Mechanisms of the inhibitory effect of epigallocatechin-3-gallate on cultured human vascular smooth muscle cell invasion
Mechanisms of the inhibitory effect of epigallocatechin-3-gallate on cultured human vascular smooth muscle cell invasion
复制标题
DOI：
10.1161/01.atv.0000179675.49619.9b
复制
发表时间：
2005-09-01
影响因子：
8.7
通讯作者：
Iguchi, A
中科院分区：
文献类型：
--
作者：
Cheng, XW;Kuzuya, M;Iguchi, A
目的：尽管我们最近发现，在大鼠球囊损伤模型中，儿茶素的应用减少了新生内膜的形成，但其确切的分子机制仍不清楚。在本研究中，我们试图利用体外SMC侵袭系统来确定这些机制。方法和结果-采用Boyden小室实验检测儿茶素对SMC侵袭行为的影响。EGCG以浓度依赖的方式抑制SMC通过胶原胶的侵袭活性。明胶酶谱、胶原酶谱和Western印迹结果显示，即使在0.1~0.3mU/L的浓度范围内，EGCG仍能抑制侵袭实验中和培养的SMC条件培养液中前基质金属蛋白酶-2的激活以及基质金属蛋白酶-2和膜型-1-基质金属蛋白酶的活性。EGCG可抑制MT1-MMPcat依赖的原-基质金属蛋白酶-2的激活。EGCG上调基质金属蛋白酶组织抑制因子-2(TIMP-2)蛋白表达。反向酶谱分析表明，TIMP-2TO的表达增加是通过活性的增加来验证的。TIMP-2活性下调提示TIMP-2表达上调可能是EGCG抑制SMC侵袭的主要机制之一。结论EGCG靶向多种基质金属蛋白酶介导的SMC细胞事件，通过上调TIMP-2表达调节基质金属蛋白酶活性，为SMC侵袭提供了新的主要机制。
Objective - Although we recently showed that the administration of catechins reduced the neointimal formation in a rat balloon-injury model, the precise molecular mechanisms are largely unknown. In the present study, we tried to determine these mechanisms using an in vitro SMC invasion system.Methods and Results - Boyden chamber assay was used to examine the effect of catechins on the invasive behavior of SMCs. The invasive activity of SMCs through collagen gel was restrained by EGCG in a concentration-dependent manner. The data from gelatin and collagen zymography and Western blot revealed that EGCG blocks the activation of pro-matrix metalloproteinase (MMP)-2 during an invasion assay and in the conditioned medium of cultured SMCs as well as the activities of MMP-2 and membrane type 1-MMP (MT1-MMP) even at 0.1 to 0.3 mu mol/L of EGCG. EGCG was found to restrain MT1-MMPcat-dependent pro-MMP-2 activation. EGCG upregulated the expression of tissue inhibitor of MMP-2 (TIMP-2) protein. Reverse zymography showed that the increased TIMP-2 to expression was validated by an increased activity. The data from decreased TIMP-2 activity using its siRNA suggested that upregulation of TIMP-2 expression may be one of the major mechanisms for inhibition of SMC invasion by EGCG.Conclusions - These results indicate that EGCG targets multiple MMP-mediated SMC cellular events and provides a new major mechanism for the SMC invasion through upregulation of TIMP-2 expression to modulate MMP activity.