Megalin binds and internalizes angiotensin-(1-7)
Megalin binds and internalizes angiotensin-(1-7)
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DOI：
10.1152/ajprenal.00164.2005
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发表时间：
2006-05-01
影响因子：
4.2
通讯作者：
Hammond, TG
中科院分区：
文献类型：
--
作者：
Gonzalez-Villalobos, R;Klassen, RB;Hammond, TG
巨蛋白是一种多配体受体，与蛋白质内吞作用密切相关。我们最近证实，巨蛋白结合并介导Ang II内化。虽然Ang II和Ang-(1-7)在结构上有很大的相似之处，但它们的生理作用和对血管紧张素1型受体(AT(1)R)的亲和力是不同的。因此，本工作的假设是测试megalin是否结合和内化Ang-(1-7)。用荧光标记的Ang-(1-7)(100 NM)处理BN/MSV细胞融合单层细胞(代表卵黄囊上皮细胞)，并用流式细胞仪测定4h后细胞相关荧光量，以确定Ang-(1-7)的摄取。抗megalin抗血清和AT1R阻断剂(奥美沙坦)分别干扰通过megalin和AT1R的摄取。抗大球蛋白抗血清(63%)对Ang(1-7)摄取的抑制程度高于奥美沙坦(13%)(P<；0.001)。用流式细胞仪对CD-1小鼠肾脏刷状缘膜囊泡结合实验进行分析时，抗巨球蛋白抗血清对Ang-(1-7)结合的干扰作用强于奥美沙坦(P<；0.05与阳性对照)。用表面等离子体共振技术研究了Meggalin与Ang-(1-7)在分子水平上的相互作用，结果表明，Ang-(1-7)与Meggalin的结合具有剂量和时间依赖性，亲和力与Ang II相似。
Megalin is a multiligand receptor heavily involved in protein endocytosis. We recently demonstrated that megalin binds and mediates internalization of ANG II. Although there is a strong structural resemblance between ANG II and ANG-(1-7), their physiological actions and their affinity for the angiotensin type 1 receptor (AT(1)R) are dissimilar. Therefore, the hypothesis of the present work was to test whether megalin binds and internalizes ANG-(1-7). The uptake of ANG-(1-7) was determined by exposure of confluent monolayers of BN/MSV cells ( a model representative of the yolk sac epithelium) to fluorescently labeled ANG-(1-7) (100 nM) and measurement of the amount of cell-associated fluorescence after 4 h by flow cytometry. Anti-megalin antisera and an AT1R blocker ( olmesartan) were used to interfere with uptake via megalin and the AT1R, respectively. ANG(1-7) uptake was prevented by anti-megalin antisera (63%) to a higher degree than olmesartan (13%) (P < 0.001). In analysis by flow cytometry of binding experiments performed in brush-border membrane vesicles isolated from kidneys of CD-1 mice, anti-megalin antisera interfered with ANG-(1-7) binding more strongly than olmesartan (P < 0.05 against positive control). Interactions of megalin with ANG-(1-7) at a molecular level were studied by surface plasmon resonance, demonstrating that ANG-(1-7) binds megalin dose and time dependently and with an affinity similar to ANG II. These results show that the scavenger receptor megalin binds and internalizes ANG-(1-7).