血管平滑肌细胞过表达人EP4受体改善血管紧张素Ⅱ诱导的高血压

Overexpression of human EP4 receptor in vascular smooth muscle cells attenuates angiotensinⅡ-induced hypertension in mice

前列腺素E_(2)(prostaglandin E_(2),PGE_(2))在心血管系统中具有重要的调节作用,PGE_(2)通过G蛋白耦联受体(EP1、EP2、EP3、EP4)参与机体血压的稳态维持。本文旨在利用血管平滑肌细胞特异性人EP4转基因小鼠(VSMC-hEP4 Tg)研究血管平滑肌细胞过表达人EP4受体对小鼠血压的作用及机制。通过转基因技术构建VSMC-hEP4 Tg小鼠并进行鉴定;使用尾套法测定小鼠在基础状态及高、低盐饮食条件下的血压水平;在小鼠背部皮下埋置血管紧张素Ⅱ(angiotensin Ⅱ,Ang Ⅱ)缓释泵后检测其血压变化;颈动脉插管法测量小鼠血压对Ang Ⅱ静脉灌注的急性反应以及两种EP4特异性激动剂(CAY10580和CAY10598,0.5 mg/kg)对小鼠血压的影响;使用血管张力仪测定小鼠肠系膜动脉血管环对Ang Ⅱ诱导的血管收缩力;运用Western blot检测EP4特异性激动剂PGE_(1)-OH对Ang Ⅱ诱导的肌球蛋白磷酸酶靶亚基1(myosin phosphatase target subunit 1,MYPT1)磷酸化的作用。结果显示:与野生型(wild type,WT)小鼠相比,VSMC-hEP4 Tg小鼠基础血压明显较低,在高、低盐饮食条件下其血压均保持较低水平;VSMC-h EP4 Tg小鼠的血压在Ang Ⅱ缓释给药及静脉灌注后均显著低于WT小鼠,其肠系膜血管环对Ang Ⅱ的反应性也明显下降。此外,CAY10580和CAY10598均能使WT小鼠血压显著降低。进一步研究发现,PGE_(1)-OH可抑制肠系膜动脉中Ang Ⅱ介导的MYPT1 Thr696位点的磷酸化。以上结果表明,血管平滑肌细胞特异性过表达人EP4基因可显著降低小鼠基础血压,并能减轻Ang Ⅱ诱导高血压的发生,其机制可能与EP4抑制AngⅡ的信号通路有关,提示基于EP4特异性的长效激动剂的开发可能在高血压防治中有重要价值。

Prostaglandin E_(2)(PGE_(2))plays an important role in cardiovascular system.PGE_(2) regulates blood pressure through its 4 G protein coupled receptors,i.e.,EP1,EP2,EP3,and EP4.The aim of this study was to investigate the role of EP4 receptors in vascular smooth muscle cells(VSMC)in blood pressure regulation.VSMC-specific human EP4 transgenic(VSMC-hEP4 Tg)mice were generated and genotyped.The systolic blood pressure(SBP)of the VSMC-hEP4 Tg mice and the wild-type(WT)littermates was measured under normal,low-salt(LSD)and high-salt diet(HSD)conditions using a tail-cuff method.Both WT and VSMC-hEP4 Tg mice were administered with a chronic infusion of angiotensin Ⅱ(Ang Ⅱ)with an osmotic pump and SBP levels were monitored every week.The mean arterial blood pressure(MAP)of WT and VSMC-hEP4 Tg mice upon Ang Ⅱ intravenous infusion was measured via carotid arterial catheterization.Ang Ⅱ-induced vasoconstriction of the mesenteric arterial rings from WT and VSMC-hEP4 Tg mice was measured using the multi myograph system.The effect of PGE_(1)-OH(a selective EP4 agonist)on Ang Ⅱ-induced phosphorylation of myosin phosphatase target subunit 1(MYPT1)was detected by Western blot.The effect of two additional EP4 specific agonists(CAY10580 and CAY10598,0.5 mg/kg)on blood pressure of WT mice was measured by carotid arterial catheterization.The results showed that the VSMC-hEP4 Tg mice were successfully generated and their basal SBP levels were lower than those of WT mice.Although blood pressure levels were significantly altered in WT mice under LSD and HSD,little change was observed in the VSMChEP4 Tg mice.After a chronic infusion and an acute intravenous injection of Ang Ⅱ,SBP levels of VSMC-hEP4 Tg mice were significantly lower than those of WT mice.In addition,both CAY10580 and CAY10598 significantly reduced MAP levels of WT mice.Ex vivo study showed that treatment of isolated mesenteric arteries with PGE_(1)-OH inhibited Ang Ⅱ-induced phosphorylation of MYPT1.Collectively,these results demonstrate that specific overexpression of human EP4 gene in VSMCs significantly reduces basal blood pressure levels and attenuates Ang Ⅱ-induced hypertension,possibly via inhibiting Ang Ⅱ/AT1 signaling pathway.Our findings suggest that EP4 may represent an attractive target for the treatment of hypertension.