EARLY EVENTS IN RENAL DEVELOPMENT

当输尿管芽侵入肾原细胞时， 间充质并开始分泌间充质的生存因子， 诱导其分化为肾单位的上皮细胞。我们有 产生了一组单克隆抗体， 间充质，并发现其中两种抗体抑制肾脏 发展 第一种抗体阻断了小管形成， 诱导间充质细胞凋亡。我们纯化了它的抗原gp- 100在诱导的间充质中表达的膜蛋白，但其 在成年后，表达逐渐消失。该蛋白的cDNA显示， 是一个新的基因，具有重要的细胞外和细胞内 信号转导结构域在上皮发育期间， 表达在病灶交界处的基底表面。我们的首要目标是 完成它的cDNA序列，检查它形成的连接类型，克隆 小鼠和人类同源物，并确定其染色体位置。 使用小鼠基因组克隆，我们将删除该基因并产生 突变小鼠并检查其表型。在生物化学研究中，我们将 纯化其胞外配体。 第二种抗体识别细胞外基质蛋白， 最初由输尿管芽和诱导间充质产生，但后来 仅限于系膜和平滑肌细胞。述抗体 抑制输尿管芽周围内皮细胞的定位。我们 纯化并克隆了该抗体的46 kDa抗原， 产生新的多克隆抗体。该蛋白质以前被 克隆作为侵袭性癌的标志物，包括肺癌、乳腺癌和 结肠 在我们的第二个目标中，我们将开发一种功能测定法， 内皮细胞和系膜细胞之间的相互作用， 这种蛋白质的迁移，毛细血管网络的形成， 血管生成 第三个目标是建立输尿管的三维模型， 基于共聚焦扫描的芽分支和肾形态发生 显微镜我们将首先培育出转基因小鼠， 荧光蛋白的表达。这种侵入性较小的 这种方法将使我们能够识别关键的新形态结构， 以及已知的诱导因子及其抑制剂在 肾小管三维结构的产生 以及它们与个体供应的关系。

The kidneys develops when the ureteric bud invades the nephrogenic mesenchyme and begins to secrete survival factors for the mesenchyme and induces it to differentiate into the epithelial of the nephron. We have generated a panel of monoclonal antibodies to induced metanephric mesenchyme and found that two of these antibodies inhibit kidney development. The first antibody blocks tubulogenesis and produces widespread apoptosis of the induced mesenchyme. We have purified its antigen, gp- 100 a membrane protein that is expressed in induced mesenchyme but whose expression fades in adult life. The cDNA of this protein shows that it is a novel gene that has important extracellular and intracellular signal transduction domains. During epithelial development it is expressed in the basal surface in a focal junction. Our First Aim is to complete its cDNA sequence, examine the type of junction it forms, clone the mouse and human homologues and identify its chromosomal location. Using the mouse genomic clone we will delete the gene and generate mutant mice and examine their phenotype. In biochemical studies, we will purify its extracellular ligand. The second antibody recognized an extracellular matrix protein that is initially produced by the ureteric bud and induced mesenchyme but later is restricted to mesangial and smooth muscle cells. The antibody inhibited localization of endothelial cells around the ureteric bud. We have purified and cloned the 46 kDa antigen for this antibody and generated new polyclonal antibodies. The protein has previously been cloned as a marker for aggressive carcinomas including lung, breast and colon. In our Second Aim, we will develop a functional assay for interaction between endothelial and mesangial cells to examine the role of this protein in migration, capillary network formation and angiogenesis. In the Third Aim, we will develop a three dimensional model of ureteric bud branching and renal morphogenesis based on confocal scanning microscopy. We will first generate transgenic mice that express gene fluorescent protein in their ureteric bud lineage. This less invasive approach will allow us to identify critical new morphological structures and the role of known inducing factors and their inhibitors in the generation of the three dimensional architecture of the kidney tubules and their relation to their individual body supply.