Pyruvate kinase M2 levels and activation as protective factors for diabetic nephropathy

丙酮酸激酶 M2 水平和激活作为糖尿病肾病的保护因素

• 批准号: 9235747
• 负责人: GEORGE L KING
• 金额: $ 227.22万
• 依托单位: JOSLIN DIABETES CENTER
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-30 至 2019-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9235747
• 关键词: Adverse effects; Agonist; Aldehyde Reductase; Angiotensins; Apoptosis; Biogenesis; Cell physiology; Cells; Clinical; Clinical Data; Clinical Markers; Creatinine; Data; Development; Diabetes Mellitus; Diabetic Nephropathy; Diglycerides; Disease; End stage renal failure; Enzymes; Epidemiologic Studies; Glomerular Filtration Rate; Glucose; Glycolysis; Glycosylated hemoglobin A; Histologic; Human; Hyperglycemia; Hypertension; Image; Individual; Inflammation; Insulin Resistance; Insulin-Dependent Diabetes Mellitus; Kidney; Kidney Diseases; Kidney Failure; Kidney Glomerulus; Knockout Mice; Laboratories; Link; M2 protein; Mass Spectrum Analysis; Measures; Microalbuminuria; Microvascular Dysfunction; Mitochondria; Mus; Nature; Neuropathy; Organ; Oxidative Phosphorylation; Oxidative Stress; Pathology; Pathway interactions; Patients; Plasma; Population; Prevention; Production; Protein Isoforms; Protein Kinase C; Proteins; Proteomics; Pyruvaldehyde; Pyruvate Kinase; Reactive Oxygen Species; Renal Tissue; Renal function; Risk Factors; Rodent; Role; Sampling; Serum; Sorbitol; Specimen; Streptozocin; Testing; Toxic effect; Tubular formation; blood pressure regulation; cohort; comparison group; diabetic; diabetic patient; glycemic control; improved; in vivo; inhibitor/antagonist; kidney cortex; knock-down; mesangial cell; metabolomics; mitochondrial dysfunction; mutant; novel; overexpression; podocyte; prevent; proliferative diabetic retinopathy; restoration; small molecule; targeted treatment; theories; type I diabetic

Diabetic nephropathy (DN) is the leading cause of renal failure in the US. Hyperglycemia (HG) has been identified as the primary risk factor for DN, but therapies targeting the known pathways of HG-induced renal damage have not shown significant benefits. We proposed endogenous protective mechanisms exist which mitigate the adverse effects of HG that contribute to DN development. Examination of a unique cohort of type 1 diabetic (T1D) patients (Joslin Medalist Study; n=1007) who have had the disease for 50–87 years, of whom only 13.2% have significant renal disease independent of HbA1c support this conclusion. Proteomic analysis of Medalists' post-mortem renal glomeruli, comparing DN class 0-I to IIB –III identified multiple enzymes in the glycolytic, aldose reductase, methylglyoxal (MG) and mitochondrial pathways as higher in those protected from DN. Additionally, metabolomic analyses comparing those with DN (eGFR <45 ml/min/1.73 m2) and those without (eGFR >90 ml/min/1.73 m2) showed many glycolytic intermediates, sorbitol, MG and DAG to be decreased. This suggests an ability to metabolize intracellular glucose via glycolysis, the aldose reductase pathway, MG degradation and mitochondrial oxidative phosphorylation. A subset comparison of metabolites from Medalists' without DN to younger individuals who progressed rapidly to DN supports PKM2's protective nature. Plasma PKM2 levels from the Medalists correlated with eGFR, MG and DN class. Therefore, activating the key regulatory allosteric glycolytic enzyme, pyruvate kinase (PK), especially the M2 isoform, may increase flux via glycolysis and the mitochondrial oxidative phosphorylation pathways and prevent the onset or stop DN progression. We have documented PKM2 activation reversed HG-induced elevations of diacylglycerol, protein kinase C, sorbitol, MG, and mitochondrial dysfunction, as well as apoptosis in podocytes. Further, knockdown of PKM2 exaggerated the toxic effects of HG. In vivo studies using a small molecule activator of PKM2 and PKM2 podocyte specific knock-outs support the importance of PKM2 to normalize mitochondrial biogenesis and function, and prevent pathology of DN in the presence of HG. These data indicate increases in glycolysis and mitochondrial function, possibly by PKM2 activation, can prevent renal damage due to HG. Specific Aim 1: Validate and reproduce the association of glomeruli PKM2 protein levels with renal cortex and serum metabolite levels in post-mortem glomerular specimens from Medalists to establish a relationship with PKM2 plasma levels and clinical markers of renal function. Specific Aim 2: Compare plasma PKM2 levels, serum metabolite profiles and clinical renal markers in samples from the Medalist population and in a shorter duration population, and establish a longitudinal relationship with PKM2 levels and the progression of renal disease. Specific Aim 3: Characterize the mechanism that causes the decreased expression or activities of PKM2 by HG and the effect of its activation to restore mitochondria and cellular function, and survival of podocytes in HG conditions.

糖尿病性肾病（DN）是美国肾衰竭的主要原因。高血糖（HG）已经 被确定为DN的主要危险因素，但针对HG诱导肾脏的已知途径的疗法 损坏尚未显示出明显的好处。我们提出了内源性保护机制，存在 减轻对DN发育有助的HG的不良影响。检查独特的1型队列 糖尿病（T1D）患者（乔斯林奖章研究； n = 1007），患有该病已有50-87年 只有13.2％的肾脏疾病与HBA1C无关，支持这一结论。蛋白质组学分析 奖章者的验证后肾肾球体，将DN类0-I与IIB –III进行比较，确定了多种酶 在受保护的人中，糖酵解，醛降低，甲基乙醛（Mg）和线粒体途径较高 Dn。此外，比较DN（EGFR <45 mL/min/1.73 m2）的代谢组分析和 没有（EGFR> 90 mL/min/1.73 m2）显示了许多糖酵解中间体，山梨糖醇，Mg和DAG为 减少。这表明可以通过糖酵解代谢细胞内葡萄糖，醛糖会减少 途径，MG降解和线粒体氧化磷酸化。代谢物的子集比较 从没有DN的奖章者到迅速发展为DN的年轻人，支持PKM2受保护的人 自然。来自奖章的血浆PKM2水平与EGFR，MG和DN类相关。因此，激活 关键的调节型糖酵解酶，丙酮酸激酶（PK），尤其是M2同工型，可能会增加 通过糖酵解和线粒体氧化磷酸化途径的通量，并防止发作或停止DN 进展。我们已经记录了PKM2激活逆转了HG诱导的二酰基甘油的升高，蛋白 激酶C，山梨糖醇，MG和线粒体功能障碍以及足细胞中的凋亡。此外，击倒 PKM2夸大了Hg的毒性作用。使用PKM2和 PKM2 Podocyte特异性敲除支持PKM2正常化线粒体生物发生的重要性 和功能，并在Hg存在下预防DN的病理。这些数据表明糖酵解的增加 线粒体功能（通过PKM2激活）可以防止由于Hg引起的肾脏损伤。具体目标 1：验证和复制肾小球PKM2蛋白水平与肾皮质和系列的关联 验证奖学金的验证后标本中的代谢物水平与PKM2建立关系 肾功能的血浆水平和临床标记。特定目标2：比较等离子体PKM2水平，串行 来自奖牌人群和较短持续时间的样品中的代谢物概况和临床肾脏标记 人口，并与PKM2水平和肾脏疾病的进展建立纵向关系。 特定目的3：表征导致PKM2表达或活性降低的机制 Hg及其激活恢复线粒体和细胞功能的影响，以及hg中足细胞的存活率 状况。