Beta-Adrenergic Receptor Modulation of Insulin Signaling

描述（由申请人提供）：糖尿病是一种日益严重的流行病。它是由于身体无法产生或调节胰岛素而导致过量葡萄糖的积累而引起的。虽然有大量的研究正在进行各种器官中的胰岛素信号传导，但很少有研究涉及眼睛中的胰岛素调节，因为它可能与糖尿病视网膜病变有关。糖尿病视网膜病变是美国工作年龄人群视力丧失的主要原因。在糖尿病发作期间，在身体的不同区域发生交感神经活动的丧失，导致肾脏、心脏和外周血管系统的功能障碍。由于已经假设交感神经在糖尿病发作时受到损害，这也可能意味着它们在眼睛中发挥作用。对交感神经支配及其在眼睛中的作用的进一步研究表明，交感神经在组织学和功能变化中起作用，这些变化与糖尿病视网膜病变中发生的变化相似。视网膜中可能对交感神经传递的变化敏感的一种细胞类型是穆勒细胞。米勒细胞作为视网膜的结构支持细胞，并跨越视网膜的整个厚度。由于Muller细胞被激活并表达增加的GFAP水平，这表明Muller细胞正在受到损害，并且还可能改变其对炎症标志物、葡萄糖转运、氧化应激、生长因子以及最终细胞存活的调节。对于本提案中的工作，大鼠Muller细胞（rMC-1）将在高糖（25 mM）或低糖（5 mM）条件下生长的DMEM培养基中培养，并用10 μ M异丙肾上腺素处理，随后进行胰岛素信号传导和凋亡的各种分析。初步数据表明，在高糖条件下培养的Muller细胞中，胰岛素受体信号传导减少。为了进一步描述胰岛素信号传导的作用，将使用针对胰岛素受体的siRNA进行实验，以进一步鉴定β-肾上腺素能受体对细胞凋亡的调节作用以及β-肾上腺素能受体与胰岛素信号传导之间的串扰。除了对培养细胞的研究外，我们还将使用β-1-肾上腺素能受体敲除小鼠及其同窝仔，以更具体地定义β-1-肾上腺素能受体信号传导对体内胰岛素信号传导和细胞凋亡的作用。这项建议与公共卫生的相关性将是确定糖尿病视网膜中β-肾上腺素能受体和胰岛素信号的相互作用。由于糖尿病患者通常需要胰岛素治疗，因此确定胰岛素和其他治疗后视网膜中细胞事件的调节至关重要。

DESCRIPTION (provided by applicant): Diabetes is a growing epidemic. It results from buildup of excess glucose caused by the body's inability to produce or regulate insulin. While there is a significant amount of research being conducted on insulin signaling in various organs, very little of that research has involved insulin regulation in the eye as it may relate to diabetic retinopathy. Diabetic retinopathy is the leading cause of vision loss in people of working age in the United States. During the onset of diabetes there is a loss of sympathetic nerve activity that takes place in different regions of the body, leading to dysfunction of the kidneys, heart, and peripheral vasculature. Since it has been postulated that sympathetic nerves are compromised with the onset of diabetes, this also could mean that they play a role in the eye. A further look into sympathetic innervation and its role in the eye has indicated that sympathetic nerves are playing a role in histological and functional changes that are similar to those occurring in diabetic retinopathy. One cell type in the retina that may be susceptible to changes in sympathetic neurotransmission is the Muller cell. Muller cells serve as structural support cells for the retina, and span the entire thickness of the retina. Since Muller become activated and express increased GFAP levels, this suggests that Muller cells are being compromised and may also alter their regulation of inflammatory markers, glucose transport, oxidative stress, growth factors, and finally cell survival. For the work in this proposa, rat Muller cells (rMC-1) will be cultured in DMEM medium grown in high glucose (25mM) or low glucose (5mM) conditions, and treated with 10uM isoproterenol followed by various analyses of insulin signaling and apoptosis. Preliminary data suggest that insulin receptor signaling is decreased in Muller cells cultured in high glucose conditions. To further delineate the role of insulin signaling, experiments will be done using siRNA against the insulin receptor to further identify roles in the regulation of apoptosis by beta-adrenergic receptors and cross-talk between beta-adrenergic receptors and insulin signaling. In addition to the work on cells in culture, we will also use beta-1-adrenergic receptor knockout mice and their littermates to more specifically define the role of beta-1-adrenergic receptor signaling on insulin signaling and apoptosis in vivo. The relevance to public health from this proposal will be in determining the interactions of beta-adrenergic receptors and insulin signaling in the diabetic retina. Since diabetic patients often require insulin treatment, it is critical to determine the regulation of cellular events in the retina following insulin and other treatments.