Mechanisms underlying metabolic syndrome in obesity

描述(申请人提供)：肥胖是造成胰岛素抵抗和代谢综合征的最常见和最强大的力量，然而，这种联系的分子基础还不是很清楚。在这项提案中，三名独立资助的研究人员--临床研究员菲利普·科恩医学博士和分别在肌肉和脂肪细胞生物学方面拥有丰富经验的夏洛特·彼得森博士和罗伯特·麦吉希博士--将把合作努力正式化，作为肥胖、胰岛素抵抗和组织脂质积累领域先前工作和共同兴趣的自然延伸。我们的总体假设是，人类的胰岛素抵抗很大程度上源于肥胖发展过程中心肌细胞内脂质(IMCL)的异位积累。此外，我们假设IMCL的过度积累依赖于脂肪组织中脂肪细胞和巨噬细胞之间复杂的相互作用所产生的分泌蛋白。为了验证这些假设，我们将检验分离和培养的脂肪细胞、巨噬细胞和肌肉细胞之间的相互作用，这些细胞来自中度肥胖伴胰岛素抵抗和糖耐量受损(IGT)的受试者，但他们还没有空腹高血糖。该研究人群的IMCL升高，是肥胖并发症的高危人群，但避免了糖毒性的病理生理并发症。这些受试者将与糖耐量正常(非)的中度肥胖受试者进行比较。目标1将探讨IMCL的发病机制，并阐明其在IGT发生发展中的作用。培养的肌肉细胞将被用来确定患有IGT和NGT的肥胖患者在不同的细胞外脂肪酸浓度下，肌肉基因表达和代谢活动是否存在内在差异。将评估脂肪堆积和氧化，以及胰岛素介导的糖原合成和信号转导。目的2将确定IMCL的积聚是否依赖于脂肪组织分泌蛋白。我们将使用脂肪细胞、成肌细胞和脂肪基质血管细胞的共培养来检测IMCL和胰岛素抵抗的发展。目的3确定IGT患者的间质部分在与肌肉细胞共培养时是否比NGT患者的间质部分更有效地促进IMCL。我们将比较间质血管部分在单核/巨噬细胞聚集和细胞因子表达方面的差异。目的4将确定吡格列酮治疗10周后糖耐量的改善是否导致IMCL减少，并确定相关的细胞机制。在吡格列酮治疗前后，肌肉和基质细胞的共培养研究也将被使用。这些实验将为肥胖和导致代谢综合征的肌肉功能之间的联系提供机械性的见解。

DESCRIPTION (provided by applicant): Obesity is the most common and powerful force for creating insulin resistance and metabolic syndrome, however, the molecular basis of this association is not well understood. In this proposal, three independently funded researchers-Philip Kern, MD a clinical investigator, and Charlotte Peterson, PhD and Robert McGehee, PhD, with significant experience in muscle and adipocyte biology, respectively- will formalize a collaborative effort as a natural extension of previous work and shared interests in the fields of obesity, insulin resistance, and tissue lipid accumulation. Our overall hypothesis is that insulin resistance in humans stems largely from ectopic accumulation of intramyocellular lipid (IMCL) during the development of obesity. Further, we hypothesize that excess IMCL accumulation is dependent on secretary proteins derived from a complex interplay between adipocytes and macrophages in adipose tissue. To test these hypotheses, we will examine the interactions among adipocytes, macrophages, and muscle cells isolated and cultured from subjects that are moderately obese with insulin resistance and impaired glucose tolerance (IGT), but who do not yet have fasting hyperglycemia. This study population has elevated IMCL and is at high risk for obesity complications, but avoids the pathophysiologic complications of glucotoxicity. These subjects will be compared to moderately obese subjects with normal glucose tolerance (NOT). Aim 1 will explore mechanisms that contribute to IMCL and elucidate its role in the development of IGT. Cultured muscle cells will be used to determine whether obese subjects with IGT versus NGT demonstrate intrinsic differences in muscle gene expression and metabolic activity under differing extracellular fatty acid concentrations. Lipid accumulation and oxidation, and insulin-mediated glycogen synthesis and signaling will be assessed. Aim 2 will determine if the IMCL accumulation is dependent on adipose tissue secretary proteins. We will use co-cultures of adipocytes, myoblasts, and adipose stromal vascular cells to examine IMCL and the development of insulin resistance. Aim 3 will determine whether the stromal fraction from IGT subjects promotes IMCL more effectively than that from NGT subjects in co-cultures with muscle cells. We will compare the stromal vascular fractions with regard to monocyte/macrophage accumulation and cytokine expression. Aim 4 will determine if improved glucose tolerance in response to a 10- week treatment with pioglitazone results in decreased IMCL and identify cellular mechanisms involved. Co-culture studies will also be used with muscle and stromal cells, before and after pioglitazone treatment. These experiments will provide mechanistic insight into the link between obesity and muscle function leading to metabolic syndrome.