Endoplasmic Reticulum Chaperone as a Regulator of Obesity and Diabetes

描述（由申请人提供）：内质网（ER）是一种细胞器，分泌蛋白和膜相关蛋白在其中合成和修饰。有人提出，肥胖会促进营养压力和慢性炎症，这涉及到许多分泌器官系统（如脂肪组织）对细胞合成机制的需求增加。因此，肥胖作为外周组织内质网应激的慢性刺激，引发胰岛素抵抗和2型糖尿病。多功能内质网伴侣蛋白GRP78/BiP通过控制蛋白质折叠和跨膜内质网应激传感器的激活而成为内质网稳态的主要调节因子。我们偶然发现，C57BL/6背景的Grp78小鼠对高脂肪饮食（HFD）诱导的肥胖表现出抵抗性，并改善了胰岛素敏感性。我们的初步研究表明，饲喂hfd的Grp78小鼠能量消耗增加，但食物摄入量和脂质吸收没有变化。初步的血糖钳夹研究显示，在Grp78小鼠的白色脂肪组织中，胰岛素刺激的葡萄糖摄取显著增加（P<0.001）。我们进一步发现，在脂肪组织中，Grp78杂合性导致饮食诱导的ER伴侣上调。相比之下，骨骼肌的葡萄糖代谢没有改变，也没有观察到伴侣蛋白上调。因此，Grp78小鼠为研究内质网完整性与能量平衡、葡萄糖稳态和脂肪细胞应激之间的基本机制提供了新的机会。我们提出，在HFD诱导的慢性应激过程中，脂肪组织中的Grp78杂合性触发代偿和保护措施，如伴侣上调和线粒体功能增加，从而导致能量消耗增加，内质网应激和炎症反应减弱，从而改善胰岛素敏感性。根据我们的初步数据，Grp78小鼠比体重相近的野生型鼠对胰岛素更敏感，Aim 1将确定Grp78改善胰岛素敏感性的机制。目的2将确定肥胖如何影响未折叠蛋白反应信号，以及Grp78杂合性如何改变能量消耗并引起对饮食诱导肥胖的抵抗。目的3将生成和表征GRP78白色脂肪组织特异性过表达或敲除的小鼠模型，以确定其在能量平衡和胰岛素敏感性中的作用。Aim 4将利用来自Grp78和小鼠的原代脂肪细胞和mef，以及3T3-L1脂肪细胞培养系统，确定hfd喂养的Grp78小鼠脂肪组织中上调的其他ER伴侣在脂肪细胞代谢功能中的功能贡献。我们研究的临床意义在于，它们可能发现饮食性肥胖和胰岛素抵抗的新调控途径，这可能为人类代谢性疾病的治疗提供新的靶点。公共卫生相关性：肥胖、胰岛素抵抗和2型糖尿病发病率的急剧增加已成为对人类健康最严重的威胁之一。因此，了解这些疾病的分子机制至关重要。这一建议是基于最近在实验室建立的一种新型小鼠模型的偶然观察，该模型可能为预防高脂肪饮食引起的肥胖和随后的胰岛素抵抗提供线索。这一建议将充分表征突变小鼠的代谢表型，并研究高脂肪饮食慢性应激下这些小鼠能量消耗增加和胰岛素敏感性改善的潜在机制。如果得到证实，我们的发现可能会为人类肥胖和2型糖尿病的治疗提供新的靶点。

DESCRIPTION (provided by applicant): The endoplasmic reticulum (ER) is a cellular organelle where secretory and membrane-associated proteins are synthesized and modified. It has been proposed that obesity promotes nutrient stress and chronic inflammation that involve increased demand on the synthetic machinery of the cells in many secretory organ systems, such as adipose tissue. Thus, obesity acts as a chronic stimulus for ER stress in peripheral tissues, triggering insulin resistance and type 2 diabetes. The multifunctional ER chaperone protein GRP78/BiP, is a master regulator of ER homeostasis due to its control of protein folding and the activation of trans-membrane ER stress sensors. Serendipitously, we discovered that the Grp78 mice in the C57BL/6 background exhibit resistance to high-fat diet (HFD)-induced obesity and improved insulin sensitivity. Our preliminary studies revealed that HFD-fed Grp78 mice showed increased energy expenditure without changes in food intake and lipid absorption. Preliminary euglycemic clamp studies showed a striking increase in insulin-stimulated glucose uptake most prominently in the white adipose tissue of the Grp78 mice (P<0.001). We further discovered that in adipose tissue, Grp78 heterozygosity leads to diet-induced upregulation of ER chaperones. In contrast, glucose metabolism was not altered in skeletal muscle and no chaperone upregulation was observed. Thus, Grp78 mice offer new opportunities to investigate the basic mechanisms linking ER integrity to energy balance, glucose homeostasis and adipocyte stress. We propose that during chronic stress induced by HFD, Grp78 heterozygosity in adipose tissue triggers compensatory and protective measures, such as upregulation of chaperones and increase in mitochondrial function, which lead to enhanced energy expenditure, attenuation of ER stress and inflammatory responses resulting in improved insulin sensitivity. Based on our preliminary data that the Grp78mice are more insulin sensitive than the wild-type littermates on chow diet with similar body weights, Aim 1 will identify the mechanism by which Grp78 improves insulin sensitivity. Aim 2 will determine how obesity affects the unfolded protein response signaling and how Grp78 heterozygosity alters energy expenditure and causes resistance to diet-induced obesity. Aim 3 will generate and characterize mouse models with white adipose tissue-specific overexpression or knockout of GRP78 to determine its role in energy balance and insulin sensitivity. Aim 4 will determine the functional contribution of other ER chaperones upregulated in adipose tissue of HFD-fed Grp78 mice in adipocyte metabolic function, utilizing primary adipocytes and MEFs from the Grp78 and mice, as well as the 3T3-L1 adipocyte culture system. The clinical relevance of our studies is that they may identify novel regulatory pathways for diet- induced obesity and insulin resistance which may represent new therapeutic targets for human metabolic diseases. PUBLIC HEALTH RELEVANCE: The dramatic increase in the incidence of obesity, insulin resistance and type 2 diabetes has become one of the most serious threats to human health. Hence, understanding the molecular mechanisms underlying these diseases is critical. This proposal is based on the serendipitous observation from a novel mouse model recently created in the laboratory that may provide clues to prevent high-fat diet-induced obesity and subsequent insulin resistance. This proposal will fully characterize the metabolic phenotypes of the mutant mice and investigate the underlying mechanisms for increased energy expenditure and improved insulin sensitivity of these mice under the chronic stress of high-fat diet. If validated, our findings may lead to new targets for therapy against obesity and type 2 diabetes in humans.