Metabolism and Growth in a Teleost Fish Model of Diabetes Mellitus: Insulin Regulation in an Ectothermic Vertebrate

[600783]一种独特的糖尿病模型已经在一种海洋鱼类中被开发出来，这种鱼就是神奇虾虎鱼（Gillichthys mirabilis）。与其他动物的胰腺内分泌细胞（即产生胰岛素和胰高血糖素的细胞）与外分泌细胞（即产生消化酶的细胞）位于同一组织的解剖结构相反，这种虾虎鱼只有一个胰腺内分泌器官，称为“胰岛”，其中包含纯粹的产生激素的细胞。因此，虾虎鱼可以进行简单的手术“胰岛切除术”，完全切除内分泌细胞，并产生胰岛素依赖型糖尿病（IDDM）的直接症状。然而，与哺乳动物模型相反，在哺乳动物模型中，IDDM是由产生胰岛素的细胞（-细胞）的单一损失引起的，虾虎鱼的胰岛切除术同时去除（-细胞）和(-细胞（胰高血糖素的来源）。在哺乳动物中，不受胰岛素影响的“反调节性”胰高血糖素被认为是IDDM发病阶段驱动高血糖的主要因素。然而，目前还没有可用的实验模型，可以将(-细胞和（-细胞）都去除，以便直接验证这一假设。该PI旨在描述胰高血糖素在虾虎鱼模型中IDDM发病过程中的致糖尿病作用，并特别关注肝脏和血液中的代谢机制。还将研究肾上腺糖皮质激素（皮质醇）的影响，皮质醇是糖尿病哺乳动物发病后重要的致糖尿病激素。除了代谢紊乱外，Ix虾虎鱼还表现出强烈的糖尿病生长抑制，包括胰岛素样生长因子（IGF）生物活性（阳性体细胞生长指标）的降低，以及低分子量IGF结合问题的显著增加，这可能与哺乳动物对代谢敏感、抑制IGF的“IGFBP-1”有关。利用igfbp -1样蛋白在Ix细胞中这种明显的代谢调节，该项目旨在表征该蛋白的内分泌/代谢调节及其生长调节作用，并尝试使用基于pcr的策略克隆其基因。PI特别感兴趣的是，这种蛋白质的测量是否可以作为自然和水产养殖鱼类种群健康、生长状态和应激的方便指标。由于先前缺乏低等脊椎动物胰岛素缺乏的模型，关于胰岛素在变温动物（“冷血”动物）中的生物学作用的研究仅限于对完整动物的激素注射研究。Ix虾虎鱼模型将使PI能够在恒温动物（“温血动物”）进化之前获得以前无法获得的关于代谢调节的见解。据推测，恒温动物的生理发展，就像哺乳动物和鸟类的进化一样，需要发展一个更有效的代谢调节系统，增加对胰岛素调节作用的依赖。在Ix虾虎鱼中，PI发现胰岛素可调节的葡萄糖转运离子外周肌肉组织产生胰岛素抵抗，并且胰岛素治疗Ix虾虎鱼使葡萄糖转运系统再次对胰岛素反应。这些数据表明，胰岛素可能对鱼类葡萄糖转运蛋白发挥与哺乳动物类似的双重调节作用，即对转运蛋白动力学/活性的急性影响，对转运蛋白基因表达的长期影响。重要的是，这些数据还建立了一个可调节的实验系统，以适应另一个特定的目的：表征虾虎鱼的胰岛素调节和葡萄糖转运系统的细胞机制，特别是与吸热哺乳动物的相关；然后，PI计划使用基于pcr的策略克隆虾虎鱼的谷氨酸-4（哺乳动物的主要胰岛素调节葡萄糖转运蛋白）。在拟议的研究中，将强调代谢和生长机制之间的相互关系，以获得这种变温动物生理系统的整体图景。

9600783 Kelley A unique model of diabetes mellitus has been developed in a marine fish, the goby Gillichthys mirabilis. In contrast to the anatomical arrangement in other animals in which pancreatic endocrine cells (i.e., those producing insulin and glucagon) are located in the same tissue as exocrine cells (i.e., digestive enzyme-producing cells), this goby possesses a single pancreatic endocrine organ, called the "islet", which contains purely hormone-producing cells. The goby thereby allows for a simple surgical "isletectomy", which removes completely the endocrine cells and produces the outright symptomatology of insulin-dependent diabetes mellitus (IDDM). In contrast to mammalian models, however, in which IDDM results from the singular loss of the insulin-producing cells ((-cells), isletectomy in the goby removes simultaneously (-cells as well as (-cells (the source of glucagon). In mammals, "counterregulatory" glucagon, acting unchallenged by insulin, is believed to be a primary factor in driving the hyperglycemia during the onset phase of IDDM. There have been no experimental models available, however, in which both (- and (-cells can be removed in order to test this hypothesis directly. The PI aims to characterize the diabetogenic role of glucagon during IDDM onset in the goby model, with special attention given to the metabolic mechanisms occurring in the liver and blood. The influence of adrenal glucocorticoid (cortisol), a diabetogenic hormone important post-onset in diabetic mammals, will also be investigated. In addition to metabolic perturbation, the Ix goby exhibits a strong diabetic growth inhibition, including reductions in insulin-like growth factor (IGF) bioactivity (an indicator of positive somatic) growth) and a significant increase in a low molecular weight IGF-binding problem that may be related to the metabolically-sensitive, IGF-inhibitory "IGFBP-1" of mammals. Taking advantage of this apparent metabolic regulation of the IGFBP-1-like protein in the Ix g oby, the PI aims to characterize this protein's endocrine/metabolic regulation and its growth-regulatory role, and to attempt the cloning of its gene using a PCR-based strategy. The PI is particularly interested in whether measurement of this protein may serve as a convenient indicator of health, growth status, and stress in natural and aquacultured fish populations. Owing to the prior lack of a model of insulin deficiency in lower vertebrates, studies on the biological role of insulin in ectotherms ("cold-blooded" animals) have been limited to hormone injection studies on intact animals. The Ix goby model will allow the PI to gain previously-unattainable insight on metabolic regulation "before" the evolution of endothermy ("warm-bloodedness"). It has been hypothesized that the development of the physiology of endothermy, as has occurred in mammalian and avian evolution, necessitated the development of a more efficient metabolic regulatory system with an increased dependence on the regulatory influence of insulin. In the Ix goby, the PI has found that insulin-regulatable glucose transport ion peripheral muscle tissues becomes insulin resistant, and that insulin treatment of Ix gobies renders the glucose transport system once again responsive to insulin action. These data suggest that insulin may exert a dual regulation on fish glucose transporters comparable to that in mammals, i.e., acute effects on transporter dynamics/activity, long-term effects on transporter gene expression. Importantly, the data also establish a regulatable experimental system amenable to another specific aim: to characterize the insulin regulation and cellular mechanisms of the glucose transport system in the goby, especially as it relates to that of the endothermic mammal; the PI then plans to use a PCR-based strategy to clone the goby counterpart of glut-4 (the primary insulin-regulated glucose transporter of mammals). In the proposed studies, interrelationships between metabolic and growth mechanisms will be e mphasized, in order to gain an integrated picture of this ectotherm's physiological system.