Functional molecules on glomerular epithelial cells

波士顿大学萨兰特博士大堂培养的肾小球上皮细胞不能被单抗5-1-6染色，用波士顿实验室制备的大鼠肾脏cDNA文库克隆目的抗原分子未成功。用体外制备和纯化的单抗5-1-6与溶解的大鼠肾小球亲和层析柱重复获得抗原分子的尝试是徒劳的。在S形体阶段，这种抗原首先在发育中的足细胞的基底部和侧边变得微弱但明显。染色强度随成熟程度的加深而增强，仅限于内脏上皮细胞。在免疫电子显微镜下，抗原在早期的毛细血管闭塞连接下方的基底面和侧面处可见，后来随着足突的交错，抗原集中在裂隙孔内。在根尖表面均未见抗原。代谢标记研究表明，抗原性分子在肾小球发育初期是活跃合成的，随着发育成熟，其合成速度显著下降。因此，我们认为从新生大鼠肾小球中制备的cDNA库更适合于克隆这种抗原分子。我们正计划利用婴儿肾小球的cDNA文库来克隆这个基因。

Cultured glomerular epithelial cells in the loboratory of Dr. Salant in Boston University could not be stained by the monoclonal antibody (mAb) 5-1-6.Cloning of the target antigenic molecule has not been succeeded, using the rat kidney cDNA library prepared in the Boston laboratory.The traial to get the antigenic molecule has been repeated in vain, using the affinity column with in vitro prepared- and purified-mAb 5-1-6 and the solublized rat glomeruli.The biogenesis of the target antigen of mAb 5-1-6 was studied in the developing glomerulus by immunolocalization and metabolic labeling. This antigen first became faintly, but clearly, detectable on the basal and lateral sides of the developing podocytes at the S-shaped body stage. Staining intensity increased with further maturation and was restricted to the visceral epithelial cells. On immunoelectro microscopy, the antigen was seen along the basal and lateral surfaces below occluding junctions at the early capillary loop stage and later, with the interdigitation of foot processes, became concentrated in the slit pores. At no stage was the antigen seen on the apical surface. Metaboric labeling studies showed that the antigenic molecule is actively synthesized during initial glomerular development and that the rate of synthesis declines substantially with maturation.From these results cDNA-library was considered to be more poperly prepared from rat glomeruli in neonatal stage for cloning of this antigenic molecule. We are planning to clone this, applying the cDNA library from baby glomeruli.