血管内皮生长因子-C在胃癌中的表达及其反义基因转染对人胃癌细胞SGC-7901的影响

目的:探讨胃癌组织中血管内皮生长因子-C(VEGF-C)的表达与肿瘤新生淋巴管形成的关系。观察以pCI-neo为载体的反义血管内皮生长因子-C(Anti-VEGF-C)转染的人胃癌细胞SGC-7901中VEGF-C蛋白的表达,及其对人胃癌细胞生长的抑制作用。方法:取72例胃癌标本的癌旁组织,通过RT-PCR检测VEGF-C的mRNA表达;免疫组化检测VEGFR-3,计算出微淋巴管的密度并行相关统计分析。以脂质体转染法转染既往试验合成的重组质粒pCI-neo-anti VEGF-C到体外培养的人胃癌细胞株,以未转染组为对照,用免疫组化和Western blot分析VEGF-C基因及蛋白的表达,最后用MTT法及流式细胞仪分析其对细胞生长的抑制作用。结果:淋巴结转移阳性组中VEGF-C mRNA阳性表达85.7%,高于阴性组的20.0%(P<0.05);VEGF-C mRNA表达阳性组淋巴管密度6.36±1.66,阴性组3.95±1.52(P<0.05);淋巴结转移阳性组淋巴管密度6.65±1.57,阴性组3.75±1.47(P<0.05)。重组质粒pCI-neo-anti VEGF-C转染体外培养的人胃癌细胞株后,免疫组化和Western blot均显示pCI-neo-anti VEGF-C转染组无VEGF-C mRNA及其蛋白的表达,MTT检测表明pCI-neo-anti VEGF-C转染组活细胞数较其它各组均低(P<0.05)。结论:VEGF-C能促进癌旁微淋巴管增生、提高胃癌侵袭力;反义VEGF-C转染可封闭人胃癌细胞SGC-7901VEGF-C mRNA,减少其VEGF-C蛋白的表达,抑制其体外生长。

Expression of Vascular Endothelial Growth Factor-C in Gastric Carcinoma and the Effect of Its Anti-sense Gene Transfection on Human Gastric Carcinoma Cell Line SGC-7901

Objective: To discuss the expression of vascular endothelial growth-C(VEGF-C) in gastric carcinomas tissue and its relation with tumor lymphangiogenesis;to observe the expression of VEGF-C protein on human gastric cancer cell line SGC-7901 which had been transfected by anti-sense VEGF-C based on the carrier of pCI-neo,and to investigate whether it could inhibit the growth of human gastric carcinoma cell line in vitro.Methods: The expressions of VEGF-C mRNA in 72 gastric carcinoma tissue specimens were determined by RT-PCR.Immunohistochemistry was used to detect the expression of VEGFR-3 in these tissues as well.Lymphatic vessel density(LVD) was counted.Using the liposome infection protocol which had been reported previously,the plasmid of pCI-neo anti-sense VEGF-C was transfected into human gastric cancer cell line SGC-7901,then immunohistochemical and Western blotting were used to detect target genes and its expression.The cell viability was detected by MTT assay.Results: The positive expression rate of VEGF-C in positive lymph node was significantly higher than that in negative lymph node(85.7% vs 20.0%,P<0.05).Compared with that in lymph node without metastasis,LVD in positive lymph node increased significantly(6.65±1.57 vs 3.75±1.47,P<0.05).LVD in positive VEGF-C tissue was significantly higher than that in negative VEGF-C tissue(6.36±1.66 vs.3.95±1.52,P<0.05).The immunohistochemical & Western blotting results showed that there was no expression of VEGF-C mRNA or protein in the transfected group.MTT assay revealed that the transfected group had a lower cell viability than the other groups(P<0.05).Conclusion: VEGF-C can promote the invasion of gastric carcinoma and lymphatic proliferation.The expression of VEGF-C mRNA and its protein in SGC-7901 cells could be inhibited by pCI-neo-antisense VEGF-C through transfection.The study showed that anti-sense VEGF-C gene could suppress the growth of human gastric cells in vitro as well.