腺病毒介导反义基质金属蛋白酶-2抑制肝癌生长和血管生成的研究

Inhibitory effect of antisense matrix metalloproteinase-2 mediated by adenovirus on hepatocellular carcinoma growth and angiogenesis in vivo

目的探讨携带反义二型基质金属蛋白酶(MMP2)基因的重组腺病毒(Ad-MMP2AS)对人肝癌动物模型生长和血管生成的抑制作用。方法用我们构建的Ad-MMP2AS感染人肝癌细胞株(Bel-7402)。Boyden Chamber检测Ad-MMP2AS对Bel-7402细胞降解人工基底膜(Matrigel)的抑制作用;Western blotting和明胶酶谱分析测定Ad-MMP2AS对Bel-7402细胞分泌MMP2的影响;用感染Ad-MMP2AS的Bel-7402细胞接种于裸鼠皮下观察其成瘤能力;Ad-MMP2AS瘤内注射,观察它对肝癌生长的抑制作用;肿瘤组织切片、HE染色观察瘤细胞生长情况;免疫组织化学分析肿瘤组织血管密度。结果Ad-MMP2AS感染的Bel-7402细胞穿过Matrigel的Bel-7402细胞数下降52%、成瘤量下降4.3倍;瘤内注射Ad-MMP2AS使瘤体生长减少63%;肿瘤组织血管密度减少2.6倍。结论Ad-MMP2AS能抑制肝癌的生长和肿瘤血管的生成,对肝癌有治疗潜力。

Objective To investigate the inhibitory effect of a recombinant adenoviral vector carrying antisense matrix metalloproteinase-2 （MMP2） on the growth and angiogenesis of human hepatocellular carcinoma （HCC） animal model in vivo. Methods The recombinant adenoviral vector carrying antisense MMP2 （Ad-MMP2AS） which was constructed previously was used to infect the human HCC cell line （Bel-7402）. The invasiveness of the Bel-7402 cells was assayed in Matrigel, and the production of MMP2 in the Bel-7402 cells was detected by using Western blotting analysis and gelatin zymography. Following the Ad-MMP2AS-infeeted cells were subcutaneously inoculated in nude mice, and injected intratumoraUy into pre-existing tumors, the tumors were removed, sectioned, stained with H＆E and stained with an anti-CD31 monoclonal antibody. Results Compared with PBS or Ad-CMV-infeeted cells, infection of Bel-7402 cells with Ad-MMP2AS significantly reduced MMP2 enzyme activity, the number of Bel-7402 cells penetrating Matrigel was decreased by 52%, and the tumor volume was reduced by 4.3-fold. Direct intratumoral injection of Ad-MMP2AS into pre-existing tumors could decrease the tumor growth by 63 % and the tumor vessel density was reduced by 2.6-fold. Conelusion The recombinant adenovirus carrying antisense MMP2 can effectively inhibit the invasiveness and growth of Bel-7402 cells in vitro and in vivo.