Expression and function of the calcium-sensing receptor in juxtaglomerular cells.
Expression and function of the calcium-sensing receptor in juxtaglomerular cells.
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肾小球旁细胞中钙敏感受体的表达和功能。
DOI：
10.1161/hypertensionaha.107.095158
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发表时间：
2007
期刊：
影响因子：
--
通讯作者：
Beierwaltes,WilliamH
中科院分区：
文献类型：
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作者：
Ortiz-Capisano,MCecilia;Ortiz,PabloA;Garvin,JeffreyL;Harding,Pamela;Beierwaltes,WilliamH
钙敏感受体感知细胞外钙的微摩尔变化，并将其转化为细胞内钙的变化。肾素是肾素-血管紧张素系统的组成部分，通过cAMP依赖的途径由肾小球旁细胞合成、储存和释放。细胞内钙的增加抑制了肾小球旁细胞腺苷环化酶亚型V型、cAMP的形成和肾素的释放。我们假设钙敏感受体在肾小球旁细胞中表达，并介导细胞内钙和肾素释放的变化。为了验证这一点，我们使用了分离的小鼠肾小球旁细胞的原代培养，在其中我们进行了RT-PCR、Western blotts和免疫荧光。RT-PCR结果显示在预期的151bp处有一条与钙敏感受体一致的阳性条带。Western blotts显示了一条130到150 kDa的条带，证实了肾小球旁细胞中的钙感受器。免疫荧光和共聚焦显微镜下观察到肾小球旁细胞钙敏感受体的免疫荧光和共聚焦显微镜显示肾素的阳性标记。为了检测钙敏感受体是否调节肾素的释放，将肾小球旁细胞与钙敏感受体激动剂Cinacalcet-HCl孵育，浓度分别为50和1000nmol/L，钙浓度为0.25 mmol/L。肾素释放量从541.9±86.2ng/毫升降至364.6±64.1ng/毫升(P<；0.001)，肾素释放为279.6±56.9ng/毫升/毫克蛋白(P<；0.001)。我们的结论是，肾小球旁细胞表达钙敏感受体，它们的激活导致腺苷环化酶-V活性的抑制，减少cAMP的形成和抑制肾素的释放。
Calcium-sensing receptors sense and translate micromolar changes of extracellular calcium into changes in intracellular calcium. Renin, a component of the renin-angiotensin system, is synthesized by, stored in, and released from the juxtaglomerular cells through a cAMP-dependent pathway. Increased intracellular calcium inhibits the adenylyl cyclase isoform type V, cAMP formation, and renin release from juxtaglomerular cells. We hypothesized that calcium-sensing receptors are expressed in juxtaglomerular cells and mediate changes in intracellular calcium and renin release. To test this we used primary cultures of isolated mouse juxtaglomerular cells in which we ran RT-PCR, Western blots, and immunofluorescence. RT-PCR showed a positive band at the expected 151 bp consistent with calcium-sensing receptor. Western blots showed a 130- to 150-kDa band confirming the calcium-sensing receptor in juxtaglomerular cells. Immunofluorescence and confocal microscopy using 2 different antibodies against the calcium-sensing receptor in juxtaglomerular cells showed positive fluorescence in the juxtaglomerular cells, which also had positive labeling for renin. To test whether calcium-sensing receptors regulate renin release, juxtaglomerular cells were incubated with a calcium-sensing receptor agonist, the calcimimetic cinacalcet-HCl, at concentrations of 50 and 1000 nmol/L in 0.25 mmol/L of calcium medium. Cinacalcet-HCl decreased juxtaglomerular cell cAMP formation to 47.3±6.8% and 44.2±9.7% of basal, respectively (P<0.001), and decreased renin release from 541.9±86.2 to 364.6±64.1 (P<0.05) and 279.6±56.9 (P<0.005) ng of angiotensin I per milliliter per hour per milligram of protein, respectively. We conclude that juxtaglomerular cells express the calcium-sensing receptor and that their activation leads to inhibition of adenylyl cyclase-V activity, decreasing cAMP formation and suppressing renin release.