整合素αVβ3在细胞外基质调控人黑色素瘤MMP-2表达和活化中的作用

doi:国家自然科学基金资助课题（30370548）

整合素α_Vβ₃在细胞外基质调控人黑色素瘤MMP-2表达和活化中的作用

梅红¹，郭玉芳²，郑贤红³ ，王心蕊¹，张丽红¹，朱桂彬¹，王建民^△，李一雷^1*

1. 吉林大学基础医学院病理生物学教育部重点实验室, 吉林长春130021；2. 陕西医学高等专科学校病理学教研室，陕西西安710068；3. 吉林大学基础医学院病原生物学教研室, 吉林长春130021

收稿日期:2005-04-08 修回日期:1900-01-01 出版日期:2006-03-28 发布日期:2006-03-28
通讯作者: 李一雷^1*

Effect of integrin α_Vβ₃ on expression and activation of matrix metalloproteinase-2 of human melanoma cells regulated by extracellular matrix

MEI Hong¹, GUO Yu-fang²,ZHENG Xian-hong³, WANG Xin-rui¹，ZHANG Li-hong¹, ZHU Gui-bin¹, WANG Jian-min^△, LI Yi-lei^1*

Received:2005-04-08 Revised:1900-01-01 Online:2006-03-28 Published:2006-03-28
Contact: LI Yi-lei^1*

摘要/Abstract

摘要： 目的：探讨整合素α_Vβ₃在细胞外基质调控人黑色素瘤细胞基质金属蛋白酶2（matrix metalloproteinase-2，MMP-2）表达和活化中的作用和地位。方法：选择黑色素瘤细胞系M21及突变型M21-L（M21-L不表达整合素α_Vβ₃），通过观察2个细胞系在同一试验条件下的差异，研究整合素α_Vβ₃在黑色素瘤细胞系M21与M21-L及其在MMP-2表达和活化中的作用；应用酶谱分析法检测黑色素瘤细胞系M21与M21-L，在包被I型胶原（type I collagen, Icol）、纤维黏连蛋白(fibronectin, FN)及ECM胶3种细胞外基质成分后，黑色素瘤细胞系M21与M21-L两者MMP‐2表达量及活性的变化；通过RT-PCR方法检测包被后黑色素瘤细胞系M21与M21-L两者MT1-MMP mRNA表达水平的变化；通过Boyden小室膜侵袭实验观察黑色素瘤细胞系M21和M21-L细胞侵袭性差异。结果：培养于三维聚合Ⅰ型胶原、纤维黏连蛋白及ECM胶的M21细胞酶谱分析结果中出现了62 000的MMP-2 的活化带，其膜型基质金属蛋白酶1(membrane‐typematrixmetalloproteinase-1,MT1-MMP) mRNA表达均较没有包被3种细胞外基质的空白对照组增高；而M21-L在包被细胞外基质成分前后未出现MMP-2的活化，其MT1-MMP mRNA表达水平变化无统计学意义；Boyden小室检测表明，M21细胞侵袭力明显高于M21-L细胞。结论：整合素α_Vβ₃参与了人黑色素瘤细胞M21的MMP-2表达和活化过程，是细胞外基质成分调控黑色素瘤细胞MMP-2表达和活化的必要条件。

关键词: 玻连蛋白, 细胞外基质, 明胶酶A

Abstract: Objective To study the role of α_Vβ₃ integrin in extracellular matrix (ECM) regulation of the expression and activation of matrix metalloproteinase-2 (MMP-2). Methods The difference between human melanoma cells M21 and M21-L cells was that M21-L didn′t express α_Vβ₃ integrin. Human melanoma cells M21 and M21-L cells were cultured on three dimensional (3D) type I collagen gel, fibronectin (FN) and ECM gel. To prepare serum-free conditioned medium (SFCM), cells were harvested by trypsinization and counted using a hemacytometer and 2×10⁵ cells/well were seeded in 24-well tissue culture plates coated with these gels,respectively. After 12　h, cells were washed three times with phosphate-buffered saline, followed by incubation in serum-free medium. SFCM was collected 48 h later and stored at -80℃ until being used. Then,the expression of MMP-2 was examined by zymography, and the expression of MT1-MMP mRNA was examined by reverse transcription polymerase chain reaction (RT-PCR). Results Induction of MMP-2 activation was specific to human melanoma cells M21 cultured on type I collagen gel， FN and ECM gel. Pro-MMP-2 protein was expressed in human melanoma cells M21-L, but no active from of MMP-2 was detectable. MT1-MMP mRNA was expressed in human melanoma cells M21, and enhanced expression of MT1-MMP mRNA was seen in cells cultured on type I collagen gel，FN and ECM gel. MT1-MMP mRNA was expressed in human melanoma cells M21-L,but cells cultured on these gels did not influence the expression of MT1-MMP mRNA. Conclusion These ECM components can regulate expression and activiation of MMP-2 by integrin α_Vβ₃.

Key words: vitronectin, extracellular matrix, gelatinase A

中图分类号:

R363

梅红，郭玉芳，郑贤红，王心蕊，张丽红，朱桂彬，王建民，李一雷. 整合素α_Vβ₃在细胞外基质调控人黑色素瘤MMP-2表达和活化中的作用[J]. J4, 2006, 32(2): 182-185.

MEI Hong, GUO Yu-fang,ZHENG Xian-hong, WANG Xin-rui，ZHANG Li-hong, ZHU Gui-bin, WANG Jian-min, LI Yi-lei. Effect of integrin α_Vβ₃ on expression and activation of matrix metalloproteinase-2 of human melanoma cells regulated by extracellular matrix[J]. J4, 2006, 32(2): 182-185.

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整合素α_Vβ₃在细胞外基质调控人黑色素瘤MMP-2表达和活化中的作用

Effect of integrin α_Vβ₃ on expression and activation of matrix metalloproteinase-2 of human melanoma cells regulated by extracellular matrix

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