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饥饿和过氧化物酶体增殖物激活受体-α激活引发大鼠胰岛中丙酮酸脱氢酶激酶同工型表达的选择性改变:对葡萄糖刺激的胰岛素分泌的影响

Selective modification of pyruvate dehydrogenase kinase isoform expression in rat pancreatic islets elicited by starvation and activation of peroxisome proliferator-activated receptor-alpha: implications for glucose-stimulated insulin secretion.

作者信息

Sugden M C, Bulmer K, Augustine D, Holness M J

机构信息

Department of Diabetes and Metabolic Medicine, Division of General and Developmental Medicine, St. Bartholomew's and the Royal London School of Medicine and Dentistry, Queen Mary, University of London, London, U.K.

出版信息

Diabetes. 2001 Dec;50(12):2729-36. doi: 10.2337/diabetes.50.12.2729.

Abstract

The pyruvate dehydrogenase complex (PDC) has a pivotal role in islet metabolism. The pyruvate dehydrogenase kinases (PDK1-4) regulate glucose oxidation through inhibitory phosphorylation of PDC. Starvation increases islet PDK activity (Am J Physiol Endocrinol Metab 270:E988-E994, 1996). In this study, using antibodies against PDK1, PDK2, and PDK4 (no sufficiently specific antibodies are as yet available for PDK3), we identified the PDK isoform profile of the pancreatic islet and delineated the effects of starvation (48 h) on protein expression of individual PDK isoforms. Rat islets were demonstrated to contain all three PDK isoforms, PDK1, PDK2, and PDK4. Using immunoblot analysis with antibodies raised against the individual recombinant PDK isoforms, we demonstrated increased islet protein expression of PDK4 in response to starvation (2.3-fold; P < 0.01). Protein expression of PDK1 and PDK2 was suppressed in response to starvation (by 27% [P < 0.01] and 10% [NS], respectively). We demonstrated that activation of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) by the selective agonist WY14,643 for 24 h in vivo leads to specific upregulation of islet PDK4 protein expression by 1.8-fold (P < 0.01), in the absence of change in islet PDK1 and PDK2 protein expression but in conjunction with a 2.2-fold increase (P < 0.01) in islet PPAR-alpha protein expression. Thus, although no changes in islet PPAR-alpha expression were observed after the starvation protocol, activation of PPAR-alpha in vivo may be a potential mechanism underlying upregulation of islet PDK4 protein expression in starvation. We evaluated the effects of antecedent changes in PDK profile and/or PPAR-alpha activation induced by starvation or PPAR-alpha activation in vivo on glucose-stimulated insulin secretion (GSIS) in isolated islets. GSIS at 20 mmol/l glucose was modestly impaired on incubation with exogenous triglyceride (1 mmol/l triolein) ( approximately 20% inhibition; P < 0.05) in islets from fed rats. Starvation (48 h) impaired GSIS in the absence of triolein (by 57%; P < 0.001), but GSIS after the further addition of triolein did not differ significantly between islets from fed or starved rats. GSIS by islets prepared from WY14,643-treated fed rats did not differ significantly from that seen with islets from control fed rats, and the response to triolein addition resembled that of islets prepared from fed rather than starved rats. PPAR-alpha activation in vivo led to increased insulin secretion at low glucose concentrations. Our results are discussed in relation to the potential impact of changes in islet PDK profile on the insulin secretory response to lipid and of PPAR-alpha activation in the cause of fasting hyperinsulinemia.

摘要

丙酮酸脱氢酶复合体(PDC)在胰岛代谢中起关键作用。丙酮酸脱氢酶激酶(PDK1 - 4)通过对PDC的抑制性磷酸化作用来调节葡萄糖氧化。饥饿会增加胰岛PDK活性(《美国生理学杂志:内分泌与代谢》270:E988 - E994,1996)。在本研究中,我们使用针对PDK1、PDK2和PDK4的抗体(目前尚无针对PDK3的足够特异性抗体),确定了胰岛中PDK同工型的分布情况,并描述了饥饿(48小时)对各个PDK同工型蛋白表达的影响。已证实大鼠胰岛含有所有三种PDK同工型,即PDK1、PDK2和PDK4。通过使用针对各个重组PDK同工型产生的抗体进行免疫印迹分析,我们发现饥饿会使胰岛中PDK4的蛋白表达增加(2.3倍;P < 0.01)。饥饿会使PDK1和PDK2的蛋白表达受到抑制(分别降低27% [P < 0.01]和10% [无统计学意义])。我们证明,在体内用选择性激动剂WY14,643激活过氧化物酶体增殖物激活受体α(PPAR - α)24小时,会导致胰岛PDK4蛋白表达特异性上调1.8倍(P < 0.01),而胰岛PDK1和PDK2的蛋白表达没有变化,但同时胰岛PPAR - α蛋白表达增加2.2倍(P < 0.01)。因此,尽管在饥饿处理后未观察到胰岛PPAR - α表达的变化,但体内PPAR - α的激活可能是饥饿时胰岛PDK4蛋白表达上调的潜在机制。我们评估了饥饿或体内PPAR - α激活诱导的PDK分布变化和/或PPAR - α激活对分离胰岛中葡萄糖刺激的胰岛素分泌(GSIS)的影响。在喂食大鼠的胰岛中,与外源性甘油三酯(1 mmol/l三油酸甘油酯)一起孵育时,20 mmol/l葡萄糖浓度下的GSIS受到适度损害(约20%抑制;P < 0.05)。饥饿(48小时)在没有三油酸甘油酯的情况下会损害GSIS(降低57%;P < 0.001),但进一步添加三油酸甘油酯后,喂食或饥饿大鼠的胰岛之间的GSIS没有显著差异。用WY14,643处理的喂食大鼠制备的胰岛的GSIS与对照喂食大鼠的胰岛没有显著差异,并且对添加三油酸甘油酯的反应类似于喂食而非饥饿大鼠制备的胰岛。体内PPAR - α激活会导致低葡萄糖浓度下胰岛素分泌增加。我们结合胰岛PDK分布变化对脂质胰岛素分泌反应的潜在影响以及PPAR - α激活在空腹高胰岛素血症病因中的作用来讨论我们的结果。

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