Chansel D, Bizet T, Vandermeersch S, Pham P, Levy B, Ardaillou R
Institut National de la Santé et de la Recherche Médicale (INSERM) 64, Hôpital Tenon, Paris, France.
Am J Physiol. 1994 Mar;266(3 Pt 2):F384-93. doi: 10.1152/ajprenal.1994.266.3.F384.
The aim of the present report was to examine the effect of several agents on angiotensin II (ANG II) and losartan receptors using 125I-[Sar1,Ala8]ANG II and [3H]losartan as radiolabeled ligand, respectively. ANG II receptors were downregulated in glomeruli from rats infused with ANG II during 3 wk or rats receiving losartan orally during 1 wk. The number of sites (Bmax) was reduced, but the dissociation constant (Kd) value was unchanged. Losartan receptors were downregulated in glomeruli from rats receiving losartan, but remained unchanged in glomeruli from rats infused with ANG II. Since in vivo administration of losartan results in increase of plasma ANG II and formation of metabolites, in vitro studies using human mesangial cells were performed to better analyze the present findings. Treatment of mesangial cells during 4 days by ANG II, losartan, or its metabolite, EXP-3174, also produced downregulation of 125I-[Sar1,Ala8]ANG II binding sites with a decreased Bmax and unchanged Kd value. Only treatment of mesangial cells by ANG II or EXP-3174 produced downregulation of [3H]losartan binding sites. In contrast, exposure of these cells to losartan resulted in upregulation of [3H]losartan binding sites. Under all conditions, only Bmax was modified. Whereas internalization of [3H]losartan in mesangial cells was negligible under all experimental conditions, there was an increase of the percentage of internalized 125I-[Sar1,Ala8]ANG II after exposure of the cells to ANG II or AT1 antagonists. No change was observed in mesangial cell AT1 receptor mRNA levels. This study demonstrates that 1) AT1 mRNA is expressed in human mesangial cells; 2) the characteristics of 125I-[Sar1,Ala8]ANG II and [3H]losartan binding sites in rat glomeruli and human mesangial cells are different, with Kd and Bmax values greater in both preparations when [3H]losartan was utilized; 3) both types of binding sites obey different regulations, and the effects of losartan in vivo are due in part to the associated increase in plasma ANG II levels and the transformation of the drug into its metabolite, EXP-3174; 4) downregulation of AT1 receptors does not depend on changes in mRNA expression but is associated with increased relative internalization.
本报告的目的是分别使用125I-[Sar1,Ala8]血管紧张素II（ANG II）和[3H]氯沙坦作为放射性标记配体，研究几种药物对ANG II和氯沙坦受体的影响。在连续3周输注ANG II的大鼠或连续1周口服氯沙坦的大鼠的肾小球中，ANG II受体下调。位点数量（Bmax）减少，但解离常数（Kd）值不变。在接受氯沙坦的大鼠的肾小球中，氯沙坦受体下调，但在输注ANG II的大鼠的肾小球中保持不变。由于氯沙坦的体内给药导致血浆ANG II增加和代谢产物形成，因此进行了使用人系膜细胞的体外研究以更好地分析当前发现。用ANG II、氯沙坦或其代谢产物EXP-3174处理系膜细胞4天，也会导致125I-[Sar1,Ala8]ANG II结合位点下调，Bmax降低而Kd值不变。只有用ANG II或EXP-3174处理系膜细胞才会导致[3H]氯沙坦结合位点下调。相反，将这些细胞暴露于氯沙坦会导致[3H]氯沙坦结合位点上调。在所有条件下，只有Bmax发生改变。在所有实验条件下，系膜细胞中[3H]氯沙坦的内化可忽略不计，而在细胞暴露于ANG II或AT1拮抗剂后，内化的125I-[Sar1,Ala8]ANG II的百分比增加。系膜细胞AT1受体mRNA水平未观察到变化。本研究表明：1）AT1 mRNA在人系膜细胞中表达；2）大鼠肾小球和人系膜细胞中125I-[Sar1,Ala8]ANG II和[3H]氯沙坦结合位点的特征不同，当使用[3H]氯沙坦时，两种制剂中的Kd和Bmax值更大；3）两种类型的结合位点遵循不同的调节方式，氯沙坦在体内的作用部分归因于血浆ANG II水平的相关增加以及药物转化为其代谢产物EXP-3174；4）AT1受体的下调不依赖于mRNA表达的变化，而是与相对内化增加有关。
