Novel Molecular Determinants of Insulin Clearance

胰岛素清除率的新分子决定因素

• 批准号: 10446927
• 负责人: Sonia M. Najjar
• 金额: $ 48.8万
• 依托单位: WESTERN UNIVERSITY OF HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-25 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10446927
• 关键词: Ablation; Adipose tissue; Affect; Animals; Antidiabetic Drugs; Area; Attention; Automobile Driving; Beta Cell; Binding; Blindness; Blood Circulation; C-Peptide; CEACAM1; Cell Line; Cells; Collection; Compensatory Hyperinsulinemia; Defect; Dependovirus; Development; Diabetes Mellitus; Diabetes prevention; Down-Regulation; Epidemic; Etiology; Excision; Foundations; Functional disorder; Genes; Genetic; Genetic Determinism; Hepatic; Hepatocyte; Heritability; Homeostasis; Human; Hybrids; Hyperinsulinism; Impairment; Inbred Strains Mice; Individual; Insulin; Insulin Receptor; Insulin Resistance; Investigation; Kidney Failure; Knowledge; Lead; Liver; Measures; Mediating; Mediator of activation protein; Metabolic; Metabolism; Molecular; Mus; Muscle; Non-Insulin-Dependent Diabetes Mellitus; Obese Mice; Obesity; Pathway interactions; Pharmacology; Phosphorylation; Physiological; Population; Prediabetes syndrome; Prevalence; Prevention; Process; Regulation; Research; Risk; Role; Signal Transduction; Testing; Text; Therapeutic; Variant; Work; base; blood glucose regulation; cardiovascular disorder risk; cell growth regulation; db/db mouse; diabetes risk; diet-induced obesity; follow-up; gene function; genetic approach; genome wide association study; hepatoma cell; insight; insulin regulation; insulin secretion; limb amputation; longitudinal analysis; loss of function; novel; novel therapeutic intervention; overexpression; pancreatic juice; receptor; trafficking; transcriptome; translational potential

Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. Type 2 diabetes (T2D) is associated with elevated risk of cardiovascular disease and a major cause of blindness, limb amputation and kidney failure. It has reached epidemic proportions worldwide with a prevalence exceeding 10% in the US. The pathophysiology of T2D involves defects in insulin homeostasis including insufficient insulin secretion from the pancreas, impaired insulin action in muscle, liver and adipose tissues (insulin resistance), and reduced insulin removal from the circulation (insulin clearance) predominantly by the liver. Understanding the cellular and molecular mechanisms of insulin secretion and insulin resistance has been a major focus of investigations for decades, which has resulted in successful antidiabetic therapeutic strategies. In contrast, insulin clearance has been a relatively understudied area in diabetes research. Its molecular determinants are incompletely understood and its role in the etiology of T2D remains unclear. Reduced insulin clearance may represent a beneficial metabolic adaptation to insulin resistance that promotes compensatory hyperinsulinemia to limit the burden on -cells and likely protect against T2D. Conversely, it has been proposed that impaired insulin clearance may exacerbate insulin resistance by driving hyperinsulinemia- mediated downregulation of insulin receptors. It has also been hypothesized that genetically reduced hepatic insulin clearance constitute a primary causal factor in the development of T2D. Clearly, the mechanisms and physiological correlates of insulin clearance deserve further attention. A limitation of experimental investigations of insulin clearance is the relative dearth of known molecular determinants that regulate this process. In previous studies, we identified CEACAM1 as a critical factor in hepatic insulin clearance. While the role of CEACAM1 in receptor-mediated insulin internalization, an initial step in cellular insulin clearance, and metabolic homeostasis is now well established, the molecular mechanisms and mediators of subsequent steps in cellular insulin trafficking are less well understood. Thus, the overall objective of the present proposal is to extend our understanding of this process through the discovery of novel molecular determinants and characterization of their role in systemic insulin/glucose homeostasis. Motivated by the high heritability of insulin clearance observed in humans, we embarked on a hypothesis-free genetic approach. Using a collection of ~100 inbred mouse strains in the Hybrid Mouse Diversity Panel, we performed transcriptome and genome-wide association studies (GWAS) to identify genes and molecular pathways associated with steady-state C-peptide/insulin (C/I) molar ratio, a surrogate measure of insulin clearance. Our preliminary results provide several novel insights. First, they implicate the AMPK pathway in the regulation of cellular insulin clearance and identify this process as a previously unrecognized metabolic target of AMPK signaling. Moreover, we identified Tmem175 as a novel genetic determinant of insulin clearance in the mouse. While Tmem175 has not previously been characterized in the context of insulin metabolism, it has been implicated in endosomal pH regulation, a key process in the degradation and intracellular trafficking of insulin and the insulin receptor (INSR). Furthermore, the TMEM175 chromosomal region is associated with T2D in human populations, which lends support to the hypothesis that variation in TMEM175 activity may affect diabetes risk and highlights the potential translational relevance of this project. We will follow up on our preliminary results as described below: Specific Aim 1: To investigate the role of AMPK pathway in insulin clearance. We will use primary hepatocytes to investigate the cellular mechanisms responsible for the regulation of insulin clearance by AMPK. Pharmacological and genetic approaches will be employed to modulate AMPK activity and the impact on insulin binding, internalization and degradation, as well as the endocytic trafficking of INSR will be assessed. To identify downstream effectors of AMPK signaling in the regulation of insulin clearance, we will test the role of direct AMPK phosphorylation targets in cellular insulin clearance using gain- and loss-of- function approaches. The effect of AMPK signaling on systemic insulin clearance will be evaluated through pharmacological activation of AMPK in C57BL/6J mice and in mice with liver-specific genetic ablation of AMPK. Specific Aim 2: To investigate the role of Tmem175 in insulin clearance and metabolic homeostasis. To gain mechanistic insights into the role of Tmem175 in cellular insulin metabolism, we will assess insulin binding, internalization, degradation and INSR trafficking in primary hepatocytes and hepatoma cell lines using gain- and loss-of-function approaches. The role of Tmem175 in systemic insulin clearance will be assessed in whole-body Tmem175-deficient mice and its impact on hepatic insulin extraction will be investigated in mice with adeno-associated virus (AAV)-mediated liver-specific suppression. Furthermore, we will evaluate the metabolic impact of Tmem175 deficiency through longitudinal analyses of insulin and glucose homeostasis in the context of pre-diabetes using diet-induced obese (DIO) mice and the genetically obese db/db mice with established T2D. We anticipate that these studies will provide novel mechanistic insights into the regulation of insulin clearance and its role in the pathophysiology of T2D. Furthermore, our work will lay the foundation for novel therapeutic approaches in T2D based on the manipulation of insulin clearance.

在此处输入文本，这是您应用程序的新摘要信息。本节必须不超过30行文本。 2型糖尿病（T2D）与心血管疾病的风险升高以及失明，肢体截肢和肾衰竭的主要原因有关。它在全球范围内达到了流行比例，美国的流行率超过10％。 T2D的病理生理涉及胰岛素稳态中的缺陷，包括胰腺中胰岛素分泌不足，胰岛素，肝脏和脂肪组织中的胰岛素作用受损（胰岛素抵抗）以及从LIVER中占循环（胰岛素清除）的减少。了解数十年来，了解胰岛素分泌和胰岛素抵抗的细胞和分子机制一直是研究的重点，这导致了成功的抗糖尿病治疗策略。相比之下，胰岛素清除一直是糖尿病研究中相对了解的领域。它的分子决定剂未完全理解，其在T2D病因中的作用尚不清楚。减少的胰岛素清除率可能代表了对胰岛素抵抗的有益代谢适应性，该适应性促进了代偿性高胰岛素清除率，以限制细胞的燃烧，并可能预防T2D。相反，已经提出，胰岛素清除受损可能会通过驱动女超胰岛素缓解剂来加剧胰岛素抵抗。还假设，通常降低肝胰岛素清除率构成T2D发展的主要因果因素。显然，胰岛素清除的机制和物理相关性值得进一步关注。 胰岛素清除实验研究的局限性是调节该过程的已知分子确定剂的相对死亡。在先前的研究中，我们将CEACAM1确定为肝炎胰岛素清除率的关键因素。尽管CEACAM1在受体介导的胰岛素内部化中的作用，现在已经确定了细胞胰岛素清除率和代谢稳态的第一步，但分子机制和随后步骤的介体在细胞胰岛素运输中的介导者的了解程度较低。这是当前建议的总体目标是通过发现新的分子决定词以及它们在系统性胰岛素/葡萄糖稳态中的作用来扩展我们对这一过程的理解。 由人类观察到的胰岛素清除率高的高遗传力的动机，我们着手采用一种假设的遗传方法。使用杂交小鼠多样性面板中约100个近交小鼠菌株的集合，我们进行了转录组和全基因组关联研究（GWAS），以识别与稳态C肽/胰岛素（C/I）摩尔比，这是一种胰岛素清除率的替代度量的基因和分子途径。我们的初步结果提供了一些新颖的见解。首先，它们将AMPK途径牵涉到细胞胰岛素清除率的调节中，并确定该过程是AMPK信号的先前未识别的代谢靶标。此外，我们将TMEM175确定为小鼠胰岛素清除率的新遗传决定因素。尽管TMEM175以前尚未在胰岛素代谢的背景下进行表征，但它与内体pH调节有关，这是胰岛素降解和细胞内运输的关键过程和胰岛素受体（INSR）。此外，TMEM175染色体区域与人群中T2D有关，这支持了以下假设：TMEM175活性的变化可能影响糖尿病的风险，并突出该项目的潜在转化相关性。我们将跟进我们的初步结果，如下所述： 具体目的1：研究AMPK途径在胰岛素清除率中的作用。我们将使用原发性肝细胞来研究负责调节AMPK胰岛素清除率的细胞机制。将使用药理学和遗传方法来调节AMPK活性以及对胰岛素结合，内在化和降解的影响，以及INSR的内吞交易。为了确定AMPK信号在调节胰岛素清除率中的下游效应，我们将使用获得功能丧失方法来测试直接AMPK磷酸化靶标在细胞胰岛素清除率中的作用。 AMPK信号传导对全身胰岛素清除率的影响将通过在C57BL/6J小鼠中的AMPK和具有肝脏特异性AMPK遗传消融的小鼠中评估。 具体目标2：研究TMEM175在胰岛素清除和代谢稳态中的作用。为了获得对TMEM175在细胞胰岛素代谢中的作用的机理见解，我们将使用增益和功能障碍方法评估胰岛素结合，内在化，降解和INSR运输。 TM175在全身TMEM175缺乏小鼠中的作用将在全身胰岛素清除率中进行评估，并且将在具有腺相关病毒（AAV）介导的肝脏特异性抑制的小鼠中研究其对肝胰岛素提取的影响。此外，我们将使用饮食诱导的肥胖（DIO）小鼠以及具有既定T2D的遗传性肥胖的肥胖小鼠（DIO）小鼠在糖尿病前进行胰岛素和葡萄糖稳态的纵向分析来评估TMEM175缺乏症的代谢影响。 我们预计这些研究将为调节胰岛素清除率及其在T2D的病理生理学中的作用提供新的机械见解。此外，我们的工作将基于操纵胰岛素清除率，为T2D的新型治疗方法奠定了基础。