基质金属蛋白酶-9在胃癌组织中的表达及其基因沉默胃癌细胞克隆株的构建  

Matrix metalloproteinase-9 expression in gastric cancer and construction of the gastric cancer cell line with silenced matrix metalloproteinase-9 gene

作　　者：刘凯华[1] 黄卫锋[1] 黄安乐[1] 蔡建春[1] 

机构地区：[1]厦门大学附属第一医院厦门市肿瘤中心肿瘤外科福建医科大学第一临床医学院,361003

出　　处：《中华实验外科杂志》2012年第10期1994-1996,共3页Chinese Journal of Experimental Surgery

基　　金：福建省自然科学基金资助项目（200919004）

摘　　要：目的观察基质金属蛋白酶-9（MMP-9）在胃癌发生、发展中的作用；构建MMP-9基因沉默质粒；建立MMP-9基因沉默的BGC823细胞。方法采用免疫组织化学法在46例人胃癌新鲜标本中检测MMP-9的蛋白表达水平，采用化学合成的方法合成3条沉默MMP-9的寡聚核苷酸序列，插入到短发卡RNA（shRNA）表达载体pGenesil上，运用荧光显微镜、明胶酶电泳、逆转录-聚合酶链反应（RT—PCR）分别检测3条质粒的沉默效率；选取沉默效率最佳的1个质粒转染BGC823进行稳定细胞株的筛选及鉴定。结果46例胃癌组织MMP-9蛋白阳性率为100％，其中57％（26／46）呈增强表达；筛选到1条MMP-9沉默最为显著的shRNA表达质粒；成功构建MMP-9基因沉默的BGC823胃癌细胞克隆。结论MMP-9基因在胃癌发生、发展及浸润、转移过程中可能扮演重要角色。Objective To investigate the roles of matrix metallopmteinase-9 （MMP-9） in gastric cancer development, to construct MMP-9 short hairpin RNA （shRNA） expression plasmids and to establish the gastric cancer cell line with silenced MMP-9 gene expression. Methods The expression level of MMP- 9 in 46 fresh specimens was detected by using immunohistoehemistry. Three oligonucleotides which silenced MMP-9 gene expression were synthesized chemically and inserted into shRNA expression vector pGenesil to construct MMP-9 shRNA plasmids. Immunofluorescence, gelatin zymography and reverse transeription-polymerase chain reaction （RT-PCR） were applied to detect the MMP-9 silencing efficiency. The MMP-9 knockdown plasmid that had a high efficiency in silencing MMP-9 gene expression was chosen and transfected into BGC823 to establish the cell line with silenced MMP-9 gene expression. Results MMP-9 protein was expressed in 46 cases of gastric cancers, and strong expression was found in 57% （26/46） of the gastric cancers. A shRNA expression plasmid with markedly silenced MMP-9 gene was screened out. BGC823 cell line with silenced MMP-9 gene was successfully constructed. Conclusion MMP-9 may play an important role in gastric cancer development, migration and invasion.

关 键 词：基质金属蛋白酶-9 RNA干扰 胃肿瘤 

分 类 号：R735.2[医药卫生—肿瘤]