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目的研究Angiostatin基因联合反义缺氧诱导因子-1α(HIF-1α)基因治疗人肝癌裸鼠皮下移植瘤的协同抑制作用。方法用人肝癌细胞株SMMC-7721建立人肝癌裸鼠皮下移植瘤动物模型。四组荷瘤裸鼠分别注射质粒PcDNA3、Angiostatin/PcDNA3、HIF-1α/PcDNA3B、Angiostatin/ PcDNA3+HIF-1α/PcDNA3B。观察肿瘤生长曲线,检测肿瘤的Angiostatin、血管内皮生长因子(VEGF)、HIF-1α表达和微血管密度(MVD),利用TUNEL染色法行原位细胞凋亡分析。结果Angiostatin基因治疗在早期具有抑制肿瘤生长的作用;Angiostatin基因治疗组的MVD(24.8±2.8)低于空质粒对照组(30.2±4.1),VEGF表达高于空质粒对照组,细胞凋亡指数(2.87±0.48)高于空质粒对照组(1.55±0.43);联合治疗组有明显抑制肿瘤生长的作用,HIF-1α局部低表达,MVD (14.6±2.1)低于Angiostatin基因治疗组(24.8±2.8),VEGF表达低于单独治疗组,细胞凋亡指数(5.12±0.63)高于单独治疗组。结论肿瘤对Angiostatin基因治疗可以产生耐受性;反义HIF-1α基因阻断肿瘤的缺氧适应途径在一定程度上解决了抗血管生成治疗的耐药性问题。
Objective To investigate the synergistic inhibitory effect of angiostatin gene combined with antisense hypoxia inducible factor-1α (HIF-1α) gene on human hepatocellular carcinoma xenografts in nude mice. Methods Human hepatocellular carcinoma xenografts in nude mice were established by human hepatocellular carcinoma cell line SMMC-7721. Four groups of tumor-bearing nude mice were injected with pcDNA3, Angiostatin / PcDNA3, HIF-1α / PcDNA3B, Angiostatin / PcDNA3 + HIF-1α / PcDNA3B respectively. Tumor growth curves were observed. Angiostatin, VEGF, HIF-1α expression and microvessel density (MVD) were detected. TUNEL staining was used to detect apoptosis in situ. Results Angiostatin gene therapy could inhibit the growth of tumor cells in early stage. The MVD (24.8 ± 2.8) in Angiostatin gene therapy group was lower than that in control group (30.2 ± 4.1), the expression of VEGF was higher than that in empty plasmid control group 2.87 ± 0.48) was higher than that of empty plasmid control group (1.55 ± 0.43). The combined treatment group had obvious inhibition on tumor growth, HIF-1α was locally low expression, MVD was lower than that of Angiostatin gene therapy group (24.8 ± 2.8) ), VEGF expression was lower than that of the untreated group. The apoptotic index (5.12 ± 0.63) was higher than that of the untreated group. Conclusions The tumor is resistant to Angiostatin gene therapy. Antisense HIF-1α gene blocks the hypoxic adaptation of tumor to a certain extent, which solves the problem of resistance to anti-angiogenesis therapy.