缺氧诱导因子-1α反义寡核苷酸抑制HBx诱导的血管内皮生长因子的表达

HIF-1α Anti-sense Oligonucleotide Inhibiting the Expression of VEGF Induced by HBx

目的 探讨缺氧诱导因子-1α(HIF-1α)反义寡核苷酸对乙型肝炎病毒x蛋白(HBx)诱导的人肝癌组织中血管内皮生长因子(VEGF)表达的抑制作用。方法 用编码HBx基因全长的pHA-HBx质粒转染肝癌HepG2细胞;Western Blot方法鉴定HBx基因的整合和表达;用转染后的细胞建立表达HBx的裸鼠人肝癌模型。实验组用HIF-1α反义寡核苷酸作肿瘤组织内注射,对照组以相同体积的生理盐水作瘤体注射,观测不同时间点裸鼠肿瘤的生长状况。采用免疫组织化学染色方法检测肿瘤组织中HIF-1α、VEGF表达水平和微血管密度。结果 HBx基因成功整合人肝癌HepG2细胞基因组并表达;实验组经反义寡核苷酸处理后肿瘤体积和重量均低于对照组,两组间差异有显著性意义(P<0.05);实验组肿瘤组织中HIF-1α表达下调;实验组肿瘤组织中VEGF表达为(21.6±6.7)％,低于对照组的(78.6±10.2)％(P<0.01),微血管密度为19.75±7.38,低于对照组的36.64±12.25(P<0.01)。结论 HIF-1α反义寡核苷酸肿瘤组织注射可以抑制肝癌组织中HBx诱导的VEGF表达,抑制表达HBx的肿瘤生长。

Objective To investigate the inhibition of hypoxia inducible factor-la (HIF-1α) antisense oligodeoxynucleotide to the expression of vascular endothelial growth factor (VEGF) induced by HBx in hepatic carcinoma tissues of nude mice. Methods The plasmid pHA-HBx encoding full length of HBx was transfected into human hepatoma HepG2 cells. The integration and expression of HBx gene in genome DNA of transfected cells were identified by Western-blot. The nude mice models of human hepatoma expressing HBx were established by injecting HBx-trandfected HepG2 cells into the flank of nude mice sub-cutaneously. The antisense oligodeoxynucleotide of HIF-1α and the saline were injected into the nude mice tumor tissues in both experimental group and control group respectively. The volume of xenograft tumors at different time points was observed. The expression of HIF-1α and VEGF in tumor tissues was detected immunohistochemically and the MVD was calculated. Results The HBx gene was integrated into the genome DNA of HepG2 cells and expressed. The antisense oligodeoxynucleotide-treated tumors were significantly smaller in volume and lower in weight than those in control group (P<0. 05) ; The expression of HIF-la was down-regulated in experimental tumor tissues; The expression of VEGF in experimental tumor cells was much decreased as compared with that in control group (P<0. 05) and the amount of MVD in experimental tumor tissues was significantly lower than that in control group. Conclusion By being injected into the tumor tissues, the HIF-1α antisense oligodeoxynucleotide could inhibit the expression of VEGF induced by HBx in human hepatoma cells of nude mice, which can inhibit the growth of tumors expressing HBx.