK和PKC？激活)导致胰岛素信号和GT增强。然而，在瘦身大鼠的肌肉中，运动预计不会显著改变它们本已很低的介体水平或近端胰岛素信号的正常水平。据报道，在瘦人中观察到的PAS160 PEX持续增加在肥胖者中较少。因此，将进行实验，以测试先前发现的瘦小鼠与肥胖大鼠更高的PEX胰岛素介导的GT是否可归因于瘦大鼠更高的PAS160 PEX。这项拟议研究的完成将为改善胰岛素敏感性的具体机制提供新的见解，这对瘦人和肥胖者都是一项主要的健康益处。 公共卫生相关性：这项拟议的研究与公共健康相关，因为骨骼肌对葡萄糖代谢的胰岛素抵抗是2型糖尿病的一个基本且可能是主要的缺陷，而运动可以极大地改善胰岛素介导的葡萄糖摄取。由于肥胖、胰岛素抵抗和糖尿病的相关流行，阐明胰岛素抵抗个体改善胰岛素敏感性的机制至关重要。了解正常个体胰岛素敏感性改善的机制也很重要，因为久坐不动的瘦弱个体的胰岛素敏感性水平可能不会带来最佳健康，而且识别健康个体的机制有可能激励新疗法的开发，使胰岛素抵抗者受益。

DESCRIPTION (provided by applicant): Increased post-exercise (PEX) insulin sensitivity, first demonstrated in 1982, is among the best documented exercise benefits linked to improved health, but identification of the mechanism has been elusive. Exercise by lean, normal individuals has little effect on proximal insulin signaling, suggesting exercise alters a distal site. Progress stalled until the discovery of a sustained PEX increase in the most distal signaling step known to be crucial for glucose transport (GT): phosphorylation of Akt Substrate of 160 kDa (AS160). The sustained PEX increase in phosphorylated AS160 (pAS160) tracks closely with elevated insulin-induced GT in lean rats, and sustained pAS160 is also found in muscle of lean humans PEX. Increased pAS160 has emerged as an attractive candidate to explain improved insulin sensitivity PEX. The broad, long-term goal is to fully elucidate the mechanisms that underlie enhanced insulin sensitivity PEX. The 3 Specific Aims are: 1) Identify the mechanism for the sustained increase in AS160 phosphorylation in skeletal muscle of lean rats after exercise. 2) By expressing in rat skeletal muscle AS160 that is genetically modified to prevent phosphorylation on key sites, determine AS160's role in exercise-induced changes in GT in lean rats. 3) Identify the similarities and differences between lean and obese rats in the specific mechanisms for their respective exercise-induced improvements in insulin-stimulated GT. Evidence indicates the sustained PEX increase in pAS160 is not attributable to persistent activation of kinases. A novel idea to be tested is that the sustained elevation in pAS160 is because of attenuated AS160 dephosphorylation by Ser/Thr protein phosphatases. Because preliminary data implicate protein phosphatase 2A (PP2A) in AS160 dephosphorylation, multiple approaches will be used for Aim 1 to probe PP2A's regulation of pAS160. Aim 2 will use an innovative method to measure GT by single muscle fibers that are matched for fiber type, but differ by expression of wildtype AS160 or AS160 genetically modified to prevent phosphorylation on key sites to learn if the improved PEX insulin-mediated GT in lean rats is attributable to greater pAS160. Aim 3 will determine if the well-known exercise benefits on insulin sensitivity for lean and obese rats are accrued by distinct mechanisms. Exercise by obese rats is predicted to reduce high muscle levels of mediators of insulin resistance (inducible nitric oxide synthase; protein S- nitrosylation; lipid metabolites; JNK, IKK & PKC? activation) leading to enhanced insulin signaling and GT. However, in muscles from lean rats, exercise is predicted to not greatly alter their already low levels of mediators or normal levels of proximal insulin signaling. The sustained increase in pAS160 PEX observed in lean humans was reported to be less in obese people. Accordingly, experiments will be performed to test if the greater PEX insulin-mediated GT previously found for lean vs. obese rats is attributable to greater pAS160 PEX in lean rats. Completion of the proposed research will provide novel insights into the specific mechanisms that account for improved insulin sensitivity, a major health benefit for both lean and obese individuals. PUBLIC HEALTH RELEVANCE: The proposed research is relevant to public health because insulin resistance for glucose disposal by skeletal muscle is an essential and perhaps primary defect for Type 2 diabetes, and exercise can greatly improve insulin-mediated glucose uptake. Because of the related epidemics of obesity, insulin resistance and diabetes, it is crucial to elucidate mechanisms for improved insulin sensitivity in insulin resistant individuals. It is also important to understand mechanisms for improved insulin sensitivity in normal individuals because the level of insulin sensitivity found in sedentary lean individuals may not confer optimal health and because identifying mechanisms in healthy individuals has the potential to inspire the development of new therapies that benefit insulin-resistant people.