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肺高压大鼠肺动脉平滑肌细胞薄荷醇引起的胞浆游离Ca2+浓度增加幅度减低

陈改英, 焦海霞, 王明月, 王瑞幸, 林默君^*

福建医科大学生理学与病理生理学系，福州 350004

摘要

本文通过观察正常(control, CON)、慢性低氧(chronic hypoxia, CH)和野百合碱(monocrotaline, MCT)预处理肺高压大鼠肺动脉平滑肌细胞(pulmonary arterial smooth muscle cells, PASMCs)中，瞬时感受器电位M8 (transient receptor potential melastatin 8, TRPM8)通道特异激动剂薄荷醇(menthol, MT)引起的胞浆游离钙浓度([Ca2+]i)变化，探讨TRPM8通道在肺高压发病过程中的作用。大鼠处于CH环境或一次性腹腔注射MCT制备肺高压大鼠模型；原代培养CON和肺高压大鼠PASMCs，观察MT激动TRPM8通道引起的PASMCs [Ca2+]i变化，以及TRPM8通道特异阻断剂N-(4-叔-丁基苯基)-4-(3-氯吡啶-2-基)哌嗪-1-甲酰胺[N-(4-tert-butylphenyl)-4-(3-chloropyridin-2-yl)piperazine-1-carboxamide, BCTC]对MT引起[Ca2+]i变化作用的抑制效应；免疫组化检测TRPM8的细胞定位。结果显示，在2 mmol/L Ca2+和无Ca2+台氏液中，MT都可以引起CON组PASMCs的[Ca2+]i增高，且这两种增高作用均可以被BCTC阻断；在2 mmol/L Ca2+和无Ca2+台氏液中，与CON组相比，CH组和MCT组MT引起PASMCs的[Ca2+]i增高幅度均显著减小。免疫组化细胞定位实验显示，PASMCs除胞核外其余部分均有TRPM8棕黄色阳性表达。以上结果提示，PASMCs上的TRPM8通道既存在于胞膜，也存在于肌浆网；CH和MCT预处理可以分别减少PASMCs上TRPM8通道介导的Ca2+内流与Ca2+释放。

关键词： 肺高压; 瞬时感受器电位M8; 钙; 低氧; 野百合碱; 薄荷醇

分类号：R363

[Decreased amplitude of [Ca2+]i elevation induced by menthol in pulmonary arterial smooth muscle cells of pulmonary hypertensive rats.] [Article in Chinese]

CHEN Gai-Ying, JIAO Hai-Xia, WANG Ming-Yue, WANG Rui-Xing, LIN Mo-Jun^*

Department of Physiology and Pathophysiology, Fujian Medical University, Fuzhou 350004, China

Abstract

The study was designed to explore the alteration of intracellular calcium concentration ([Ca²⁺]_i), induced by transient receptor potential melastatin 8 (TRPM8) channel-specific agonist menthol, in pulmonary arterial smooth muscle cells (PASMCs) between control and pulmonary hypertensive (PH) rats. PH rat models were established by means of chronic hypoxia (CH) and monocrotaline (MCT) injection, respectively. PASMCs from control and PH rats were cultured. The change of [Ca²⁺]_i in PASMCs induced by menthol, and the effect of TRPM8 channel-specific antagonist BCTC on the change of [Ca²⁺]_i, were observed. Cellular localization of TRPM8 was examined by using immunohistochemistry. Results showed that menthol increased [Ca²⁺]_i in the control PASMCs both in Ca²⁺-normal and Ca²⁺-free Tyrode's solutions, and at the same time BCTC could inhibit these two kinds of elevations. Compared with the control group, elevations of [Ca²⁺]_i were decreased notably in CH- and MCT-pretreated PASMCs superfused with 2 mmol/L Ca²⁺- or 0 Ca²⁺-Tyrode's solutions. Immunohistochemical localization experiments showed that the whole PASMCs were dyed brown except for the nucleus. This study verified that TRPM8 exists both in membrane and sarcoplasmic reticulum of PASMCs. In addition, CH- and MCT-pretreatment could independently down-regulate the Ca²⁺ influx and Ca²⁺ release mediated by TRPM8 channel.

Key words: hypertension; pulmonary; TRPM8; calcium; hypoxia; monocrotaline

收稿日期：2013-09-25　　录用日期：2013-11-19

通讯作者：林默君　　E-mail: mjlin@mail.fjmu.edu.cn

引用本文：

陈改英, 焦海霞, 王明月, 王瑞幸, 林默君. 肺高压大鼠肺动脉平滑肌细胞薄荷醇引起的胞浆游离Ca2+浓度增加幅度减低[J]. 生理学报 2014; 66 (3): 267-275.

CHEN Gai-Ying, JIAO Hai-Xia, WANG Ming-Yue, WANG Rui-Xing, LIN Mo-Jun. [Decreased amplitude of [Ca2+]i elevation induced by menthol in pulmonary arterial smooth muscle cells of pulmonary hypertensive rats.] [Article in Chinese]. Acta Physiol Sin 2014; 66 (3): 267-275 (in Chinese with English abstract).