Angiotensin-converting enzyme inhibitor ramiprilat interferes with the sequestration of the B2 kinin receptor within the plasma membrane of native endothelial cells
Angiotensin-converting enzyme inhibitor ramiprilat interferes with the sequestration of the B2 kinin receptor within the plasma membrane of native endothelial cells
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DOI：
10.1161/01.cir.99.15.2034
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发表时间：
1999-04-20
期刊：
影响因子：
37.8
通讯作者：
Busse, R
中科院分区：
文献类型：
--
作者：
Benzing, T;Fleming, I;Busse, R
背景：ace （kininase II）抑制剂已被证明通过抑制血管紧张素II的形成和缓激肽的分解来发挥其有益的心血管作用。由于最近的证据表明，ACE抑制剂也可能干扰B-2激肽受体信号，从而增强血管对缓激肽的反应，我们研究了天然内皮细胞质膜内B-2激肽受体的分布是否受到ACE抑制剂的影响。方法与结果：采用[H-3]缓激肽特异性结合和B-2受体免疫沉淀的方法，研究了B-2受体在猪主动脉内皮细胞制备膜中的定位。缓激肽和雷米普利特对细胞内信号传导的影响是通过监测细胞外调节激酶Erk1和Erk2的激活以及fura 2负载内皮细胞中[Ca2+](i)的增加来确定的。用100 nmol/L的缓激肽刺激天然内皮细胞，导致B-2受体在富小窝蛋白（CR）膜上的时间依赖性隔离，在5分钟后达到最大。雷米普利特100 nmol/L预处理15分钟显著降低了CR膜中B-2激肽受体的恢复，而增加了缺乏小窝蛋白膜中B-2激肽受体的恢复。这种效果不是由于抑制缓激肽的降解，因为在合成ACE底物的抑制浓度存在时，没有看到任何效果-l -组氨酸-l -亮氨酸。在缓激肽刺激之前或之后使用雷米普利特也能降低[H-3]缓激肽与CR膜的结合。此外，雷米普利特在缓激肽刺激的细胞中导致B-2受体的再激活，并诱导[Ca2+](i)的第二个峰值和Erk1/2的再激活。结论：血管紧张素转换酶抑制剂雷米普利特干扰天然内皮细胞B-2激肽受体对CR膜结构域的靶向作用。因此，雷米普利特和其他ACE抑制剂对血管系统的影响可能是由于抑制激酶II以外的作用。
Background-ACE (kininase II) inhibitors have been shown to exert their beneficial cardiovascular effects via the inhibition of both angiotensin II formation and bradykinin breakdown. Because recent evidence suggests that ACE inhibitors may also interfere with B-2 kinin receptor signaling and thus enhance the vascular response to bradykinin, we examined whether the distribution of B-2 kinin receptors within the plasma membrane of native endothelial cells is affected by an ACE inhibitor.Methods and Results-Localization of the B-2 kinin receptor in membranes prepared from native porcine aortic endothelial cells was evaluated by means of specific [H-3]bradykinin binding and immunoprecipitation of the B-2 receptor from isolated membranes. Effects of bradykinin and ramiprilat on intracellular signaling were determined by monitoring the activation of the extracellularly regulated kinases Erk1 and Erk2 as well as [Ca2+](i) increases in fura 2-loaded endothelial cells. Stimulation of native endothelial cells with bradykinin 100 nmol/L resulted in the time-dependent sequestration of-the B-2 receptor to caveolin-rich (CR) membranes, which was maximal after 5 minutes. Pretreatment with ramiprilat 100 nmol/L for 15 minutes significantly attenuated the recovery of B-2 kinin receptors in CR membranes while increasing that from membranes lacking caveolin. This effect was not due to the inhibition of bradykinin degradation, because no effect was seen in the presence of an inhibitory concentration of the synthetic ACE substrate hippuryl-L-histidyl-L-leucine. Ramiprilat also decreased [H-3]bradykinin binding to CR membranes when applied either before or after bradykinin stimulation. Moreover, ramiprilat resulted in reactivation of the B-2 receptor in bradykinin-stimulated cells and induced a second peak in [Ca2+](i) and reactivation of Erk1/2.Conclusions-The ACE inhibitor ramiprilat interferes with the targeting of the B-2 kinin receptor to CR membrane domains in native endothelial cells. Therefore, effects other than the inhibition of kininase II may account for the effects of ramiprilat and other ACE inhibitors on the vascular system.