Wang Liming, Flannery Patrick J, Spurney Robert F
Division of Nephrology, Department of Medicine, Duke University Medical Center and Durham Veterans Affairs Medical Center, NC 27710, USA.
J Lab Clin Med. 2003 Nov;142(5):313-21. doi: 10.1016/S0022-2143(03)00139-2.
Glomerular podocytes play a key role in maintaining the integrity of the glomerular filtration barrier. This function may be regulated by angiotensin II (Ang II) through activation of cell-surface receptors. Although studies suggest that podocytes express receptors for Ang II, the Ang II binding site has not been characterized with radioligand binding techniques. We therefore used iodine 125-labeled Ang II to monitor Ang II-receptor density during differentiation of a mouse podocyte cell line. Scatchard analyses of equilibrium binding data revealed a single class of high-affinity binding sites (dissociation constant approximately 3 nmol/L) in both differentiated and nondifferentiated cells. During differentiation, the density of Ang II-receptor sites increased roughly 15-fold in differentiated podocytes (maximal density of specific binding sites 881 fmol/mg protein) compared with that in nondifferentiated cells (52 fmol/mg protein; P<.005). Glomerular podocytes expressed messenger RNA for AT1A, AT1B, and AT2 receptor subtypes, and competitive binding studies found that differentiated podocytes expressed mostly AT1 receptors (approximately 75%) with lesser amounts of AT2 (approximately 25%). Up-regulation of Ang II-receptor number was associated with increased Ang II-receptor responsiveness, as evidenced by enhanced Ang II-stimulated inositol phosphate (IP) generation and incorporation of tritiated thymidine. Both [3H]thymidine incorporation and IP generation were mediated by AT1-receptor activation. These data suggest that glomerular podocytes express a high-affinity binding site for Ang II with pharmacologic characteristics of both AT1 and AT2 receptors. This receptor site is up-regulated during podocyte differentiation, and receptor activation induces both IP generation and DNA synthesis by AT1-dependent mechanisms. We speculate that activation of podocyte Ang II receptors contributes to glomerular damage in disease states.
肾小球足细胞在维持肾小球滤过屏障的完整性方面发挥着关键作用。该功能可能受血管紧张素II（Ang II）通过激活细胞表面受体来调节。尽管研究表明足细胞表达Ang II受体，但Ang II结合位点尚未用放射性配体结合技术进行表征。因此，我们使用125碘标记的Ang II来监测小鼠足细胞系分化过程中Ang II受体密度。对平衡结合数据的Scatchard分析显示，在分化和未分化细胞中均存在一类高亲和力结合位点（解离常数约为3 nmol/L）。在分化过程中，与未分化细胞（52 fmol/mg蛋白）相比，分化的足细胞中Ang II受体位点密度增加了约15倍（特异性结合位点的最大密度为881 fmol/mg蛋白；P<0.005）。肾小球足细胞表达AT1A、AT1B和AT2受体亚型的信使RNA，竞争性结合研究发现，分化的足细胞主要表达AT1受体（约75%），而AT2受体表达量较少（约25%）。Ang II受体数量的上调与Ang II受体反应性增加有关，这表现为Ang II刺激的肌醇磷酸（IP）生成增加和氚标记胸腺嘧啶掺入增加。[3H]胸腺嘧啶掺入和IP生成均由AT1受体激活介导。这些数据表明，肾小球足细胞表达具有AT1和AT2受体药理学特征的Ang II高亲和力结合位点。该受体位点在足细胞分化过程中上调，受体激活通过AT1依赖机制诱导IP生成和DNA合成。我们推测，足细胞Ang II受体的激活在疾病状态下促成肾小球损伤。
