·药理·

Y-27632对低氧性肺动脉收缩的作用

刘蕾，沈歆，张志武，肖雄，李小强*

(第四军医大学药学院药理学教研室，西安710032)

摘要：目的探讨Y-27632对低氧性肺动脉收缩的作用及其机制。方法建立小鼠低氧损伤模型；采用微血管张力实验测定肺动脉收缩力，WB检测TRPC6蛋白表达，离子影像技术测定平滑肌细胞全细胞钙变化。结果与对照组比较，CPA诱导低氧组肺动脉的收缩(加强到了167.9%±68.6%, `*P<0.05；Y-27632可显著抑制这一变化(抑制率为47.3%±17.8%, `#P<0.05)；低氧可致肺动脉血管TRPC6蛋白表达显著增加(增加到154.8%±34.3%, `*P<0.05)，这一变化可被Y-27632显著降低(降低到134.2%±18.2%, `#P<0.05)；CPA诱导低氧组肺动脉平滑肌细胞[Ca`(2+)]i显著升高(荧光比率升高到188.9%±46.6%, `*P<0.05)，此作用可被Y-27632显著抑制(荧光比率降低了46.7%±14.5%, `#P<0.05)。结论ROCK抑制剂Y-27632可通过调控TRPC6蛋白表达及其介导的[Ca`(2+)]i变化，重要性地参与低氧性肺动脉收缩。

关键词:钙离子；低氧性肺动脉收缩；经典瞬时受体电位通道蛋白；Y-27632

doi:10.3969/j.issn.1004-2407.2015.03.011

中图分类号：R965

文献标志码：A

文章编号：1004-2407(2015)03-0253-04

Abstract:ObjectiveTo investigate the effect of Y-27632 on hypoxic pulmonary vasoconstriction and the potential mechanism. Methods Hypoxia-induced injury model was made in mouse. CPA-induced muscle contraction was detected by capillary tension experiment. To explore the expression of TRPC6, WB and immunofluorescence staining experiments were performed. Ca`(2+) was measured using a fluorescence imaging system. Results CPA-induced muscle contraction was enhanced by 167.9%±68.6% in hypoxic pulmonary artery compared with control, which was markedly inhibited by Y-27632(inhibition ratio was 47.3%±17.8%, `#P<0.05). The expression of TRPC6 induced by hypoxia was markedly increased (to 154.8%±34.3%, `*P<0.05). After treatment with Y-27632, the expression of TRPC6 was significantly decreased compared with hypoxia mice (to 134.2%±18.2%, `#P<0.05). Ca`(2+) influx in pulmonary artery smooth muscle cells elicited by store depletion using CPA were dramatically augmented in hypoxia group by 188.9%±46.6% in the ratio of 340 nm/380 nm(`*P<0.05). However, Y-27632 obviously attenuated this change by 46.7%±14.5%(`#P<0.05).Conclusion Inhibition of ROCK by Y-27632 can importantly take part in HPV via the regulation of TRPC6 expression in pulmonary artery, which mediated extracellular Ca`(2+) influx.

基金项目：国家自然科学基金(No.81170107,No.30800433)

作者简介：刘蕾，女，硕士研究生

收稿日期：(2015-01-30)

Effect of Y-27632 on hypoxic pulmonary vasoconstriction

LIU Lei, SHEN Xin, ZHANG Zhiwu, XIAO Xiong, LI Xiaoqiang*(Department of Pharmacology, School of Pharmacy,the Fourth Military Medical University, Xi′an 710032, China)

Key words: calcium; hypoxic pulmonary vasoconstriction; TRPC channel; Y-27632

*通信作者：李小强，男，副教授，硕士研究生导师

低氧性肺血管收缩(hypoxic pulmonary vasoconstriction，HPV) 是低氧性肺动脉高压(hypoxic pulmonary hypertension，HPH)形成的基本病理生理特征之一[1,2]。寻找针对HPV的防治药物是肺动脉高压治疗的重要策略。研究发现[3]，RhoA蛋白及其下游效应因子Rho激酶(Rho-kinase, ROCK)通路参与了HPH的发生、发展，ROCK抑制剂是一种很有希望的PH治疗药物。Y-27632作为一种ROCK抑制剂，是否能在HPV中发挥作用，目前尚不清楚，有待深入研究[4]。本研究通过检测Y-27632对环匹阿尼酸(CPA)诱导的肺动脉血管收缩，经典瞬时受体电位通道6(transient receptor potential canonical channel, TRPC6)的蛋白表达及其介导的细胞内Ca2+动员等的影响，以明确其对HPV的作用，探讨其可能的机制。

1仪器与材料

1.1仪器WB化学发光成像检测系统(Tanon公司)；雷磁PHS-3C pH计(上海精密科学化学有限公司)；AB135-S精密天平(METTLER公司)；SCIENTZ-IID型超声波细胞粉碎仪(宁波新芝生物有限公司)；5415R低温离心机(Eppendorf公司)；TILL离子影像系统(TILL公司)；Power Lab系统(AD公司)；激光共聚焦显微镜(Olympus公司)。……