STEROID REGULATION OF B-ADRENORECEPTOR GENES IN OBESITY

肥胖中 B 肾上腺素受体基因的类固醇调节

• 批准号: 2444084
• 负责人: SHEILA COLLINS
• 金额: $ 10.49万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-07-01 至 1999-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2444084
• 关键词: G protein; RNase protection assay; adenylate cyclase; adipocytes; adipose tissue; adrenalectomy; beta adrenergic receptor; biological signal transduction; cell differentiation; corticosteroid inhibitor; gel mobility shift assay; gene expression; genetic models; genetic promoter element; genetic transcription; glucocorticoids; hormone regulation /control mechanism; laboratory mouse; lipolysis; northern blottings; nuclear runoff assay; obesity; receptor binding; receptor expression; tissue /cell culture

The congenital obesity, beta-adrenergic receptor (betaAR) stimulation of lipolysis and thermogenesis are impaired. Both white and brown adipocytes express beta1-, beta2-, and beta3AR (the latter is expressed exclusively in fat). Our preliminary data demonstrate that beta1AR and beta3AR are impaired in white and brown adipocytes from genetically obese (ob/ob) mice compared to lean controls. This defect may contribute to the obese state. In addition, the stimulatory G-protein (Gs) alpha subunit, to which betaARs are coupled, is depressed in obese animals. Since elevated corticosteroid (CORT) levels are an essential feature promoting obesity, and betaAR subtype expression can be modulated by CORTs, regulation of betaAR subtype gene transcription by this steroid may be an important component of obesity. There are 4 major aims of this research plan. (1) Changes in betaAR and Gsalpha will be monitored during the development of obesity. The mRNA levels of the beta1AR, beta2AR beta3AR and Gsalpha genes, radioligand binding and beta-agonist- stimulated adenylyl cyclase activity will be measured in white and brown adipose tissue from lean and obese mice from 1 week to 14 weeks of age. Adipocyte-specific markers will also be monitored. (2) The effect of CORT on expression of betaAR subtypes and Gsalpha genes in brown and white fat, and their effects on obesity will be assessed in 2 ways: (i) Adrenalectomized obese mice will be studied over the same developmental period outlined in Aim 1. (ii) Lean and obese mice will be treated chronically with the CORT antagonist RU38486 over the same period as in Aim 1. Results from Aims 1 and 2 will provide a basis for examining more specific molecular and cellular mechanisms regulating betaAR and Gsalpha gene expression. (3) Established murine preadipocyte cell lines of white and brown fat origin will be used to examine the molecular mechanisms regulating transcription of betaAR subtypes and Gsalpha genes during adipocyte differentiation and CORT exposure. Nuclear run-on transcription, mRNA half-life, and cycloheximide sensitivity studies will determine specific molecular mechanisms responsible for observed changes in individual betaAR and Gsalpha transcripts. These data will be related to results obtained from lean and obese mice in vivo in Aims 1 and 2. (4) Transfection studies with betaAR promoter-reporter constructs in pre- and differentiated adipocytes are designed to identify regulatory elements involved in adipocyte differentiation and CORT regulation of betaAR transcription. Together, these integrated studies should more directly define the physiological, cellular and molecular mechanisms regulating betaAR signal transduction in adipocytes during the development of obesity.

先天性肥胖，β-肾上腺素能受体（betaAR）刺激 脂肪分解和产热作用受损。 有白色和棕色两种 脂肪细胞表达 beta1-、beta2- 和 beta3AR（后者表达 仅存在于脂肪中）。 我们的初步数据表明 beta1AR 和 beta3AR 在遗传性肥胖的白色和棕色脂肪细胞中受损 (ob/ob) 小鼠与瘦对照小鼠相比。 此缺陷可能会导致 肥胖状态。 此外，刺激性 G 蛋白 (Gs) α 与 betaAR 偶联的亚基在肥胖动物中受到抑制。 由于皮质类固醇 (CORT) 水平升高是一个重要特征 促进肥胖，并且 betaAR 亚型表达可以通过调节 CORT，通过该类固醇调节 betaAR 亚型基因转录 可能是肥胖的重要组成部分。 此举有4个主要目标 研究计划。 (1) 监测betaAR和Gsalpha的变化 在肥胖的发展过程中。 beta1AR 的 mRNA 水平， beta2AR beta3AR 和 Gsalpha 基因、放射性配体结合和 β-激动剂- 刺激的腺苷酸环化酶活性将以白色和棕色测量 来自 1 周至 14 周龄瘦小鼠和肥胖小鼠的脂肪组织。 还将监测脂肪细胞特异性标记物。 (2) 效果 CORT 对棕色和棕色中 betaAR 亚型和 Gsalpha 基因表达的影响 白色脂肪及其对肥胖的影响将通过两种方式进行评估：（i） 肾上腺切除的肥胖小鼠将在相同的发育过程中进行研究 目标 1 中概述的时期。 (ii) 将治疗瘦小鼠和肥胖小鼠 在同一时期长期使用 CORT 拮抗剂 RU38486 目标 1. 目标 1 和 2 的结果将为检查更多内容提供基础 调节 betaAR 和 Gsalpha 的特定分子和细胞机制 基因表达。 (3)白色小鼠前脂肪细胞系的建立 棕色脂肪来源将用于检查分子机制 调节 betaAR 亚型和 Gsalpha 基因的转录 脂肪细胞分化和 CORT 暴露。 核连续运行 转录、mRNA 半衰期和放线菌酮敏感性研究将 确定导致观察到的变化的具体分子机制 在单独的 betaAR 和 Gsalpha 转录本中。 这些数据将是相关的 目标 1 和 2 中从瘦小鼠和肥胖小鼠体内获得的结果。 (4) 用 betaAR 启动子-报告基因构建体进行预转染研究 和分化的脂肪细胞被设计来识别调节 参与脂肪细胞分化和 CORT 调节的元件 βAR 转录。 总之，这些综合研究应该更多 直接定义生理、细胞和分子机制 调节脂肪细胞中的βAR信号转导 肥胖的发展。