摘要
慢性低氧性肺动脉高压(CHPH)是大部分慢性阻塞性肺疾病(COPD)患者最终发展成为肺心病而死亡的主要原因,关于其发病机制已经进行了大量的研究,临床以此为基础的治疗也取得了一定的疗效,然而,临床治疗CHPH的手段仍然有限,疗效依然欠佳。肺血管低氧早期(急性低氧)的收缩反应和慢性低氧性肺血管结构重建是CHPH发生的主要机制,造成CHPH的这两个主要环节不仅发生在肺动脉,而且包括肺静脉。尽管关于肺动脉急性低氧性收缩反应和慢性低氧性肺血管结构重建的机制已经进行了大量的研究,然而,关于肺静脉的此类研究却少有报道。大量的研究证明:细胞内Ca~(2+)在血管平滑肌细胞的收缩和增生的复杂机制中起至关重要的作用。但是,关于肺静脉平滑肌细胞(PVSMC)Ca~(2+)信号转导以及低氧对PVSMC细胞内Ca~(2+)浓度([Ca~(2+)]_i)作用的研究却鲜有报道,我们推测:低氧可能通过某些机制使PVSMC的[Ca~(2+)]_i改变,参与CHPH的发生、发展过程。因此,我们建立了大鼠远端PVSMC原代培养模型及CHPH大鼠模型,对急慢性低氧诱导大鼠远端肺静脉平滑肌[Ca~(2+)]_i改变的作用及机理进行了研究。从以往研究甚少的肺静脉方面来阐述CHPH发生的可能机制,为寻找有效的防治肺动脉高压的方法打下坚实的理论基础。
方法
一、急性低氧对大鼠远端PVSMC[Ca~(2+)]_i的影响:首先利用胶原酶消化法分离、原代培养大鼠远端PVSMC,通过细胞形态学观察、平滑肌细胞α-actin免疫荧光和western-blot免疫印迹分析以及[Ca~(2+)]_i对高钾KRBS溶液的反应来鉴定PVSMC,从而建立大鼠远端PVSMC原代培养模型。然后利用InCyte细胞内钙浓度检测系统,观测急性低氧对大鼠远端PVSMC[Ca~(2+)]_i的影响
二、急性低氧对大鼠远端PVSMC[Ca~(2+)]_i影响的机制:1、利用InCyte细胞内钙浓度检测系统,观测电压依赖性钙通道(VDCC)拮抗剂硝苯地平对急性低氧诱导大鼠远端PVSMC[Ca~(2+)]_i改变的干预作用,明确是否有VDCC以外的钙通道参与急性低氧诱导的大鼠远端PVSMC[Ca~(2+)]_i改变过程。2、分离大鼠远端肺静脉平滑肌,原代培养大鼠远端PVSMC,通过荧光定量PCR和western-blot免疫印迹分析来检测经典瞬时受体电位(TRPC)蛋白在大鼠远端肺静脉平滑肌和培养的大鼠远端PVSMC的mRNA水平和蛋白水平的表达,明确大鼠远端肺静脉平滑肌是否表达TRPC蛋白。3、利用InCyte细胞内钙浓度检测系统,通过恢复细胞外钙法、锰荧光焠灭法,并利用储量操纵性钙通道(SOCC)拮抗剂SKF-96365和NiCl2,在大鼠远端PVSMC上检测Ca~(2+)通过由TRPC蛋白组成的SOCC而形成的储量操作性钙内流(SOCE),明确大鼠远端肺静脉平滑肌是否存在SOCE。4、利用InCyte细胞内钙浓度检测系统,通过恢复细胞外钙法、锰荧光焠灭法,并利用SOCC拮抗剂SKF-96365和NiCl2观测急性低氧对大鼠远端PVSMC的SOCE的影响,明确SOCE在急性低氧引起的大鼠远端PVSMC[Ca~(2+)]_i改变中的作用。
三、慢性低氧对大鼠远端PVSMC的[Ca~(2+)]_i的影响:首先,通过慢性持续低氧(10%O2)条件下饲养大鼠3周建立CHPH大鼠模型;然后,分离正常大鼠和CHPH大鼠远端PVSMC;最后,利用InCyte细胞内钙浓度检测系统,分别检测正常大鼠和CHPH大鼠远端PVSMC的[Ca~(2+)]_i,从而明确慢性低氧对大鼠远端PVSMC的[Ca~(2+)]_i的影响。
四、慢性低氧对大鼠远端PVSMC的[Ca~(2+)]_i的影响的机制:1、分离正常大鼠和CHPH大鼠远端肺静脉平滑肌,通过荧光定量PCR和Western-blot免疫印迹分析来检测TRPC在正常大鼠和CHPH大鼠远端肺静脉平滑肌的表达,明确慢性低氧对大鼠远端肺静脉平滑肌TRPC表达的影响。2、分离正常大鼠和CHPH大鼠远端PVSMC,利用InCyte细胞内钙浓度检测系统,首先分别检测由环匹阿尼酸(CPA)诱导的正常大鼠和CHPH大鼠远端PVSMC的[Ca~(2+)]_i变化,然后分别检测由CPA诱导的正常大鼠和CHPH大鼠远端PVSMC的锰荧光焠灭,从而明确慢性低氧对大鼠远端肺静脉平滑肌SOCE的影响;接着分别检测由油酰基乙酰甘油(OAG)诱导的正常大鼠和CHPH大鼠远端PVSMC的[Ca~(2+)]_i变化,最后分别检测由OAG诱导的正常大鼠和CHPH大鼠远端PVSMC的锰荧光焠灭,从而明确慢性低氧对大鼠远端肺静脉平滑肌由受体操纵性钙通道(ROCC)介导的Ca~(2+)内流的影响。
结果
一、急性低氧对大鼠远端PVSMC[Ca~(2+)]_i的影响:
1、成功建立了大鼠远端PVSMC原代培养模型,原代培养的大鼠远端PVSMC不仅表现出典型的血管平滑肌细胞的形态学和免疫学特征,而且具有完好的生理学功能。
2、急性低氧使大鼠远端PVSMC[Ca~(2+)]_i升高。
二、急性低氧对大鼠远端PVSMC[Ca~(2+)]_i影响的机制:
1、作为VDCC拮抗剂,硝苯地平能完全阻断高钾KRBS溶液引起的大鼠远端PVSMC [Ca~(2+)]_i升高,却只能部分抑制急性低氧引起的大鼠远端PVSMC的[Ca~(2+)]_i升高,提示VDCC以外的钙通道参与急性低氧引起的大鼠远端PVSMC[Ca~(2+)]_i升高。
2、在大鼠远端肺静脉平滑肌组织和培养的PVSMC都有TRPC的mRNA表达,其中以TRPC-1、TRPC-4和TRPC-6,尤其是TRPC-1和TRPC-6的mRNA表达最强。Western-blot免疫印迹分析进一步表明,在大鼠远端肺静脉平滑肌组织和培养的PVSMC都有TRPC蛋白的表达,且尤其以TRPC-1和TRPC-6的蛋白表达最强。
3、在去除细胞外Ca~(2+)、用硝苯地平阻断VDCC并用CPA使Ca~(2+)从PVSMC肌浆网完全清空的条件下,恢复PVSMC细胞外Ca~(2+)后, PVSMC [Ca~(2+)]_i可迅速显著升高;在去除细胞外Ca~(2+)并用硝苯地平阻断VDCC的条件下,CPA使Mn~(2+)引起的PVSMC的荧光焠灭增强;作为SOCC拮抗剂,SKF-96365和NiCl2均能明显抑制由CPA诱导的PVSMC [Ca~(2+)]_i升高和锰荧光焠灭增强,但对高钾KRBS溶液引起的PVSMC [Ca~(2+)]_i升高没有影响,提示大鼠远端PVSMC存在SOCE。
4、急性低氧能显著增加由CPA诱导的、恢复细胞外钙后所引起的大鼠远端PVSMC[Ca~(2+)]_i升高,并能增强由CPA诱导的大鼠远端PVSMC的锰荧光焠灭; SKF-96365和NiCl2均能明显抑制急性低氧引起的大鼠远端PVSMC的SOCE增加;在常氧情况下,用不含CPA但含有硝苯地平的无钙溶液孵育PVSMC,锰荧光焠灭与不加Mn~(2+)的细胞的荧光自发衰减量没有差异;而在急性低氧情况下,用不含CPA但含有硝苯地平的无钙溶液孵育PVSMC,锰荧光焠灭显著增加,而且此反应能被SKF-96365和NiCl2明显抑制,提示急性低氧自身能诱发大鼠远端PVSMC的SOCE,而且能够通放大SOCE,引起PVSMC [Ca~(2+)]_i升高。
三、慢性低氧对大鼠远端PVSMC的[Ca~(2+)]_i的影响:
1、成功建立CHPH大鼠模型,慢性持续性低氧3周大鼠的MPAP、RVSP、RV/(LV+S)都明显增高,肺内血管壁明显增厚。
2、CHPH大鼠远端PVSMC[Ca~(2+)]_i较正常大鼠明显增高,提示慢性低氧使大鼠远端PVSMC的[Ca~(2+)]_i升高。
四、慢性低氧对大鼠远端PVSMC[Ca~(2+)]_i影响的机制:
1、荧光定量PCR和western-blot免疫印迹分析证实:在CHPH大鼠远端肺静脉平滑肌组织TRPC-6蛋白的表达较正常大鼠明显增强,而TRPC-1和TRPC-4蛋白的表达较正常大鼠没有明显改变,提示慢性低氧上调大鼠远端肺静脉平滑肌组织TRPC-6蛋白的表达。
2、用硝苯地平阻断VDCC并恢复细胞外Ca~(2+)后,CPA不仅能导致CHPH大鼠远端PVSMC [Ca~(2+)]_i迅速升高,而且能使正常大鼠远端PVSMC的[Ca~(2+)]_i也迅速升高,两者升高幅度无差别;用含CPA和硝苯地平的无钙溶液孵育细胞,CHPH大鼠远端PVSMC的锰荧光焠灭与正常大鼠的也无差别。然而,用硝苯地平阻断VDCC并恢复细胞外Ca~(2+)后,OAG能使正常大鼠远端PVSMC的[Ca~(2+)]_i升高,而CHPH大鼠远端PVSMC由OAG诱导的[Ca~(2+)]_i升高较正常大鼠更明显;同时,OAG诱导的锰荧光焠灭在CHPH大鼠远端PVSMC也比正常大鼠表现的更强,提示慢性低氧通过增加由ROCC介导的Ca~(2+)内流使大鼠远端PVSMC的[Ca~(2+)]_i升高。
结论
1、急性和慢性低氧都能使大鼠远端肺静脉平滑肌细胞内钙浓度升高。
2、大鼠远端肺静脉平滑肌主要表达TRPC-1、TRPC-4和TRPC-6,尤以TRPC-1和TRPC-6为主;大鼠远端肺静脉平滑肌存在SOCE。
3、急性低氧不仅自身能够诱发大鼠远端肺静脉平滑肌细胞的SOCE,而且能够放大SOCE,从而引起细胞内钙浓度升高。
4、慢性低氧能够上调大鼠远端肺静脉平滑肌TRPC-6的表达,使主要由TRPC-6蛋白组成的ROCC增加,导致由ROCC介导的Ca~(2+)内流增加,从而引起细胞内钙浓度升高。
Objective
Hypoxia causes remodeling and alters contractile responses in both pulmonary arteries and pulmonary veins (PV). Although pulmonary arteries have been studied extensively in this disorder, the mechanisms by which pulmonary veins respond to hypoxia and whether these responses contribute to chronic hypoxic pulmonary hypertension (CHPH) remain poorly understood. In pulmonary arterial smooth muscle, influx of Ca~(2+) through store-operated calcium channels thought to be composed of canonical transient receptor potential (TRPC) proteins is likely to play an important role in development of CHPH. The aim of this study is to determine whether this mechanism could also be operative in pulmonary venous smooth muscle.
Methods
The rat distal PVSMC were isolated and cultured by collagenase digestion and characterized by morphological activity, by immunoblotting and immunofluorescence of smooth muscle -actin and by response of intracellular Ca~(2+) concentration ([Ca~(2+)]_i) to KCL.
The effect of acute hypoxia on [Ca~(2+)]_i in PVSMC and the inhibition of nifedipine were measured by Incyte intracellular Ca~(2+) concentration system. The expression of TRPC in distal pulmonary venous smooth muscle or PVSMC was detected by real-time PCR and western blotting. Store-operated Ca~(2+) entry (SOCE) and the effect of acute hypoxia on SOCE were assessed by measurement of increase in [Ca~(2+)]_i caused by restoration of extracellular Ca~(2+) and the rate at which extracellular Mn~(2+) quenched fura-2 fluorescence.
CHPH rats were obtained using a hypoxic (10 % O2) chamber for 21 days. PVSMC were isolated from CHPH rats or normal rats. The effect of chronic hypoxia on [Ca~(2+)]_i in PVSMC was assessed by comparison of [Ca~(2+)]_i in PVSMC from CHPH rats with that from normal rats.
The effect of chronic hypoxia on TRPC expression was measured by comparison of TRPC mRNA and protein expression in pulmonary venous smooth muscle from CHPH rats with that from normal rats using real-time PCR and western blotting. PVSMC from CHPH rats or normal rats were isolated. The mechanisms by which [Ca~(2+)]_i in PVSMC respond to chronic hypoxia were assessed by comparison of CPA-induced Ca~(2+) influx and Mn~(2+) quenching in PVSMC isolated from CHPH rats with that from normal rats and by comparison of OAG-induced Ca~(2+) influx and Mn~(2+) quenching in PVSMC isolated from CHPH rats with that from normal rats.
Results
Cells isolated from rat distal pulmonary veins displayed morphological characteristic of vascular smooth muscle cells and expressed smooth muscle -actin and the functional voltage-depended Ca~(2+) channels (VDCC). Acute hypoxia could increase [Ca~(2+)]_i in rat distal PVSMC.
VDCC antagonist nifedipine could block KCL-induced increase of [Ca~(2+)]_i in rat distal PVSMC. However, nifedipine partially prevented the effect of acute hypoxia on [Ca~(2+)]_i , indicating acute hypoxia actived influx of Ca~(2+) through VDCC and another independent pathway. Real-time PCR and western blotting both indicated that TRPC were expressed in rat distal pulmonary venous smooth muscle or PVSMC with most abundant expression of TRPC1, TRPC6 and TRPC4.
Restoration of extracellular Ca~(2+) caused a large increase of [Ca~(2+)]_i in PVSMC perfused with Ca~(2+)-free KRB solution containing CPA and nifedipine. In the presence of CPA, fura-2 fluorescence decreased more than that in the absence of CPA after administration of Mn~(2+) in PVSMC perfused with Ca~(2+)-free KRB solution containing nifedipine. The store-operated Ca~(2+) channels (SOCC) antagonists SKF-96365 and NiCl2 both inhibited CPA-induced Ca~(2+) influx and Mn~(2+) quenching, but had no effect on KCL-induced increase of [Ca~(2+)]_i in PVSMC. These data indicated SOCE in rat distal PVSMC.
Acute hypoxia caused enhancement of CPA-induced increase in [Ca~(2+)]_i elicited by Ca~(2+) restoration and the rate of Mn~(2+) quenching. SKF-96365 and NiCl2 blocked hypoxic enhancement of both the [Ca~(2+)]_i response to extracellular Ca~(2+) restoration and the rate of Mn~(2+) quenching. In normoxic PVSMC treated with nifedipine but not CPA, Mn~(2+) quenching did not differ from the spontaneous decline of fluorescence in cells not exposed to Mn~(2+). During acute hypoxia, however, the rate of Mn~(2+) quenching in the absence of CPA increased, and this increase was blocked by SKF-96365 or NiCl2.
The MPAP, RVSP and RV/(LV+S) were all increased in rats exposed to chronic hypoxia. The pulmonary vessels from CHPH rats were thicker than that from normal rats. [Ca~(2+)]_i in PVSMC from CHPH rats was higher than that from normal rats, indicating chronic hypoxia could increase [Ca~(2+)]_i in rat distal PVSMC.
Both TRPC6 mRNA and protein expression in pulmonary venous smooth muscle from CHPH rats were stronger than that from normal rats. But neither TRPC1 nor TRPC4 mRNA or protein expression in pulmonary venous smooth muscle from CHPH rats differed from that from normal rats.
In the presence of nifedipine and CPA, neither the increase of [Ca~(2+)]_i elicited by Ca~(2+) restoration nor the rate of Mn~(2+) quenching in PVSMC from CHPH rats differed from that from normal rats. In the presence of nifedipine and OAG, however, restoration of extracellular Ca~(2+) caused a larger increase of [Ca~(2+)]_i in PVSMC from CHPH rats than that from normal rats. Otherwise, in the presence of nifedipine and OAG, the rate of Mn~(2+) quenching in PVSMC from CHPH rats was also larger than that from normal rats.
Conclusion
1. Both acute and chronic hypoxia could increase [Ca~(2+)]_i in rat distal PVSMC.
2. SOCE and TRPC proteins were expressed in rat distal pulmonary venous smooth muscle with most abundant expression of TRPC1, TRPC6 and TRPC4.
3. Acute hypoxia was able to evoke SOCE in rat distal PVSMC on its own, as well as amplify SOCE.
4. Chronic hypoxia could increase Ca~(2+) influx through receptor-operated Ca~(2+) channels (ROCC) thought to be composed of TRPC6 protein by upregulation of TRPC6 expression in rat distal PVSMC.
引文
1.慢性阻塞性肺疾病防治全球倡仪,2006。
2.冉丕鑫。慢性阻塞性肺疾病的患病危险因素及其预防。中华结核和呼吸杂志,2007;30:141-143。
3.实用内科学(第11版),1449。
4.内科学(第6版),86。
5.Zhao Y, Packer CS, and Rhoades RA. Pulmonary vein contracts in response to hypoxia. Am J Physiol Lung Cell Mol Physiol.1993;265: L87–L92.
6.Tracey WR, Hamilton JT, Craig ID, and Paterson NA. Responses of isolated guinea pig pulmonary venules to hypoxia and anoxia. J Appl Physiol.1989; 67:2147–2153.
7.Hillier SC, Graham JA, Hanger CC, Godbey PS, Glenny RW, and Wagner WW Jr. Hypoxic vasoconstriction in pulmonary arterioles and venules. J Appl Physiol. 1997;82: 1084–1090.
8.Raj JU and Chen P. Micropuncture measurement of microvascular pressures in isolated lamb lungs during hypoxia. Circ Res. 1986;59: 398–404.
9. Sheehan DW, Farhi LE, and Russell JA. Prolonged lobar hypoxia in vivo enhances the responsivity of isolated pulmonary veins to hypoxia. Am Rev Respir Dis. 1992;145: 640–645.
10. Raj JU, Hillyard R, Kaapa P, Gropper M, and Anderson J. Pulmonary arterial and venous constriction during hypoxia in 3- to 5-wk-old and adult ferrets. J Appl Physiol. 1990;69: 2183–2189.
11. Feletou M, Girard V, and Canet E. Different involvement of nitric oxide in endothelium-dependent relaxation of porcine pulmonary artery and vein: influence of hypoxia. J Cardiovasc Pharmacol. 1995;25: 665–673.
12.Chazova I, Loyd JE, Zhdanov VS, Newman JH, Belenkov Y, and Meyrick B.Pulmonary artery adventitial changes and venous involvement in primarypulmonary hypertension. Am J Pathol. 1995;146: 389–397.
13. Wagenvoort CA and Wagenvoort N. Pulmonary venous changes in chronic hypoxia. Virchows Arch. 1976;372: 51–56.
14. Dingemans KP and Wagenvoort CA. Pulmonary arteries and veins in experimental hypoxia. An ultrastructural study. Am J Pathol. 1978;93: 353–368.
15. Takahashi H, Soma S, Muramatsu M, Oka M, and Fukuchi Y. Upregulation of ET-1 and its receptors and remodeling in small pulmonary veins under hypoxic conditions. Am J Physiol Lung Cell Mol Physiol. 2001;280: L1104–L1114.
16. Johnson JE, Perkett EA, and Meyrick B. Pulmonary veins and bronchial vessels undergo remodeling in sustained pulmonary hypertension induced by continuous air embolization into sheep. Exp Lung Res. 1997;23: 459–473.
17. Chazova I, Loyd JE, Zhdanov VS, Newman JH, Belenkov Y, and Meyrick B. Pulmonary artery adventitial changes and venous involvement in primary pulmonary hypertension. Am J Pathol. 1995;146: 389–397.
18.Doi, S., D.S. Damron, M. Horibe and P.A. Murray. Capacitative Ca~(2+) entry and tyrosine kinase activation in canine pulmonary arterial smooth muscle cells. Am J Physiol Lung Cell Mol Physiol. 2000;278: L118-130.
19. Gonzalez De La Fuente, P., J.P. Savineau and R. Marthan. Control of pulmonary vascular smooth muscle tone by sarcoplasmic reticulum Ca~(2+) pump blockers: thapsigargin and cyclopiazonic acid. Pflugers Arch.1995;429: 617-624.
20. McDaniel, S.S., O. Platoshyn, Wang J,Y. Yu, M. Sweeney, S. Krick, L.J. Rubin and J.X. Yuan. Capacitative Ca~(2+) entry in agonist-induced pulmonary vasoconstriction. Am J Physiol Lung Cell Mol Physiol. 2001;280: L870-880.
21. Ng, L.C. and A.M. Gurney. Store-operated channels mediate Ca~(2+) influx and contraction in rat pulmonary artery. Circ Res. 2001;89: 923-929.
22. Robertson, T.P., D. Hague, P.I. Aaronson and J.P. Ward.Voltage-independent calcium entry in hypoxic pulmonary vasoconstriction of intrapulmonary arteries of the rat. J Physiol. 2000;525: 669-680.
23. Golovina, V.A., O. Platoshyn, C.L. Bailey, Wang J, A. Limsuwan, M. Sweeney, L.J. Rubin and J.X. Yuan. Upregulated TRP and enhanced capacitative Ca(~(2+)) entry in human pulmonary artery myocytes during proliferation. Am J Physiol Heart Circ Physiol. 2001;280: H746-755.
24. Sweeney, M., Y. Yu, O. Platoshyn, S. Zhang, S.S. McDaniel and J.X. Yuan. Inhibition of endogenous TRP1 decreases capacitative Ca~(2+) entry and attenuates pulmonary artery smooth muscle cell proliferation. Am J Physiol Lung Cell Mol Physiol. 2002;283: L144-155.
25. Yu, Y., M. Sweeney, S. Zhang, O. Platoshyn, J. Landsberg, A. Rothman and J.X. Yuan. PDGF stimulates pulmonary vascular smooth muscle cell proliferation by upregulating TRPC6 expression. Am J Physiol Cell Physiol. 2003;284: C316-330.
26. Sham, J.S., B.R. Crenshaw, L.H. Deng, L.A. Shimoda and J.T. Sylvester. Effects of hypoxia in porcine pulmonary arterial myocytes: roles of KV channel and endothelin-1. Am J Physiol Lung Cell Mol Physiol. 2000;279: L262-272.
27. Archer, S.L. Diversity of phenotype and function of vascular smooth muscle cells. J Lab Clin Med. 1996;127: 524-529.
28. Barman, S.A. Potassium channels modulate hypoxic pulmonary vasoconstriction. Am J Physiol. 1998;275: L64-70.
29. Hasunuma, K., D.M. Rodman and I.F. McMurtry. Effects of K+ channel blockers on vascular tone in the perfused rat lung. Am Rev Respir Dis. 1991;144: 884-887.
30. McMurtry, I.F., A.B. Davidson, J.T. Reeves and R.F. Grover. Inhibition of hypoxic pulmonary vasoconstriction by calcium antagonists in isolated rat lungs. Circ Res. 1976;38: 99-104.
31.Cornfield, D.N., T. Stevens, I.F. McMurtry, S.H. Abman and D.M. Rodman. Acute hypoxia causes membrane depolarization and calcium influx in fetalpulmonary artery smooth muscle cells. Am J Physiol. 1994;266: L469-475.
32. Salvaterra, C.G. and W.F. Goldman. Acute hypoxia increases cytosolic calcium in cultured pulmonary arterial myocytes. Am J Physiol. 1993;264: L323-328.
33. Post, J.M., C.H. Gelband and J.R. Hume. [Ca~(2+)]_i inhibition of K+ channels in canine pulmonary artery. Novel mechanism for hypoxia-induced membrane depolarization. Circ Res.1995;77: 131-139.
34. Gelband, C.H. and H. Gelband. Ca~(2+) release from intracellular stores is an initial step in hypoxic pulmonary vasoconstriction of rat pulmonary artery resistance vessels. Circulation.1997;96: 3647-3654.
35. Madden, J.A., M.S. Vadula and V.P. Kurup. Effects of hypoxia and other vasoactive agents on pulmonary and cerebral artery smooth muscle cells. Am J Physiol. 1992;263: L384-393.
36. Vadula, M.S., J.G. Kleinman and J.A. Madden. Effect of hypoxia and norepinephrine on cytoplasmic free Ca~(2+) in pulmonary and cerebral arterial myocytes. Am J Physiol. 1993;265: L591-597.
37. Jabr, R.I., H. Toland, C.H. Gelband, X.X. Wang and J.R. Hume. Prominent role of intracellular Ca~(2+) release in hypoxic vasoconstriction of canine pulmonary artery. Br J Pharmacol. 1997;122: 21-30.
38. Liu, Q., J.S. Sham, L.A. Shimoda and J.T. Sylvester. Hypoxic constriction of porcine distal pulmonary arteries: endothelium and endothelin dependence. Am J Physiol Lung Cell Mol Physiol. 2001;280: L856-865.
39. Morio, Y. and I.F. McMurtry. Ca~(2+) release from ryanodine-sensitive store contributes to mechanism of hypoxic vasoconstriction in rat lungs. J Appl Physiol. 2002;92: 527-534.
40.Wang J, Weigand L, Sylvester JT, Semenza GL, and Shimoda LA. HIF-1 Mediates hypoxia-induced TRPC expression and elevated intracellular Ca~(2+) in pulmonary arterial smooth muscle cells. Circulation Research. 2006;98:1528–1537.
41.Shimoda, L.A., D.J. Manalo, J.S. Sham, G.L. Semenza and J.T. Sylvester. Partial HIF-1alpha deficiency impairs pulmonary arterial myocyte electrophysiological responses to hypoxia. Am J Physiol Lung Cell Mol Physiol. 2001;281: L202-208.
42. Smirnov, S.V., T.P. Robertson, J.P. Ward and P.I. Aaronson. Chronic hypoxia is associated with reduced delayed rectifier K+ current in rat pulmonary artery muscle cells. Am J Physiol. 1994;266: H365-370.
43. Suzuki, H. and B.M. Twarog. Membrane properties of smooth muscle cells in pulmonary hypertensive rats. Am J Physiol. 1982;242: H907-915.
44. Michael, J.R., T.P. Kennedy, P. Buescher, I. Farrukh, R. Lodato, P.C. Rock, J. Gottlieb, G. Gurtner, S.M. de la Monte and G.M. Hutchins. Nitrendipine attenuates the pulmonary vascular remodeling and right ventricular hypertrophy caused by intermittent hypoxia in rats. Am Rev Respir Dis. 1986;133: 375-379.
45. Morio, Y., K.G. Morris and I.F. McMurtry. Acute hemodynamic effects of Y27632, a selective Rho-kinase inhibitor, in chronically hypoxic pulmonary hypertensive rats (abstract). FASEB J. 2002;16: A74.
46. Shimoda, L.A., J.S. Sham, T.H. Shimoda and J.T. Sylvester. L-type Ca~(2+) channels, resting [Ca~(2+)]_i, and ET-1-induced responses in chronically hypoxic pulmonary myocytes. Am J Physiol Lung Cell Mol Physiol. 2000;279: L884-894.
47.Wang J, Shimoda LA, and Sylvester JT. Capacitative calcium entry and TRPC channel proteins are expressed in rat distal pulmonary arterial smooth muscle. Am J Physiol. Lung Cell Mol Physiol. 2004;286: L848-L858.
48.Hall KL, Harding JW, Hosick HL. Isolation and characterization of clonal vascular smooth muscle cell lines from spontaneously hypertensive and normotensive rat aortas. In Vitro Cell Dev Biol. 1991;27A:791-798.
49.Heimli H, K?hler H, Endresen MJ, Henriksen T, Lyberg T. A new method for isolation of smooth muscle cells from human umbilical cord arteries. Scand J Clin Lab Invest.1997;57:21-29.
50.Seidel MF, Simard JM, Hunter SF, Campbell GA. Isolation of arteriolar microvessels and culture of smooth muscle cells from cerebral cortex of guinea pig.Cell Tissue Res. 1991;265:579-87.
51.Cairr?o E, Santos-Silva AJ, Alvarez E, Correia I, Verde I. Isolation and culture of human umbilical artery smooth muscle cells expressing functional calcium channels.In Vitro Cell Dev Biol Anim. 2009;45:175-184.
52.Leik CE, Willey A, Graham MF, Walsh SW. Isolation and culture of arterial smooth muscle cells from human placenta. Hypertension. 2004;43:837-840
53.Moore SA, Strauch AR, Yoder EJ, Rubenstein PA, Hart MN. Cerebral microvascular smooth muscle in tissue culture. In Vitro. 1984 ;20:512-20.
54.Heaps CL, Parker JL, Sturek M, Bowles DK. Altered calcium sensitivity contributes to enhanced contractility of collateral-dependent coronary arteries.J Appl Physiol. 2004;97:310-6.
55.Zhao Y, Packer CS, Rhoades RA. Pulmonary vein contracts in response to hypoxia. Am J Physiol. 1993;265:L87-92.
56.彭公永,何志义,刘启才,李冰,钟南山,冉丕鑫。尾加压素Ⅱ促兔肺动脉平滑肌细胞增殖及机理探讨。中国病理生理杂志,2004,20(9):1597-1600。
57. Bakhramov A, Evans AM, and Kozlowski RZ. Differential effects of hypoxia on the intracellular Ca~(2+) concentration of myocytes isolated from different regions of the rat pulmonary arterial tree. Exp Physiol.1998;83: 337–347.
58. Cornfield DN, Stevens T, McMurtry IF, Abman SH, and Rodman DM. Acute hypoxia increases cytosolic calcium in fetal pulmonary artery smooth muscle cells. Am J Physiol Lung Cell Mol Physiol. 1993;265: L53–L56.
59. Olschewski A, Hong Z, Nelson DP, and Weir EK. Graded response of K+ current, membrane potential, and [Ca~(2+)]_i to hypoxia in pulmonary arterial smooth muscle. Am J Physiol Lung Cell Mol Physiol. 2002;283: L1143–L1150.
60. Harder DR, Madden JA, and Dawson C. Hypoxic induction of Ca~(2+)- dependent action potentials in small pulmonary arteries of the cat. J ApplPhysiol 59: 1389–1393, 1985.
61. Hoshino Y, Obara H, Kusunoki M, Fujii Y, and Iwai S. Hypoxic contractile response in isolated human pulmonary artery: role of calcium ion. J Appl Physiol. 1988;65:2468–2474.
62. Pfaffl MW. A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 2001;29:E45.
63.Wang J, Shimoda LA, Weigand L, Wang W, Sun D and Sylvester JT. Acute hypoxia increases intracellular [Ca~(2+)]_i in pulmonary arterial smooth muscle by enhancing capacitative Ca~(2+) entry. Am J Physiol Lung Cell Mol Physiol. 2005;288: L1059-L1069.
64.Kang TM, Park MK, and Uhm DY. Characterization of hypoxiainduced [Ca~(2+)]_i rise in rabbit pulmonary arterial smooth muscle cells. Life Sci. 2002;70: 2321–2333.
65.J. Roos, P.J. DiGregorio, A.V. Yeromin, K. Ohlsen, M. Lioudyno, S. Zhang, O. Safrina, J.A. Kozak, S.L. Wagner, M.D. Cahalan, G. Velicelebi, K.A. Stauderman. STIM1, an essential and conserved component of store-operated Ca~(2+) channel function. J Cell Biol. 2005;169:435–445.
66.J. Liou, M.L. Kim, W.D. Heo, J.T. Jones, J.W. Myers, J.E. Ferrell Jr.,T. Meyer. STIM is a Ca~(2+) sensor essential for Ca~(2+)-store- depletiontriggered Ca~(2+) influx. Curr Biol.2005;15: 1235–1241.
67. G.N. Huang, W. Zeng, J.Y. Kim, J.P. Yuan, L. Han, S. Muallem, P.F. Worley. STIM1 carboxyl-terminus activates native SOC, I(crac) and TRPC1 channels. Nat Cell Biol.2006;8;1003–1010.
68. S.L. Zhang, Y. Yu, J. Roos, J.A. Kozak, T.J. Deerinck, M.H. Ellisman, K.A. Stauderman, M.D. Cahalan. STIM1 is a Ca~(2+) sensor that activates CRAC channels and migrates from the Ca~(2+) store to the plasma membrane. Nature. 2005;437: 902–905.
69. M.M. Wu, J. Buchanan, R.M. Luik, R.S. Lewis. Ca~(2+) store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane. J Cell Biol. 2006;174;803–813.
70. Y. Baba, K. Hayashi,Y. Fujii, A. Mizushima, H.Watarai, M.Wakamori, T. Numaga, Y. Mori, M. Iino, M. Hikida, T. Kurosaki. Coupling of STIM1 to store-operated Ca~(2+) entry through its constitutive and inducible movement in the endoplasmic reticulum. Proc Natl Acad Sci USA. 2006;103:16704–16709.
71. M.A. Spassova, J. Soboloff, L.P. He, W. Xu, M.A. Dziadek, D.L. Gill. STIM1 has a plasma membrane role in the activation of store-operated Ca~(2+) channels. Proc Natl Acad Sci USA. 2006;103:4040–4045.
72. Smith IF, Plant LD, Boyle JP, Skinner RA, Pearson HA, Peers C. Chronic hypoxia potentiates capacitative Ca~(2+) entry in type-I cortical astrocytes. J Neurochem. 2003;85:1109-1116.
73. Lo KJ, Luk HN, Chin TY, Chueh SH. Store depletion-induced calcium influx in rat cerebellar astrocytes. Br J Pharmacol. 2002;135:1383-1392.
74. Sweeney M, McDaniel SS, Platoshyn O, Zhang S, Yu Y, Lapp BR, Zhao Y, Thistlethwaite PA, Yuan JX. Role of capacitative Ca~(2+) entry in bronchial contraction and remodeling. J Appl Physiol. 2002;92:1594-1602.
75. Hunton DL, Zou L, Pang Y, Marchase RB. Adult rat cardiomyocytes exhibit capacitative calcium entry. Am J Physiol Heart Circ Physiol. 2004;286:H1124-1132.
76. Smani T, Patel T, Bolotina VM. Complex regulation of store-operated Ca~(2+) entry pathway by PKC-epsilon in vascular SMCs. Am J Physiol Cell Physiol.2008;294:C1499-1508.
77. Lu W, Wang J, Shimoda LA, Sylvester JT.Differences in STIM1 and TRPC expression in proximal and distal pulmonary arterial smooth muscle are associated with differences in Ca~(2+) responses to hypoxia. Am J Physiol Lung Cell Mol Physiol. 2008;295:L104-113.
78. Kunichika N, Yu Y, Remillard CV, Platoshyn O, Zhang S, Yuan JX. Overexpression of TRPC1 enhances pulmonary vasoconstriction induced bycapacitative Ca~(2+) entry. Am J Physiol Lung Cell Mol Physiol. 2004;287:L962-969.
79. McElroy SP, Gurney AM, Drummond RM. Pharmacological profile of store-operated Ca(~(2+)) entry in intrapulmonary artery smooth muscle cells. Eur J Pharmacol.2008;584:10-20.
80.Fantozzi I, Zhang S, Platoshyn O, Remillard CV, Cowling RT, Yuan JX. Hypoxia increases AP-1 binding activity by enhancing capacitative Ca~(2+) entry in human pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2003 285:L740-754.
81. Lin MJ, Leung GP, Zhang WM, Yang XR, Yip KP, Tse CM, Sham JS. Chronic hypoxia-induced upregulation of store-operated and receptor-operated Ca~(2+) channels in pulmonary arterial smooth muscle cells: a novel mechanism of hypoxic pulmonary hypertension. Circ Res. 2004;95:496-505.
82. X. Wu, G. Babnigg, M.L. Villereal. Functional significance of human trp1 and trp3 in store-operated Ca~(2+) entry in HEK-293 cells. Am J Physiol Cell Physiol. 2000;278;C526–C536.
83.X. Wu, T.K. Zagranichnaya, G.T. Gurda, E.M. Eves, M.L. Villereal. A TRPC1/TRPC3-mediated increase in store-operated calcium entry is required for differentiation of H19-7 hippocampal neuronal cells. J Biol Chem. 2004;279 :43392–43402.
84. A. Fiorio Pla, D. Maric, S.C. Brazer, P. Giacobini, X. Liu, Y.H. Chang, I.S. Ambudkar, J.L. Barker. Canonical transient receptor potential 1 plays a role in basic fibroblast growth factor (bFGF)/FGF receptor-1- induced Ca~(2+) entry and embryonic rat neural stem cell proliferation. J Neurosci. 2005;25:2687–2701.
85. L. Vaca, A. Sampieri. Calmodulin modulates the delay period between release of calcium from internal stores and activation of calcium influx via endogenous TRP1 channels. J Biol Chem. 2002;277:42178–42187.
86. T.K. Zagranichnaya, X. Wu, M.L. Villereal. Endogenous TRPC1, TRPC3,and TRPC7 proteins combine to form native store-operated channels in HEK-293 cells. J Biol Chem. 2005;280:29559–29569.
87. L.I. Brueggemann, D.R. Markun, J.A. Barakat, H. Chen, K.L. Byron. Evidence against reciprocal regulation of Ca~(2+) entry by vasopressin in A7r5 rat aortic smooth-muscle cells. Biochem J. 2005;388:237-244.
88. C.L. Tu,W. Chang, D.D. Bikle. Phospholipase cgamma1 is required for activation of store-operated channels in human keratinocytes. J Invest Dermatol. 2005;124:187–197.
89. S. Thebault, A. Zholos, A. Enfissi, C. Slomianny, E. Dewailly, M. Roudbaraki, J. Parys, N. Prevarskaya. Receptor-operated Ca~(2+) entry mediated by TRPC3/TRPC6 proteins in rat prostate smooth muscle (PS1) cell line. J Cell Physiol. 2005;204:320–328.
90. X. Wang, J.L. Pluznick, P. Wei, B.J. Padanilam, S.C. Sansom. TRPC4 forms store-operated Ca~(2+) channels in mouse mesangial cells. Am J Physiol Cell Physiol. 2004;287:C357–C364.
91. H. Yang, S. Mergler, X. Sun, Z.Wang, L. Lu, J.A. Bonanno, U. Pleyer, P.S. Reinach. TRPC4 knockdown suppresses epidermal growth factorinduced store-operated channel activation and growth in human corneal epithelial cells. J Biol Chem. 2005;280:32230–32237.
92. G. Vazquez, G.S. Bird, Y. Mori, J.W. Putney Jr.Native TRPC7 channel activation by an inositol trisphosphate receptor-dependent mechanism. J Biol Chem. 2006;281:25250–25258.
93. J.P. Lievremont, G.S. Bird, J.W. Putney Jr.Canonical transient receptor potential TRPC7 can function as both a receptor- and store-operated channel in HEK-293 cells. Am J Physiol Cell Physiol. 2004;287:C1709–C1716.
94. J.P. Lievremont, T. Numaga, G. Vazquez, L. Lemonnier, Y. Hara, E. Mori, M. Trebak, S.E. Moss, G.S. Bird, Y. Mori, J.W. Putney Jr. The role of canonical transient receptor potential 7 in B-cell receptoractivated channels. J Biol Chem. 2005;280:35346–35351.
95. K. Kiselyov, X. Xu, G. Mozhayeva, T. Kuo, I. Pessah, G. Mignery, X. Zhu, L. Birnbaumer, S. Muallem. Functional interaction between InsP3 receptors and store-operated Htrp3 channels. Nature. 1998;396:478–482.
96. G. Vazquez, B.J. Wedel, M. Trebak, G. St John Bird, J.W. Putney Jr. Expression level of the canonical transient receptor potential 3 (TRPC3) channel determines its mechanism of activation. J Biol Chem. 2003;278: 21649–21654.
97. G. Vazquez, J.P. Lievremont, G. St John Bird, J.W. Putney Jr. Human Trp3 forms both inositol trisphosphate receptor-dependent and receptor-independent store-operated cation channels in DT40 avian B lymphocytes. Proc Natl Acad Sci USA. 2001; 98:11777–11782.
98.Salvaterra CG, Rubin LJ, Schaeffer J, and Blaustein MP. The influence of the transmembrane sodium gradient on the responses of pulmonary arteries to decreases in oxygen tension. Am Rev Respir Dis. 1989;139: 933–939.
99. Wang YX, Dhulipala PK, and Kotlikoff MI. Hypoxia inhibits the Na~+/Ca~(2+) exchanger in pulmonary artery smooth muscle cells. FASEB J. 2000;14: 1731–1740.
100.Kang TM, Park MK, and Uhm DY. Effects of hypoxia and mitochondrial inhibition on the capacitative calcium entry in rabbit pulmonary arterial smooth muscle cells. Life Sci. 2003;72: 1467–1479.
101. Dipp M, Nye PC, and Evans AM. Hypoxic release of calcium from the sarcoplasmic reticulum of pulmonary artery smooth muscle. Am J Physiol Lung Cell Mol Physiol. 2001;281: L318–L325.
102. Parekh AB and Penner R. Store depletion and calcium influx. Physiol Rev. 1997;77: 901–930.
103.Taraseviciene-Stewart L, Kasahara Y, Alger L, et al. Inhibition of the VEGF receptor 2 combined with chronic hypoxia causes cell death-dependent pulmonary endothelial cell proliferation and severe pulmonary hypertension. FASEB J.2001;15:427–438.
104. Satoh K, Kagaya Y, Nakano M, et al. Important role of endogenous erythropoietin system in recruitment of endothelial progenitor cells in hypoxia-induced pulmonary hypertension in mice. Circulation. 2006;113:1442–1450.
105. Meyrick B, Reid L. Development of Crotalaria pulmonary hypertension: hemodynamic and structural study. Am J Physiol.1980;239:692–702.
106. Buermans HP, Redout EM, Schiel AE, et al. Microarray analysis reveals pivotal divergent mRNA expression profiles early in the development of either compensated ventricular hypertrophy or heart failure. Physiol Genomics.2005;21:314–323.
107. Dorfmuller P, Perros F, Balabanian K, Humbert M. Inflammation in pulmonary arterial hypertension. Eur Respir J.2003;22:358–363.
108. Wright JL, Lawson L, Pare PD, et al. The structure and function of the pulmonary vasculature in mild chronic obstructive pulmonary disease. The effect of oxygen and exercise. Am Rev Respir Dis.1983;128:702–707.
109. Kessler R, Faller M, Weitzenblum E, et al. "Natural history" of pulmonary hypertension in a series of 131 patients with chronic obstructive lung disease. Am J Respir Crit Care Med.2001;164:219–224.
110.冉丕鑫,段生福。常压缺氧性大鼠肺动脉高压模型的建立。衡阳医学院学报,1994;3:229-231。
111.梁国容,邓静敏。建立缺氧性大鼠肺动脉高压模型的简易方法。广西医学,1997;2:244-245。
112.肖诗亮,高思海。常压缺氧性大鼠肺动脉高压模型的改进。临床心血管病杂志,2001;4:190。
113.Adnot S, Raffestin B, Eddahibi S, Braquet P, Chabrier PE. Loss of endothelium-dependent relaxant activity in the pulmonary circulation of rats exposed to chronic hypoxia. J Clin Invest.1991;87:155–162.
114. Savale L, Tu L, Rideau D, Izziki M, Maitre B, Adnot S, Eddahibi S. Impact of interleukin-6 on hypoxia-induced pulmonary hypertension and lung inflammation in mice. Respir Res.2009; 27:10:6.
115. Xu SZ, Beech DJ. TrpC1 is a membrane-spanning subunit of store-operated Ca~(2+) channels in native vascular smooth muscle cells. Circ Res.2001; 88: 84–87.
116. Brough GH, Wu S, Cioffi D, Moore TM, Li M, Dean N, Stevens T. Contribution of endogenously expressed Trp1 to a Ca~(2+)-selective, store-operated Ca~(2+) entry pathway. FASEB J. 2001; 15: 1727–1738.
117. Inoue R, Okada T, Onoue H, Hara Y, Shimizu S, Naitoh S, Ito Y, Mori Y. The transient receptor potential protein homologue TRP6 is the essential component of vascular alpha1-adrenoceptor-activated Ca~(2+)-permeable cation channel. Circ Res. 2001; 88: 325–332.
118. Jung S, Strotmann R, Schultz G, Plant TD .TRPC6 is a candidate channel involved in receptor-stimulated cation currents in A7r5 smooth muscle cells. Am J Physiol Cell Physiol. 2002; 282:C347–359.
119. Yu Y, Sweeney M, Zhang S, Platoshyn O, Landsberg J, Rothman A, Yuan JX ,PDGF stimulates pulmonary vascular smoothmuscle cell proliferation by upregulating TRPC6 expression. Am J Physiol Cell Physiol.2003; 284:C316–330.
120. Yu Y, Fantozzi I, Remillard CV, Landsberg JW, Kunichika N, Platoshyn O, Tigno DD,Thistlethwaite PA, Rubin LJ, Yuan JX. Enhanced expression of transient receptor potential channels in idiopathic pulmonary arterial hypertension. Proc Natl Acad Sci U S A. 2004;101:13861–13866.
121. Guilbert A, Dhennin-Duthille I, Hiani YE, Haren N, Khorsi H, Sevestre H, Ahidouch A, Ouadid-Ahidouch H. Expression of TRPC6 channels in human epithelial breast cancer cells. BMC Cancer. 2008;8:125.
122. Li M, Zacharia J, Sun X, Wier WG. Effects of siRNA knock-down of TRPC6 and InsP(3)R1 in vasopressin-induced Ca(~(2+)) oscillations of A7r5 vascular smooth muscle cells. Pharmacol Res. 2008;58:308-315.
1.J. Roos, P.J. DiGregorio, A.V. Yeromin, K. Ohlsen, M. Lioudyno, S. Zhang, O. Safrina, J.A. Kozak, S.L. Wagner, M.D. Cahalan, G. Velicelebi, K.A. Stauderman. STIM1, an essential and conserved component of store-operated Ca~(2+) channel function. J Cell Biol. 2005;169:435–445.
2.J. Liou, M.L. Kim, W.D. Heo, J.T. Jones, J.W. Myers, J.E. Ferrell Jr.,T. Meyer. STIM is a Ca~(2+) sensor essential for Ca~(2+)-store- depletiontriggered Ca~(2+) influx. Curr Biol.2005;15: 1235–1241.
3. G.N. Huang, W. Zeng, J.Y. Kim, J.P. Yuan, L. Han, S. Muallem, P.F. Worley. STIM1 carboxyl-terminus activates native SOC, I(crac) and TRPC1 channels. Nat Cell Biol.2006;8;1003–1010.
4. S.L. Zhang, Y. Yu, J. Roos, J.A. Kozak, T.J. Deerinck, M.H. Ellisman, K.A. Stauderman, M.D. Cahalan. STIM1 is a Ca~(2+) sensor that activates CRAC channels and migrates from the Ca~(2+) store to the plasma membrane. Nature. 2005;437: 902–905.
5. M.M. Wu, J. Buchanan, R.M. Luik, R.S. Lewis. Ca~(2+) store depletion causes STIM1 to accumulate in ER regions closely associated with the plasma membrane. J Cell Biol. 2006;174;803–813.
6. Y. Baba, K. Hayashi,Y. Fujii, A. Mizushima, H.Watarai, M.Wakamori, T. Numaga, Y. Mori, M. Iino, M. Hikida, T. Kurosaki. Coupling of STIM1 to store-operated Ca~(2+) entry through its constitutive and inducible movement in the endoplasmic reticulum. Proc Natl Acad Sci USA. 2006;103:16704–16709.
7. M.A. Spassova, J. Soboloff, L.P. He, W. Xu, M.A. Dziadek, D.L. Gill. STIM1 has a plasma membrane role in the activation of store-operated Ca~(2+) channels. Proc Natl Acad Sci USA.2006;103:4040–4045.
8. Smith IF, Plant LD, Boyle JP, Skinner RA, Pearson HA, Peers C. Chronic hypoxia potentiates capacitative Ca~(2+) entry in type-I cortical astrocytes. J Neurochem. 2003;85:1109-1116.
9. Lo KJ, Luk HN, Chin TY, Chueh SH. Store depletion-induced calcium influx in rat cerebellar astrocytes. Br J Pharmacol. 2002;135:1383-1392.
10. Sweeney M, McDaniel SS, Platoshyn O, Zhang S, Yu Y, Lapp BR, Zhao Y, Thistlethwaite PA, Yuan JX. Role of capacitative Ca~(2+) entry in bronchial contraction and remodeling. J Appl Physiol. 2002;92:1594-1602.
11. Hunton DL, Zou L, Pang Y, Marchase RB. Adult rat cardiomyocytes exhibit capacitative calcium entry. Am J Physiol Heart Circ Physiol. 2004;286:H1124-1132.
12. Smani T, Patel T, Bolotina VM. Complex regulation of store-operated Ca~(2+) entry pathway by PKC-epsilon in vascular SMCs. Am J Physiol Cell Physiol.2008;294:C1499-1508.
13. Lu W, Wang J, Shimoda LA, Sylvester JT.Differences in STIM1 and TRPC expression in proximal and distal pulmonary arterial smooth muscle are associated with differences in Ca~(2+) responses to hypoxia. Am J Physiol Lung Cell Mol Physiol. 2008;295:L104-113.
14. Kunichika N, Yu Y, Remillard CV, Platoshyn O, Zhang S, Yuan JX. Overexpression of TRPC1 enhances pulmonary vasoconstriction induced by capacitative Ca~(2+) entry. Am J Physiol Lung Cell Mol Physiol. 2004;287:L962-969.
15. McElroy SP, Gurney AM, Drummond RM. Pharmacological profile of store-operated Ca(~(2+)) entry in intrapulmonary artery smooth muscle cells. Eur J Pharmacol.2008;584:10-20.
16.Fantozzi I, Zhang S, Platoshyn O, Remillard CV, Cowling RT, Yuan JX. Hypoxia increases AP-1 binding activity by enhancing capacitative Ca~(2+) entry in human pulmonary artery endothelial cells. Am J Physiol Lung Cell Mol Physiol. 2003 285:L740-754.
17. X. Wu, G. Babnigg, M.L. Villereal. Functional significance of human trp1 and trp3 in store-operated Ca~(2+) entry in HEK-293 cells. Am J Physiol Cell Physiol. 2000;278;C526–C536.
18.X. Wu, T.K. Zagranichnaya, G.T. Gurda, E.M. Eves, M.L. Villereal. A TRPC1/TRPC3-mediated increase in store-operated calcium entry is required for differentiation of H19-7 hippocampal neuronal cells. J Biol Chem. 2004;279 :43392–43402.
19. A. Fiorio Pla, D. Maric, S.C. Brazer, P. Giacobini, X. Liu, Y.H. Chang, I.S. Ambudkar, J.L. Barker. Canonical transient receptor potential 1 plays a role in basic fibroblast growth factor (bFGF)/FGF receptor-1- induced Ca~(2+) entry and embryonic rat neural stem cell proliferation. J Neurosci. 2005;25:2687–2701.
20. L. Vaca, A. Sampieri. Calmodulin modulates the delay period between release of calcium from internal stores and activation of calcium influx via endogenous TRP1 channels. J Biol Chem. 2002;277:42178–42187.
21. T.K. Zagranichnaya, X. Wu, M.L. Villereal. Endogenous TRPC1, TRPC3, and TRPC7 proteins combine to form native store-operated channels in HEK-293 cells. J Biol Chem. 2005;280:29559–29569.
22. Golovina, V.A., O. Platoshyn, C.L. Bailey, Wang J, A. Limsuwan, M. Sweeney, L.J. Rubin and J.X. Yuan. Upregulated TRP and enhanced capacitative Ca(~(2+)) entry in human pulmonary artery myocytes during proliferation. Am J Physiol Heart Circ Physiol. 2001;280: H746-755.
23. Sweeney, M., Y. Yu, O. Platoshyn, S. Zhang, S.S. McDaniel and J.X. Yuan. Inhibition of endogenous TRP1 decreases capacitative Ca~(2+) entry and attenuates pulmonary artery smooth muscle cell proliferation. Am J Physiol Lung Cell Mol Physiol. 2002;283: L144-155.
24. L.I. Brueggemann, D.R. Markun, J.A. Barakat, H. Chen, K.L. Byron. Evidence against reciprocal regulation of Ca~(2+) entry by vasopressin in A7r5 rat aortic smooth-muscle cells. Biochem J. 2005;388:237-244.
25. C.L. Tu,W. Chang, D.D. Bikle. Phospholipase cgamma1 is required for activation of store-operated channels in human keratinocytes. J Invest Dermatol. 2005;124:187–197.
26. X. Wang, J.L. Pluznick, P. Wei, B.J. Padanilam, S.C. Sansom. TRPC4 forms store-operated Ca~(2+) channels in mouse mesangial cells. Am J Physiol Cell Physiol. 2004;287:C357–C364.
27. H. Yang, S. Mergler, X. Sun, Z.Wang, L. Lu, J.A. Bonanno, U. Pleyer, P.S. Reinach. TRPC4 knockdown suppresses epidermal growth factorinduced store-operated channel activation and growth in human corneal epithelial cells. J Biol Chem. 2005;280:32230–32237.
28. Lin MJ, Leung GP, Zhang WM, Yang XR, Yip KP, Tse CM, Sham JS. Chronic hypoxia-induced upregulation of store-operated and receptor-operated Ca~(2+) channels in pulmonary arterial smooth muscle cells: a novel mechanism of hypoxic pulmonary hypertension. Circ Res. 2004;95:496-505.
29. Guilbert A, Dhennin-Duthille I, Hiani YE, Haren N, Khorsi H, Sevestre H, Ahidouch A, Ouadid-Ahidouch H. Expression of TRPC6 channels in human epithelial breast cancer cells. BMC Cancer. 2008;8:125.
30. Li M, Zacharia J, Sun X, Wier WG. Effects of siRNA knock-down of TRPC6 and InsP(3)R1 in vasopressin-induced Ca(~(2+)) oscillations of A7r5 vascular smooth muscle cells. Pharmacol Res. 2008;58:308-315.
31. G. Vazquez, G.S. Bird, Y. Mori, J.W. Putney Jr.Native TRPC7 channel activation by an inositol trisphosphate receptor-dependent mechanism. J Biol Chem. 2006;281:25250–25258.
32. J.P. Lievremont, G.S. Bird, J.W. Putney Jr.Canonical transient receptor potential TRPC7 can function as both a receptor- and store-operated channel in HEK-293 cells. Am J Physiol Cell Physiol. 2004;287:C1709–C1716.
33. J.P. Lievremont, T. Numaga, G. Vazquez, L. Lemonnier, Y. Hara, E. Mori, M. Trebak, S.E. Moss, G.S. Bird, Y. Mori, J.W. Putney Jr. The role of canonical transient receptor potential 7 in B-cell receptoractivated channels. J Biol Chem. 2005;280:35346–35351.
34. K. Kiselyov, X. Xu, G. Mozhayeva, T. Kuo, I. Pessah, G. Mignery, X. Zhu, L. Birnbaumer, S. Muallem. Functional interaction between InsP3 receptors and store-operated Htrp3 channels. Nature. 1998;396:478–482.
35. G. Vazquez, B.J. Wedel, M. Trebak, G. St John Bird, J.W. Putney Jr. Expression level of the canonical transient receptor potential 3 (TRPC3) channel determines its mechanism of activation. J Biol Chem. 2003;278: 21649–21654.
36. G. Vazquez, J.P. Lievremont, G. St John Bird, J.W. Putney Jr. Human Trp3 forms both inositol trisphosphate receptor-dependent and receptor-independent store-operated cation channels in DT40 avian B lymphocytes. Proc Natl Acad Sci USA. 2001; 98:11777–11782.
37. Sham, J.S., B.R. Crenshaw, L.H. Deng, L.A. Shimoda and J.T. Sylvester. Effects of hypoxia in porcine pulmonary arterial myocytes: roles of KV channel and endothelin-1. Am J Physiol Lung Cell Mol Physiol. 2000;279: L262-272.
38. Archer, S.L. Diversity of phenotype and function of vascular smooth muscle cells. J Lab Clin Med. 1996;127: 524-529.
39. Barman, S.A. Potassium channels modulate hypoxic pulmonary vasoconstriction. Am J Physiol. 1998;275: L64-70.
40. Hasunuma, K., D.M. Rodman and I.F. McMurtry. Effects of K+ channel blockers on vasculartone in the perfused rat lung. Am Rev Respir Dis. 1991;144: 884-887.
41. Robertson, T.P., D. Hague, P.I. Aaronson and J.P. Ward. Voltage-independent calcium entry in hypoxic pulmonary vasoconstriction of intrapulmonary arteries of the rat. J Physiol. 2000;525: 669-680.
42. McMurtry, I.F., A.B. Davidson, J.T. Reeves and R.F. Grover. Inhibition of hypoxic pulmonary vasoconstriction by calcium antagonists in isolated rat lungs. Circ Res. 1976;38: 99-104.
43. Cornfield, D.N., T. Stevens, I.F. McMurtry, S.H. Abman and D.M. Rodman. Acute hypoxia causes membrane depolarization and calcium influx in fetal pulmonary artery smooth muscle cells. Am J Physiol. 1994;266: L469-475.
44. Salvaterra, C.G. and W.F. Goldman. Acute hypoxia increases cytosolic calcium in cultured pulmonary arterial myocytes. Am J Physiol. 1993;264: L323-328.
45. Post, J.M., C.H. Gelband and J.R. Hume. [Ca~(2+)]_i inhibition of K+ channels in canine pulmonary artery. Novel mechanism for hypoxia-induced membrane depolarization. Circ Res.1995;77: 131-139.
46. Gelband, C.H. and H. Gelband. Ca~(2+) release from intracellular stores is an initial step in hypoxic pulmonary vasoconstriction of rat pulmonary artery resistance vessels. Circulation.1997;96: 3647-3654.
47. Madden, J.A., M.S. Vadula and V.P. Kurup. Effects of hypoxia and other vasoactive agents on pulmonary and cerebral artery smooth muscle cells. Am J Physiol. 1992;263: L384-393.
48. Vadula, M.S., J.G. Kleinman and J.A. Madden. Effect of hypoxia and norepinephrine on cytoplasmic free Ca~(2+) in pulmonary and cerebral arterial myocytes. Am J Physiol. 1993;265: L591-597.
49. Jabr, R.I., H. Toland, C.H. Gelband, X.X. Wang and J.R. Hume. Prominent role of intracellular Ca~(2+) release in hypoxic vasoconstriction of canine pulmonary artery. Br J Pharmacol. 1997;122: 21-30.
50. Liu, Q., J.S. Sham, L.A. Shimoda and J.T. Sylvester. Hypoxic constriction of porcine distal pulmonary arteries: endothelium and endothelin dependence. Am J Physiol Lung Cell Mol Physiol. 2001;280: L856-865.
51. Morio, Y. and I.F. McMurtry. Ca~(2+) release from ryanodine-sensitive store contributes to mechanism of hypoxic vasoconstriction in rat lungs. J Appl Physiol. 2002;92: 527-534.
52. Yu, Y., M. Sweeney, S. Zhang, O. Platoshyn, J. Landsberg, A. Rothman and J.X. Yuan. PDGF stimulates pulmonary vascular smooth muscle cell proliferation by upregulating TRPC6 expression. Am J Physiol Cell Physiol. 2003;284: C316-330.
53.Wang J, Weigand L, Sylvester JT, Semenza GL, and Shimoda LA. HIF-1 Mediates hypoxia-induced TRPC expression and elevated intracellular Ca~(2+) in pulmonary arterial smooth muscle cells. Circulation Research. 2006;98:1528–1537.
54. Shimoda, L.A., D.J. Manalo, J.S. Sham, G.L. Semenza and J.T. Sylvester. Partial HIF-1alpha deficiency impairs pulmonary arterial myocyte electrophysiological responses to hypoxia. Am J Physiol Lung Cell Mol Physiol. 2001;281: L202-208.
55. Smirnov, S.V., T.P. Robertson, J.P. Ward and P.I. Aaronson. Chronic hypoxia is associated with reduced delayed rectifier K+ current in rat pulmonary artery muscle cells. Am J Physiol. 1994;266: H365-370.
56. Suzuki, H. and B.M. Twarog. Membrane properties of smooth muscle cells in pulmonary hypertensive rats. Am J Physiol. 1982;242: H907-915.
57. Michael, J.R., T.P. Kennedy, P. Buescher, I. Farrukh, R. Lodato, P.C. Rock, J. Gottlieb, G.Gurtner, S.M. de la Monte and G.M. Hutchins. Nitrendipine attenuates the pulmonary vascular remodeling and right ventricular hypertrophy caused by intermittent hypoxia in rats. Am Rev Respir Dis. 1986;133: 375-379.
58. Morio, Y., K.G. Morris and I.F. McMurtry. Acute hemodynamic effects of Y27632, a selective Rho-kinase inhibitor, in chronically hypoxic pulmonary hypertensive rats (abstract). FASEB J. 2002;16: A74.
59. Shimoda, L.A., J.S. Sham, T.H. Shimoda and J.T. Sylvester. L-type Ca~(2+) channels, resting [Ca~(2+)]_i, and ET-1-induced responses in chronically hypoxic pulmonary myocytes. Am J Physiol Lung Cell Mol Physiol. 2000;279: L884-894.
60.Wang J, Shimoda LA, and Sylvester JT. Capacitative calcium entry and TRPC channel proteins are expressed in rat distal pulmonary arterial smooth muscle. Am J Physiol. Lung Cell Mol Physiol. 2004;286: L848-L858.
61. Yu Y, Sweeney M, Zhang S, Platoshyn O, Landsberg J, Rothman A, Yuan JX ,PDGF stimulates pulmonary vascular smoothmuscle cell proliferation by upregulating TRPC6 expression. Am J Physiol Cell Physiol.2003; 284:C316–330.
62. Yu Y, Fantozzi I, Remillard CV, Landsberg JW, Kunichika N, Platoshyn O, Tigno DD,Thistlethwaite PA, Rubin LJ, Yuan JX. Enhanced expression of transient receptor potential channels in idiopathic pulmonary arterial hypertension. Proc Natl Acad Sci U S A. 2004;101:13861–13866