静压力下兔髁突软骨细胞COL2A1、SOX9、COL1A1和ALP的表达变化

中国口腔颌面外科杂志 ›› 2014, Vol. 12 ›› Issue (4): 295-300.

静压力下兔髁突软骨细胞COL2A1、SOX9、COL1A1和ALP的表达变化

黄林剑¹, 李辉¹, 谢千阳¹, 张旻², 陈永进², 杨驰¹, 蔡协艺¹

1.上海交通大学医学院附属第九人民医院·口腔医学院口腔外科,上海市口腔医学重点实验室,上海 200011;
2.第四军医大学口腔医学院,陕西西安 710032

收稿日期:2014-01-15 出版日期:2014-07-10 发布日期:2014-08-20
通讯作者: 蔡协艺,Tel:021-23271699-5705,E-mail:caixieyi27@126.com
作者简介:黄林剑（1987-）,男,在读硕士研究生,E-mail:dshuanglinjian@126.com
基金资助:
国家自然科学基金（81200766,81070848）; 上海市科学技术委员会资助项目（08DZ2271100,114119a3800,13140902702,13XD1402300）

Effect of static pressure on expression of COL2A1, SOX9, COL1A1 and ALP in temporomandibular condylar chondrocytes in rabbit

HUANG Lin-jian¹, LI Hui¹, XIE Qian-yang¹, ZHANG Min², CHENG Yong-jin², YANG Chi¹, CAI Xie-yi

1.Department of Oral Surgery, Ninth People’s Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology. Shanghai 200011;
2. College of Stomatology, The Fourth Military Medical University. Xi’an 710032, Shaanxi Province, China

Received:2014-01-15 Online:2014-07-10 Published:2014-08-20
Supported by:
Supported by National Natural Science Foundation of China (81200766,81070848), and Research Fund of Science and Technology Committee of Shanghai Municipality (08DZ2271100, 114119a3800, 13140902702, 13XD1402300)

摘要/Abstract

摘要： 目的研究兔髁突软骨细胞对静压力的分子生物学响应。方法：体外分离培养4周龄健康雌性新西兰大白兔髁突软骨细胞,使用形态学观察,COL2A1和SOX9免疫细胞化学染色方法对P2代髁突软骨细胞进行鉴定;100kPa静压力条件下,分别对P2代髁突软骨细胞进行0、1、2、3和4 h的加压处理;采用CCK-8检测压力加载后各组细胞活性的变化;通过Western免疫印迹分析髁突软骨细胞COL2A1、SOX9、COL1A1和ALP的表达变化。采用SPSS19.0软件包对数据进行统计学分析。结果：兔髁突软骨细胞呈多角形,“铺路石”样排列,细胞爬片COL2A1和SOX9免疫细胞化学染色鉴定结果为阳性。100kPa静压力加载1 h时,细胞活性显著降低（P=0.04）,但在2~4 h后,细胞活性恢复并与对照组无显著差异。Western印迹结果显示,压力加载3 h和4 h时,COL2A1、SOX9、COL1A1和ALP的表达显著升高。其中,在压力加载4 h组ALP的表达量比3 h组低。结论：兔髁突软骨细胞对压力微环境的改变具有一定的适应能力,100kPa静压力加载4 h不会对髁突软骨细胞活性产生不可逆损伤。适宜的压力加载对髁突软骨细胞的成软骨和成骨能力有促进作用。髁突软骨细胞压力微环境的改变,可能影响颞下颌关节适应性改建和颞下颌关节紊乱病的病理过程。

关键词: 髁突, 软骨细胞, 静压力, COL2A1, SOX9, COL1A1, ALP

Abstract: PURPOSE: To study the cellular events of condylar chondrocytes during the alteration of pressure microenvironment. METHODS: Condylar chondrocytes were aseptically dissected and cultured from the temporomandibular joint (TMJ) of 4-week-old New Zealand white rabbit in vitro. The chondrocytes of P2 were identified by morphological analysis, immunocytochemical staining of COL2A1 and SOX9. The monolayer of condylar chondrocytes was subjected to static pressure of 100kPa for different times (0 h, 1 h, 2 h, 3 h and 4 h). The viability of chondrocytes was assessed by CCK-8 staining. Expression of COL2A1, SOX9, COL1A1 and ALP was examined by Western blot. One-way ANOVA was performed using SPSS19.0 software package. RESULTS: The condylar chondrocytes of rabbits were polygon in shape and displayed cobble-stone morphology. Immunocytochemical staining of COL2A1 and SOX9 was positive in the slides of cells. Compared with 0 h, the viability of chondrocytes significantly decreased after being compressed for 1 h (P=0.04), whereas it returned to normal after 2 to 4 h. Western blot results showed that exposure of chondrocytes to 100 kPa for 3 h and 4 h resulted in significant up-regulation of COL2A1, SOX9, COL1A1 and ALP. Moreover, the expression of ALP at 4 h reduced compared with at 3 h. CONCLUSIONS: The condylar chondrocytes are good at adapting to the alteration of pressure microenvironment. It is sure that a suitable static pressure loading can accelerate chondrogenesis and osteogenesis. And this may influence the growth remodeling of TMJ and the pathological condition of temporomandibular joint disorder (TMD).

Key words: Condyle, Chondrocytes, Static pressure, COL2A1, SOX9, COL1A1, ALP

中图分类号:

R782.2

黄林剑, 李辉, 谢千阳, 张旻, 陈永进, 杨驰, 蔡协艺. 静压力下兔髁突软骨细胞COL2A1、SOX9、COL1A1和ALP的表达变化[J]. 中国口腔颌面外科杂志, 2014, 12(4): 295-300.

HUANG Lin-jian, LI Hui, XIE Qian-yang, ZHANG Min, CHENG Yong-jin, YANG Chi, CAI Xie-yi. Effect of static pressure on expression of COL2A1, SOX9, COL1A1 and ALP in temporomandibular condylar chondrocytes in rabbit[J]. China J Oral Maxillofac Surg, 2014, 12(4): 295-300.

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