Impact of Inflammation on the Control of Muscle Glucose Uptake

描述（由申请人提供）：即使没有糖尿病史，高血糖在脓毒症患者中也很常见。骨骼肌葡萄糖摄取（MGU）的胰岛素抵抗是这种高血糖症的主要原因。MGU的控制分布在葡萄糖输送到肌肉、葡萄糖转运到肌肉和肌肉内葡萄糖磷酸化之间；胰岛素抵抗是由于其中一个或多个存在缺陷。许多研究小组已经研究了肥胖对MGU的影响，但炎症对MGU控制分布的影响尚不清楚。本实验将研究脂多糖（LPS）诱导的炎症在多大程度上重新分配对MGU的控制。在本提案中，转运和磷酸化在控制MGU中的作用将通过使用种系操作（部分敲除或过表达）转运和磷酸化能力（例如己糖激酶）来调节单个步骤或多个步骤并测量对MGU的影响来评估。我们假设葡萄糖磷酸化能力的缺陷在炎症诱导的胰岛素抵抗中起核心作用。实验将在长期插管、有意识的小鼠中进行。这种方法允许在没有压力的情况下对体内MGU进行全面的代谢评估。实验策略是干扰参与控制MGU的蛋白质或过程，并测量扰动对葡萄糖内流的影响。全身葡萄糖摄取和MGU将分别使用[3-3H]葡萄糖和[14C] 2-脱氧葡萄糖，结合血液和组织取样和测量肌肉血流量的方法进行测量。MGU与长链脂肪酸（LCFA）摄取的关系将同时使用放射性标记脂肪酸类似物进行测量。肌肉ATP通量将使用31NMR波谱进行评估。组织将分析糖原合成、胰岛素信号、氧化应激和GLUT4易位。我们的具体目标是确定：1；脂多糖对相对控制转运和葡萄糖磷酸化的影响决定了MGU 2。如果LPS放大了NEFA和葡萄糖可用性在调节MGU 3中的影响。我们的长期目标是确定受炎症影响的控制MGU的步骤，并评估哪些步骤对氧化应激的变化更敏感。在脓毒症等炎症应激期间，未来的治疗方法可以更有针对性地纠正MGU。公共卫生相关性：高血糖在住院患者中非常常见，临床试验表明，如果高血糖可以最小化，发病率和死亡率都可以改善。高血糖的一个主要原因是骨骼肌葡萄糖摄取的胰岛素抵抗，该建议将确定缺陷在哪里，并解决以下问题：是由于胰岛素激活信号通路失败，还是由于炎症加剧和限制胰岛素发挥有益作用的能力而导致的线粒体潜在缺陷。

DESCRIPTION (provided by applicant): Hyperglycemia is very common in patients with sepsis even if there is no history of diabetes. Insulin resistance of skeletal muscle glucose uptake (MGU) is a major cause of this hyperglycemia. Control of MGU is distributed between delivery of glucose to muscle, glucose transport into muscle, and glucose phosphorylation within muscle; insulin resistance is due to defects in one or more of them. The impact of obesity on MGU has been studied by a number of groups, but the impact of inflammation on the distribution of control of MGU is unknown. The experiments described in this proposal will examine the extent to which inflammation induced by lipopolysaccharide (LPS) redistributes the control of MGU. In this proposal the roles transport and phosphorylation play in controlling MGU will be assessed by using germline manipulation (partial knockout or over expression) of transport and phosphorylation capacity (e.g. hexokinase) to modulate a single step or multiple steps and measure the impact on MGU. We hypothesize that defects in glucose phosphorylation capacity play a central role in the inflammation induced insulin resistance. Experiments will be performed in chronically catheterized, conscious mice. This approach allows for comprehensive metabolic assessment of MGU in vivo in the absence of stress. The experimental strategy is to perturb proteins or processes involved in control of MGU and measure the effect of the perturbation on glucose influx. Whole body glucose uptake and MGU will be measured using [3-3H] glucose and [14C] 2- deoxyglucose, respectively, in combination with methods for sampling blood and tissues and measuring muscle blood flow. The relationship of MGU to long chain fatty acid (LCFA) uptake will simultaneously be measured using a radiolabeled fatty acid analog. Muscle ATP flux will be assessed using 31NMR spectroscopy. Tissues will be analyzed for glycogen synthesis, insulin signaling, oxidative stress and GLUT4 translocation. Our specific aims are to determine: 1. The impact of LPS on the relative control transport and glucose phosphorylation have in determining MGU 2. If LPS amplifies the impact NEFA and glucose availability have in modulating MGU 3. If modulating oxidative stress (NO availability and NF-:B activation) following LPS will improve MGU by augmenting glucose phosphorylation and mitochondrial ATP flux Our long term goal is to identify the steps controlling MGU that are impacted by inflammation and assess which of those steps are more responsive to changes in oxidative stress. Future therapies can then have a more targeted approach in correcting MGU during an inflammatory stress such as sepsis. PUBLIC HEALTH RELEVANCE: Hyperglycemia is very common in hospitalized patients and clinical trials suggest if the hyperglycemia can be minimized morbidity and mortality are improved. A major cause of the hyperglycemia is insulin resistance of skeletal muscle glucose uptake This proposal will determine where the defect is and address the questions is it due simply to a failure of insulin to activate it signaling pathway or are underlying defects in the mitochondria caused by the inflammation aggravating and limiting the ability of insulin to exert is beneficial effects.