In vivo analysis of the regulation of renin production

我们研究了血管紧张素（A）Ⅱ在两个不同系统中的局部作用与全身作用。首先，我们用AII受体（AT 1）缺陷和完整细胞制备嵌合小鼠。持续输注AII可诱导肾小球细胞增殖。当ATI缺陷和完整的肾小球进行比较，在一个给定的嵌合小鼠，两种类型的肾小球被发现containe增殖细胞的数量相似。另一方面，当用ATI反义寡核苷酸处理Thyl肾炎大鼠的一侧肾脏时，仅在处理的肾脏中抑制系膜细胞的增殖，而在对侧肾脏中不抑制系膜细胞的增殖。这表明AII单独的局部作用对细胞增殖没有影响，但它与组织损伤中激活的其他生长因子协同合作。我们计划建立一个转基因小鼠系统，在体内分析肾素生产的调节。5.5将小鼠肾素基因（Renld）的5'侧翼区的kb DNA片段与具有或不具有突变的报告基因（lacZ或EGFP）组合，并导入小鼠ES细胞。这些转基因被设计为包括loxP序列，使得它们被掺入小鼠基因组的相同位点。将ES细胞注射到胚泡中以产生嵌合小鼠，其中在各种条件下分析报告基因表达。主要的技术问题是DNA的多次转染常常使ES细胞分化。该项目目前正在进行中，以克服这一问题。

We studied the role of local vs. systemic action of angiotensin (A)II in two different system. First, we generated chimeric mice made up with AII receptor (AT1) deficient and intact cells. Continuous infusion of AII induced proliferation of glomerular cells. When ATI deficient and intact glomeruli were compared within a given chimeric mouse, both types of glomeruli were found to containe similar number of proliferated cells. On the other hand, when one side of the kidney was treated with ATI antisense oligo-DNA in rats with Thyl nephritis, proliferation of mesangial cell were decreased only in the treated kidney, but not in the contra-lateral kidney. These indicate that the local action of AII alone does not have an impact on the cell proliferation, but it synergistically cooperates with other growth factors activated in tissue injury. We plan to establish a transgenic mouse system in which the regulation of renin production is analyzed in vivo. 5.5 kb DNA fragment of 5' flanking region of the mouse renin gene (Renld) was combined with reporter genes (lacZ or EGFP) with or without mutations, and introduced into mouse ES cells. These transgenes were designed to include loxP sequences so that they are incorporated into the same site of the mouse genome. The ES cells will be injected into blastocyst to generate chimeric mice, in which the reporter gene expression is analyzed under various conditions. The major technical problem is that multiple transfection of DNA often differentiated ES cells. The project is now ongoing to overcome this problem.