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高糖促进成年大鼠心脏成纤维细胞periostin 表达及其信号转导机制

邹平, 吴立玲, 吴丹, 凡栋, 崔晓兵, 周允, 王程, 李丽

北京大学医学部生理学与病理生理学系，教育部分子心血管病重点实验室，北京 100191；泸州医学院病理生理学教研室，泸州 646000

摘要

心肌纤维化是糖尿病心肌病时心室舒缩功能障碍的主要机制之一。心脏成纤维细胞分泌的一种细胞外基质蛋白periostin 与心肌纤维化及间质重塑密切相关。本研究旨在观察高糖对心脏成纤维细胞periostin 表达的影响及其相关信号转导机制。在离体培养的成年大鼠心脏成纤维细胞，采用低糖(5.5 mmol/L)和高糖(25 mmol/L)干预，RT-PCR 检测periostinmRNA 表达，免疫印迹法检测periostin 蛋白表达，荧光法检测细胞内活性氧(reactive oxygen species, ROS)含量。结果显示，高糖刺激12 h 可使periostin mRNA表达增加117.26%，刺激24 h 可使其蛋白表达上调93.12%，高糖刺激30 min 可显著促进心脏成纤维细胞内ROS 的产生，蛋白激酶C (protein kinase C, PKC)抑制剂白屈菜红碱可减少高糖诱导的ROS 生成增加。高糖诱导的periostin 蛋白表达增加可被白屈菜红碱或ROS 清除剂N- 乙酰-L- 半胱氨酸(N-acetylcysteine, NAC)所抑制。高糖刺激30 和60 min 可显著增加磷酸化c-Jun 氨基末端激酶(c-jun N-terminal protein kinase, JNK)的含量，应用白屈菜红碱或NAC 可抑制高糖诱导的磷酸化JNK 含量增加。JNK 抑制剂SP600125 预孵育明显抑制了高糖诱导的periostin 蛋白表达增加。上述结果提示，高糖刺激可促进periostin 的mRNA和蛋白表达，激活PKC/ROS/JNK 通路介导高糖诱导的periostin 表达增加。

关键词： 高糖; 心脏成纤维细胞; p e r ios t in; 信号通路

分类号：R363.2

[High glucose increases periostin expression and the related signal pathway in adult rat cardiac fibroblasts.] [Ariticle in Chinese]

ZOU Ping, WU Li-Ling, WU Dan, FAN Dong, CUI Xiao-Bing, ZHOU Yun, WANG Cheng, LI Li

Department of Physiology and Pathophysiology, Peking University Health Science Center and Key Laboratory of Molecular Cardiovascular Sciences, Ministry of Education, Beijing 100191, China； Department of Pathophysiology, Luzhou Medical College, Luzhou 646000, China

Abstract

Cardiac fibrosis is a major mechanism contributing to myocardial systolic and diastolic dysfunction in diabetic cardiomyopathy.Periostin is a novel extracellular matrix protein, secreted from cardiac fibroblasts, and closely related with cardiac fibrosis and remodeling.The present study aimed to investigate the effect of high glucose on periostin expression and the related signal transduction pathwayin cardiac fibroblasts. Adult rat cardiac fibroblasts were cultured and stimulated with high glucose (25 mmol/L). The mRNA and proteinexpressions of periostin were detected by RT-PCR and Western blot, respectively. Intracellular reactive oxygen species (ROS)production was measured using 2, 7-dichlorofluorescein diacetate (DCF-DA), an oxidant-sensitive fluorescent probe. Results showedthat the mRNA expression of periostin in adult rat cardiac fibroblasts was increased by 117.26% when treated with high glucose for 12 h.Incubation with high glucose for 24 h enhanced periostin protein expression by up to 93.12%. High glucose induced the production ofROS in adult rat cardiac fibroblasts, which was reduced by chelerythrine (CLT), a protein kinase C (PKC) inhibitor. High glucoseinducedperiostin protein expression was decreased significantly when pretreated with CLT or N-acetylcysteine (NAC), a ROS scavenger. The phosphorylation of c-jun N-terminal protein kinase (JNK) was increased markedly when stimulated with high glucosefor 30 and 60 min, which was abolished when pretreated with CLT or NAC. SP600125, a specific JNK inhibitor, significantly decreasedperiostin expression induced by high glucose. In conclusion, high glucose stimulates periostin protein expression via a PKC/ROS/JNKdependentpathway in adult rat cardiac fibroblasts.

Key words: high glucose; cardiac fibroblasts; periostin; signal pathway

收稿日期：2009-12-22　　录用日期：2010-04-02

通讯作者：李丽　　E-mail: lilyby@163.com

引用本文：

邹平, 吴立玲, 吴丹, 凡栋, 崔晓兵, 周允, 王程, 李丽. 高糖促进成年大鼠心脏成纤维细胞periostin 表达及其信号转导机制[J]. 生理学报 2010; 62 (3): 247-254.

ZOU Ping, WU Li-Ling, WU Dan, FAN Dong, CUI Xiao-Bing, ZHOU Yun, WANG Cheng, LI Li. [High glucose increases periostin expression and the related signal pathway in adult rat cardiac fibroblasts.] [Ariticle in Chinese] . Acta Physiol Sin 2010; 62 (3): 247-254 (in Chinese with English abstract).