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.

发育中的肾脏的收集系统是通过输尿管芽的分支形态发生。大约100万由于多达20次迭代，诱导肾单位形成该结构的二分分支。因此，分支形态发生发育中的尿收集系统直接影响最终成人肾脏中包含的肾单位数。由于肾单位减少质量可能是对易感性和肾脏疾病的临床过程，分支机构的分子机制形态发生具有相当大的临床和基本兴趣。在此外，多达10％的人口具有一定的发展尿路异常，其中很大一部分涉及收集系统。在三维胶原蛋白基质中生长的MDCK和IMCD细胞是理想的体外模型系统，用于鉴定分子决定因素发育中的肾脏中的微管发生和分支。暴露于肝细胞生长因子（HGF），胶原蛋白凝胶形式的MDCK细胞分支管状结构。另一方面，IMCD单元格形成分支响应EGF和TGF-Alpha以及HGF的管状结构。赞助商实验室的初步数据，同时又采用了MDCK和 IMCD细胞模型表明细胞外基质（ECM）组件和胚胎肾脏中存在的基质降解蛋白酶很关键这些细胞中分支的决定因素。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。该方法将是结合基板-SDS-PAGE的组合分析和北方分析以识别候选形态发生蛋白酶和TIMP显示出反应HGF的表达改变的蛋白酶和TIMP MDCK和IMCD单元格中的EGF和TGF-Alpha，也显示胚胎肾脏表达的时空模式通过原位杂交与在收集形成中的作用一致系统。确定了可能的形态发生蛋白酶，其功能将在两种培养物中使用反义寡核苷酸的破坏分支的细胞模型和胚胎肾脏的器官培养通过反义RNA在MDCK和IMCD中产生转染构建体细胞模型。赞助商的实验室是理想的环境执行本提案中描述的工作。赞助商是公认的上皮分支形态发生的领导者在所有方面具有专业知识完成这些研究所需的技术。实验室很好装备和肾部门的学术环境哈佛医学院妇女医院的肾脏非常强大上皮生物学。

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

Multiple fibroblast growth factors support growth of the ureteric bud but have different effects on branching morphogenesis.

多种成纤维细胞生长因子支持输尿管芽的生长，但对分支形态发生有不同的影响。