胞外酸化对体外培养大鼠关节软骨细胞增殖与凋亡及组织蛋白酶K mRNA表达的影响

中国临床药理学与治疗学 ›› 2012, Vol. 17 ›› Issue (1): 34-39.

胞外酸化对体外培养大鼠关节软骨细胞增殖与凋亡及组织蛋白酶K mRNA表达的影响

荣超¹, 陈飞虎¹, 江晟¹, 胡伟^1,2, 葛金芳¹, 张晨晨¹, 吴繁荣¹

¹安徽医科大学药学院,合肥 230032,安徽;
²安徽医科大学第二附属医院药剂科,合肥 230601,安徽

收稿日期:2011-10-28 修回日期:2011-11-24 出版日期:2012-01-26 发布日期:2012-02-16
通讯作者: 陈飞虎,男,博士,教授,博士生导师,研究方向:分子药理学。Tel: 0551-5161115 E-mail: cfhchina@sohu.com
作者简介:荣超,男,硕士,研究方向:分子药理学。Tel: 13515645451 E-mail: rongchaochina@163.com
基金资助:
国家自然科学基金资助(30873080)

Effect of extracellular acidification on the proliferation, apoptosis and Cathepsin K mRNA of rat articular chondrocytes in vitro

RONG Chao¹, CHEN Fei-hu¹, JIANG Sheng¹, HU Wei^1,2, ZHANG Cheng-cheng¹, WU Fan-rong¹

¹School of Pharmacy, Anhui Medical University, Hefei 230032, Anhui, China;
²the Second Hospital Affiliated to Anhui Medical University, Hefei 230601, Anhui, China

Received:2011-10-28 Revised:2011-11-24 Online:2012-01-26 Published:2012-02-16

摘要/Abstract

摘要： 目的: 探讨不同胞外pH环境对体外培养的大鼠关节软骨细胞增殖、凋亡的影响,以及大鼠关节软骨细胞中组织蛋白酶K的表达与胞外酸化的关系。方法: Ⅱ型胶原酶消化法从大鼠关节软骨中分离软骨细胞;细胞传代培养后分为4组分别在pH7.4、pH6.5、pH6.0、pH5.5不同的胞外酸化环境下培养3 h,MTT法检测软骨细胞活力,Hoechst33258染色观察细胞凋亡形态,Annexin-V/PI双染法检测软骨细胞的凋亡率,实时荧光定量PCR检测组织蛋白酶K基因的表达。结果: 与pH7.4 组比较其他各组软骨细胞增殖均受到抑制,其中pH5.5、pH6.0与pH7.4相比差异有重要的统计学意义(P<0.01),pH5.5和pH6.0组可见较多的凋亡细胞,且流式细胞仪检测的凋亡率明显高于pH7.4组(P<0.01);组织蛋白酶K的表达与酸性程度成正相关。结论: 胞外酸化环境下能明显抑制体外培养软骨细胞增殖、促进软骨细胞凋亡,并且酸性环境能增加组织蛋白酶K的表达,这为类风湿关节炎中关节软骨的破坏发生机制提供了实验依据。

关键词: 胞外酸化, 细胞凋亡, 类风湿关节炎, 软骨细胞

Abstract: AIM: To investigate the effects of various extracellular solutions on rat chondrocytes proliferation and apoptosis in vitro, and relation of Cathepsin K mRNA expression and extracellular acidosis.METHODS: Articular chondrocytes were obtained from rats using type II collagenase digestion method. The chondrocytes were divided into four groups and incubated in different pH extracelluar solution (pH 7.4, pH6.5, pH6.0, pH5.5) for three hours. MTT assay was employed to determine the proliferation of articular chondrocytes in acidic solutions. Effects of extracellular acidosis on chondrocyte apoptosis were observed by Hoechst 33258 and Annexin V/PI staining. The expression of Cathepsin K mRNA were detected by real time reverse transcription PCR.RESULTS: Proliferation of articular chondrocytes were inhibited observably by acidosis (P<0.05). Apoptosis rate of articular chondrocytes were increased comparing with control group (P<0.05). The expression of Cathepsin K mRNA level gradually increased following the degree of acidity in extracellular solution.CONCLUSION: Extracellular acidosis can restrain the proliferation and promote the apoptosis of articular chondrocytes in vitro, and increase the expression of Cathepsin K mRNA. It has provided an experimental evidence on mechanism bone destruction in Rheumatoid arthritis.

Key words: Extracellular acidification, Apoptosis, Rheumatoid arthritis, Chondrocytes

中图分类号:

R965.2

荣超, 陈飞虎, 江晟, 胡伟, 葛金芳, 张晨晨, 吴繁荣. 胞外酸化对体外培养大鼠关节软骨细胞增殖与凋亡及组织蛋白酶K mRNA表达的影响[J]. 中国临床药理学与治疗学, 2012, 17(1): 34-39.

RONG Chao, CHEN Fei-hu, JIANG Sheng, HU Wei, ZHANG Cheng-cheng, WU Fan-rong. Effect of extracellular acidification on the proliferation, apoptosis and Cathepsin K mRNA of rat articular chondrocytes in vitro[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2012, 17(1): 34-39.

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