PROTEINASES IN KIDNEY EPITHELIAL BRANCHING MORPHOGENESIS

肾上皮分支形态发生中的蛋白酶

基本信息

批准号：
6176831
负责人：
ROBERT O STUART
金额：
$ 11.64万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
1996
资助国家：
美国
起止时间：
1996-07-01 至 2001-06-30
项目状态：
已结题

项目摘要

The collecting system of the developing kidney is generated through branching morphogenesis of the ureteric bud. Approximately 1 million nephrons are induced to form as the result of as many as 20 iterations of dichotomous branching of this structure. Thus, branching morphogenesis in the developing urinary collecting system directly influences the eventual number of nephrons contained in the adult kidney. Since, reduced nephron mass may be critical determinant of both the susceptibility to and clinical course of renal disease, molecular mechanisms of branching morphogenesis are of considerable clinical and basic interest. In addition, as much as 10% of the population possess some developmental abnormality of the urinary tract with a large percentage of these involving the collecting system. MDCK and IMCD cells grown in a three-dimensional collagen matrix are ideal in vitro model systems for the identification of molecular determinants of tubulogenesis and branching in the developing kidney. When exposed to hepatocyte growth factor (HGF), MDCK cells in collagen-I gels form branching tubular structures. On the other hand, IMCD cells form branching tubular structures in response to EGF and TGF-alpha, as well as HGF. Preliminary data from the sponsor's laboratory employing both the MDCK and IMCD cell models indicates that extracellular matrix (ECM) components and matrix degrading proteinases present in the embryonic kidney are critical determinants of branching in these cells. The purpose of the work described in this proposal is to identify critical morphogenetic ECM degrading matrix metalloproteinases (MMPs), serine proteases, and tissue inhibitors of MMPs involved in branching morphogenesis of the ureteric bud. The approach will be to use a combination of substrate-SDS-PAGE analysis and Northern analysis to identify candidate morphogenetic proteinases and TIMPs which show altered expression in response to HGF, EGF, and TGF-alpha in the MDCK and IMCD cell models and which also display spatiotemporal patterns of expression in the embryonic kidney by in situ hybridization consistent with a role in formation of the collecting system. Having identified likely morphogenetic proteinases, their function will be disrupted using antisense oligonucleotides in both the cultured cell models of branching and in organ culture of the embryonic kidney and by antisense RNA producing transfection constructs in the MDCK and IMCD cell models. The Sponsor's laboratory is the ideal environment in which to carry out the work described in this proposal. The Sponsor is a recognized leader in epithelial branching morphogenesis with expertise in all of the techniques required to complete these studies. The laboratory is well equipped and the academic environment of the Renal Division at Brigham and Women's Hospital, Harvard Medical School is extremely strong in renal epithelial biology.

发育中肾脏的收集系统是通过以下方式产生的输尿管芽的分支形态发生。约100万经过多达 20 次迭代后，诱导形成肾单位该结构的二分分支。因此，分支形态发生泌尿收集系统的发育直接影响最终的结果成人肾脏中所含肾单位的数量。因为，肾单位减少质量可能是敏感性和敏感性的关键决定因素肾脏疾病的临床过程、分支的分子机制形态发生具有相当大的临床和基础意义。在此外，多达 10% 的人口具有一定的发育能力尿路异常，其中很大一部分是这样的涉及到采集系统。在三维胶原蛋白基质中生长的 MDCK 和 IMCD 细胞是理想的选择用于鉴定分子决定因素的体外模型系统发育中肾脏的肾小管发生和分支。当暴露于肝细胞生长因子 (HGF)、MDCK 细胞以 I 型胶原凝胶形式存在分支管状结构。另一方面，IMCD细胞形成分支管状结构对 EGF 和 TGF-α 以及 HGF 作出反应。来自申办者实验室使用 MDCK 和 IMCD 细胞模型表明细胞外基质 (ECM) 成分和胚胎肾脏中存在的基质降解蛋白酶至关重要这些细胞分支的决定因素。工作目的该提案中描述的是确定关键的形态发生 ECM 降解基质金属蛋白酶 (MMP)、丝氨酸蛋白酶和组织参与输尿管分支形态发生的 MMP 抑制剂芽。该方法将结合使用底物-SDS-PAGE 分析和 Northern 分析以确定候选形态发生蛋白酶和 TIMP 在 HGF 的作用下表现出表达改变， MDCK 和 IMCD 细胞模型中的 EGF 和 TGF-α 也显示胚胎肾脏中原位表达的时空模式杂交与收集形成中的作用一致系统。鉴定出可能的形态发生蛋白酶后，它们的功能将在培养的两个细胞中使用反义寡核苷酸进行破坏胚胎肾的分支和器官培养的细胞模型通过在 MDCK 和 IMCD 中产生转染构建体的反义 RNA 细胞模型。申办者的实验室是进行研究的理想环境开展本提案中所述的工作。赞助商是公认的上皮分支形态发生领域的领导者，拥有所有领域的专业知识完成这些研究所需的技术。实验室很好布莱根医学院肾脏科的设备和学术环境哈佛医学院女子医院肾病极强上皮生物学。

项目成果

期刊论文数量（7）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Rival penalized competitive learning (RPCL): a topology-determining algorithm for analyzing gene expression data.

竞争对手惩罚竞争学习 (RPCL)：一种用于分析基因表达数据的拓扑确定算法。

DOI：
10.1016/j.compbiolchem.2003.09.006
发表时间：
2003
期刊：
Computational biology and chemistry
影响因子：
3.1
作者：
Nair,TMurlidharan;Zheng,ChristinaL;Fink,JLynn;Stuart,RobertO;Gribskov,Michael
通讯作者：
Gribskov,Michael

Evolution of gene expression patterns in a model of branching morphogenesis.

分支形态发生模型中基因表达模式的演变。