Mechanisms of Renal Cell Injury

肾细胞损伤的机制

基本信息

批准号：
10013592
负责人：
GOUTAM GHOSH CHOUDHURY
金额：
--
依托单位：
SOUTH TEXAS VETERANS HEALTH CARE SYSTEM
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-07-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10013592
关键词：
20 year old Accounting Address Affect Automobile Driving Binding CRISPR/Cas technology Cell Membrane Permeability Cells Chromosome 10 Codon Nucleotides Complex Complications of Diabetes Mellitus Data Development Diabetes Mellitus Diabetic Nephropathy Diabetic mouse Disease Dissociation Down-Regulation End stage renal failure Epithelial Cells Event Exposure to FRAP1 gene Fibronectins Glucose Healthcare Human Hyperglycemia Hypertrophy Immune Immunoblotting Immunofluorescence Immunologic Immunohistochemistry Immunoprecipitation Kidney Kidney Diseases Knock-out Lead Maps Methods Modality Modeling Molecular Mus PTEN gene Pathogenesis Pathologic Pathology Patients Permeability Phosphorylation Phosphotransferases Plasminogen Activator Inhibitor 1 Platelet Activation Platelet-Derived Growth Factor B Platelet-Derived Growth Factor beta Receptor Process Protein Isoforms Protein Tyrosine Kinase Proteins Recombinant Proteins Recombinants Regulation Renal Tissue Renal function Reporting Resources Rodent Rodent Model Role Signal Transduction Small Interfering RNA Streptozocin Techniques Testing Therapeutic Therapeutic Agents Transfection Transforming Growth Factors Translations Tricarboxylic Acids Tubular formation Type 2 diabetic Tyrosine United States Variant Veterans age group cell injury cost db/db mouse demographics diabetic diabetic rat falls inhibitor/antagonist kidney cell kidney cortex mesangial cell military veteran morphometry novel novel therapeutic intervention pandemic disease prevent protein expression response small molecule therapeutic protein transfection/expression vector type I diabetic

项目摘要

Pathologic manifestations of diabetic nephropathy (DN) include glomerular and tubular hypertrophy and matrix protein fibronectin expression. These changes occur concomitant with increased expression of TGFb (transforming growth factor-b) that contributes to the pathogenesis of human and experimental DN. We interrogate the molecular signaling events by which high glucose and TGFb drive the pathologies and provide new potential therapeutic strategies for complications of DN. We have reported that Akt kinase/mTORC1 (mechanistic target of rapamycin complex 1) axis contributes to renal hypertrophy and fibronectin expression in mesangial and proximal tubular epithelial (PTE) cells, and in kidneys of type 1 and type 2 diabetic mice. Recently, a novel longer translational variant of PTEN (phosphatase and tensin homolog deleted in chromosome 10), PTEN-Long, a negative regulator of Akt kinase, has been identified as a secretory and membrane permeable protein. Our preliminary data show markedly reduced levels of PTEN-Long along with increased Akt kinase activity in the kidney cortex of type 1 and type 2 diabetic mice and in high glucose- or TGFb-treated mesangial and PTE cells. Moreover, we show that high glucose and TGFb activate PDGFRb (platelet-derived growth factor-b). We find PDGFRb as a substrate for PTEN-Long; thus downregulation of PTEN-Long results in PDGFRb activation by high glucose and TGFb. Furthermore, in the kidney and in mesangial and PTE cells, we identify Akt-2 as the predominant isotype of Akt kinase that acts downstream of PTEN-Long/PDGFRb. Also, Akt-2 is activated in kidneys of type 1 and type 2 diabetic mice kidneys. We plan to exploit the intrinsic protective function of PTEN-long during the progression of DN. In this proposal, using cultured mesangial and PTE cells and renal tissues from diabetic OVE26 and db/db mice, we will test the hypothesis that hyperglycemia/TGFb-induced inappropriate downregulation of PTEN-Long results in PDGFRb/Akt-2 activation that contributes to renal hypertrophy and matrix expansion in diabetic kidney disease. Probing the novel negative regulatory function of PTEN-Long, we will use it as a therapeutic agent for DN. In the first specific aim, we will determine the role of PTEN-Long in hypertrophy and, fibronectin and PAI-1 (plasminogen-activator inhibitor-1) expression. In the second aim, how PTEN-Long forces activation of PDGFRb to regulate mesangial and PTE cell hypertrophy, and matrix protein expression will be investigated. In specific aim 3, we will study the contribution of Akt-2 isotype to hypertrophy and fibronectin/PAI-1 expression in response to high glucose and TGFb in mesangial and PTE cells and in diabetic mice kidneys. To address these aims, techniques including immunoblotting, immunoprecipitation, morphometry, immunohistochemistry, transfection of expression vectors and siRNAs, administration of recombinant therapeutic protein and small molecule compound will be employed. Our study will establish a network of interconnections involving PTEN- Long, PDGFRb and Akt-2 for driving DN.

糖尿病肾病（DN）的病理表现包括肾小球和肾小管肥大，基质蛋白纤连蛋白表达。这些变化伴随着 TGFb 表达的增加而发生（转化生长因子-b）有助于人类和实验性 DN 的发病机制。我们探究高葡萄糖和 TGFb 驱动病理的分子信号传导事件并提供 DN 并发症的新潜在治疗策略。我们报道了 Akt 激酶/mTORC1 （雷帕霉素复合物 1 的机械靶标）轴有助于肾肥大和纤连蛋白表达系膜和近端肾小管上皮 (PTE) 细胞，以及 1 型和 2 型糖尿病小鼠的肾脏。最近，PTEN 的一种新的较长翻译变体（磷酸酶和张力蛋白同源物在染色体 10），PTEN-Long，Akt 激酶的负调节因子，已被确定为分泌和膜渗透性蛋白质。我们的初步数据显示 PTEN-Long 水平显着降低 1 型和 2 型糖尿病小鼠以及高血糖或高血糖小鼠肾皮质中的 Akt 激酶活性增加 TGFb 处理的系膜细胞和 PTE 细胞。此外，我们发现高葡萄糖和 TGFb 会激活 PDGFRb （血小板衍生生长因子-b）。我们发现 PDGFRb 作为 PTEN-Long 的底物；从而下调 PTEN-Long 导致高葡萄糖和 TGFb 激活 PDGFRb。此外，在肾脏和系膜细胞和 PTE 细胞，我们将 Akt-2 确定为 Akt 激酶的主要同种型，作用于下游 PTEN-长/PDGFRb。此外，Akt-2 在 1 型和 2 型糖尿病小鼠肾脏中被激活。我们计划在 DN 进展过程中利用 PTEN-long 的内在保护功能。在本提案中，使用培养糖尿病 OVE26 和 db/db 小鼠的系膜细胞和 PTE 细胞以及肾组织，我们将测试假设高血糖/TGFb 诱导 PTEN-Long 的不适当下调导致 PDGFRb/Akt-2 激活导致糖尿病肾脏肾肥大和基质扩张疾病。探索 PTEN-Long 的新型负调节功能，我们将用它作为治疗剂 DN。在第一个具体目标中，我们将确定 PTEN-Long 在肥大中的作用，以及纤连蛋白和 PAI-1 （纤溶酶原激活剂抑制剂-1）表达。在第二个目标中，PTEN-Long 如何强制激活将研究 PDGFRb 调节系膜和 PTE 细胞肥大以及基质蛋白表达的作用。在具体目标 3 中，我们将研究 Akt-2 同种型对肥大和纤连蛋白/PAI-1 表达的贡献系膜细胞和 PTE 细胞以及糖尿病小鼠肾脏中对高葡萄糖和 TGFb 的反应。致地址这些目标和技术包括免疫印迹、免疫沉淀、形态测定、免疫组织化学、表达载体和 siRNA 的转染、重组治疗蛋白和小分子的施用将采用分子化合物。我们的研究将建立一个涉及 PTEN 的互连网络 Long、PDGFRb 和 Akt-2 用于驱动 DN。