Cell Cycle Dependent Mechanisms of Renal Hypertrophy

肾肥大的细胞周期依赖性机制

• 批准号: 6466738
• 负责人: DANIEL J RILEY
• 金额: $ 21.08万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-05-15 至 2007-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6466738
• 关键词: AP1 protein; cell cycle; cell growth regulation; cyclin dependent kinase; diabetic nephropathy; gel mobility shift assay; gene expression; genetic regulation; genetically modified animals; glomerulosclerosis; glucose; immunoprecipitation; insulin dependent diabetes mellitus; kidney cell; kidney hypertrophy; laboratory mouse; mesangium; phosphorylation; protooncogene; retinoblastoma protein; site directed mutagenesis; western blottings

DESCRIPTION (Provided by applicant): Early in diabetic nephropathy glomerular mesangium and renal tubules hypertrophy. Mesangial hypertrophy is an important, perhaps reversible, step preceding glomeruloscierosis and overt nephropathy. Using unique transgenic mice overexpressing retinoblastoma protein (Rb) and mesangial cells expressing Rb from a tetracycline-confrollable promoter system, we have discovered that renal and glomerular mesangial cell hypertrophy occur with both diabetes mellitus and with over expression of hypophosphorylated Rb. Preliminary data has shown that when Rh is over expressed, neither kidneys nor mesangial cells hypertrophy further in diabetic conditions. Our hypothesis to explain these observations is that excess glucose results in specific, cyclin-dependent phosphorylation of Rb protein during early Gi phase, and that Rb is involved distally in a pathway of glomerular mesangial cell hypertrophy. To test this hypothesis, three Specific Aims are proposed: (1) Determine whether renal and glomerular hypertrophy caused by diabetes mellitus in vivo and mesangial cell hypertrophy caused by high glucose concentrations ex vivo are dependent on activation of specific C1 phase cyclin-cdk complexes and specific phosphorylations of Rb protein. (2) Determine how high glucose regulates cdk4-cyclin Dl activity and Rb phosphorylation in mesangial cells by examining patterns of gene regulation including transcription of the cyclin D1 gene itself and of events controlled by relevant transcription factors including AP-1. (3) Test the requirement for Cl cyclin dependent kinase activation and specific cdk4-dependent phosphorylations of Rb protein for diabetic renal hypertrophy in vivo and mesangial cell hypertrophy in culture. Transgenic mice with strategic phosphorylation sites in Rb inactivated by site-directed mutagenesis will be generated and the effects of type 1 diabetes mellitus will be examined in them. Primary mesangial cells will also be cultured from the mice and more detailed studies on hypertrophy and GI cell cycle regulation by cdks and Rb conducted in vitro.

描述（由申请人提供）：糖尿病肾病肾小球早期 系膜和肾小管肥大。系膜肥大是一个重要的 也许是可逆的，先于肾小球硬化和明显的肾病。 使用独特的转基因小鼠过度表达视网膜母细胞瘤蛋白（Rb）和 从四环素可控启动子系统表达 Rb 的系膜细胞， 我们发现肾脏和肾小球系膜细胞发生肥大 患有糖尿病和低磷酸化 Rb 过度表达。 初步数据表明，当 Rh 过度表达时，肾脏和 在糖尿病情况下，系膜细胞进一步肥大。我们的假设是 解释这些观察结果的是，过量的葡萄糖会导致特定的、 早期 Gi 期 Rb 蛋白依赖于细胞周期蛋白的磷酸化，并且 Rb 在远端参与肾小球系膜细胞肥大的途径。 为了检验这一假设，提出了三个具体目标： (1) 确定 体内是否有糖尿病引起的肾及肾小球肥大 离体高浓度葡萄糖引起的肾小球系膜细胞肥大 依赖于特定 C1 期细胞周期蛋白-cdk 复合物的激活， Rb 蛋白的特异性磷酸化。 (2)确定血糖有多高 通过调节系膜细胞中的 cdk4-cyclin Dl 活性和 Rb 磷酸化 检查基因调控模式，包括细胞周期蛋白 D1 的转录 基因本身以及相关转录因子控制的事件 包括AP-1。 (3)测试C1细胞周期蛋白依赖性激酶的要求 Rb 蛋白的激活和特异性 cdk4 依赖性磷酸化 体内糖尿病肾肥大和培养物肾小球系膜细胞肥大。 Rb 中具有策略性磷酸化位点的转基因小鼠被灭活 将产生定点突变并对 1 型糖尿病产生影响 将在其中进行检查。原代系膜细胞也将 从小鼠中培养，并对肥大和胃肠道细胞进行更详细的研究 cdks 和 Rb 的循环调节在体外进行。