Mechanisms of Renal Tubulogenesis

肾小管发生机制

基本信息

批准号：
7544935
负责人：
Keith E Mostov
金额：
$ 31.67万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-01-01 至 2010-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7544935
关键词：
3-Dimensional 4-Hydroxy-Tamoxifen Abbreviations Acute Kidney Failure Acute Kidney Tubular Necrosis Affect Binding Biochemical Biological Assay Biological Models Canis familiaris Cell Culture Techniques Cell Line Cell surface Cells Cholera Toxin Collagen Complex Conditioned Culture Media Confocal Microscopy Cyst Defect Development Disease Dominant-Negative Mutation Doxycycline Epithelial Epithelial Cells Estrogen Receptor 1 Extracellular Matrix Family Fibrosis Four-dimensional GTPase-Activating Proteins Gel Genes Growth Factor Guanine Nucleotide Exchange Factors Guanosine Triphosphate Phosphohydrolases Health Hepatocyte Growth Factor Hereditary Disease Immunofluorescence Immunologic Injury Kidney Kidney Diseases Kinetics Learning MAP Kinase Gene MAP Kinase Modules Matrix Metalloproteinases Membrane Microdomains Mesenchymal Microtubules Mitogen-Activated Protein Kinases Molecular Myosin Light Chains Natural regeneration Organ Pathway interactions Phosphatidylinositols Phosphotransferases Physiological Process RNA Interference Ras/Raf Receptor Protein-Tyrosine Kinases Recovery Renal tubule structure Research Personnel Rho-associated kinase Role Signal Pathway Signal Transduction Staging System Testing Thick Tight Junctions Time Tube Video Microscopy Work cell type in vivo kidney cell meetings methyl-beta-cyclodextrin monolayer nephrogenesis phosphoinositide-3,4,5-triphosphate phosphoinositide-3,4-bisphosphate prevent programs rho small hairpin RNA stem time use two-dimensional

项目摘要

The kidney, like many epithelial organs, consists mainly of tubules lined by a monolayer of polarized epithelial cells. Though we have learned a great deal about the genes that control kidney development, we know much less about the mechanisms by which cells rearrange themselves into tubules. Weuse a simplified model system of Madin-Darbycanine kidney (MDCK) cells grown in a 3 dimensionalcollagen gel. Single MDCK cellsgrow to form hollow cysts lined by a monolayer of epithelial cells. When these cysts are stimulated with Hepatocyte Growth Factor (HGF), they form tubules over a period of ~3 days. This is a good model system for studying renal tubule formation, and can also provide informationon several pathophysiological processes, such as recovery from acute tubular necrosis, renal fibrosis, polycystickidney disease and perhaps renal regeneration from stem cells.We will study the signaling pathways by which cells respond to HGF. The kinetics of activation of ERKsuggest that it is activated by a MAPK cascadeinvolving B-Raf, rather than the better known Raf-1. We will test this hypothesis by studying activation of components of the B-Raf pathway and by using dominant negatives and RNAi. We will test the involvement of Rho family GTPases, Rho Kinase and lipid rafts in the early stages of tubulogenesis, especially the formation of extensions from the basolateral surface of cells in cysts. We will use time lapse confocal microscopy to observe the effects of perturbing these molecular components on extension formation.We will test the involvement of phosphatidyl inositol (3,4,5)P3 [PI(3,4,5)P3]in extension formation. We have found that exogenous PI(3,4,5)P3 induces extension formation and we will test if these extensions are identical to those produced by HGF. We will analyze possible effectors of PI(3,4,5)P3. Kidney diseases, including developmental defects, kidney injury and genetic conditions that form multiple cysts in the kidney, are major health problems. The kidney consists of many narrow tubes lined by cells. We are using a simple model system where kidney cells are grown in culture and produce such tubes, to study tube formation and how we can influence this to treat and prevent kidney diseases.

与许多上皮器官一样，肾脏主要由排列有单层极化的肾小管组成。上皮细胞。尽管我们对控制肾脏发育的基因了解很多，但我们对细胞重新排列成小管的机制了解甚少。我们使用一个在 3 维胶原蛋白中生长的 Madin-Darbycanine 肾 (MDCK) 细胞的简化模型系统凝胶。单个 MDCK 细胞生长形成空心囊肿，内衬单层上皮细胞。当这些囊肿受到肝细胞生长因子 (HGF) 的刺激，它们会在约 3 天的时间内形成小管。这是一个研究肾小管形成的良好模型系统，还可以提供一些信息病理生理过程，例如从急性肾小管坏死、肾纤维化、多囊肾中恢复疾病，也许还有干细胞的肾再生。我们将研究细胞的信号传导途径对 HGF 做出反应。 ERK 激活动力学表明它是由涉及 MAPK 级联的激活 B-Raf，而不是更为人所知的 Raf-1。我们将通过研究组件的激活来检验这个假设 B-Raf 途径并通过使用显性失活和 RNAi。我们将测试 Rho 的参与情况 GTPase 家族、Rho 激酶和脂筏在肾小管发生的早期阶段，尤其是形成囊肿细胞基底外侧表面的延伸。我们将使用延时共焦显微镜观察扰动这些分子成分对延伸形成的影响。我们将测试磷脂酰肌醇 (3,4,5)P3 [PI(3,4,5)P3] 参与延伸形成。我们发现外源 PI(3,4,5)P3 诱导延伸形成，我们将测试这些延伸是否与那些相同由HGF生产。我们将分析 PI(3,4,5)P3 可能的效应器。肾脏疾病，包括发育缺陷、肾损伤和形成多种遗传性疾病肾脏囊肿是主要的健康问题。肾脏由许多排列有细胞的狭窄管组成。我们正在使用一个简单的模型系统来研究肾细胞在培养物中生长并产生这样的管管的形成以及我们如何影响它来治疗和预防肾脏疾病。