肝动脉结扎对转移性人肝癌裸鼠原位移植瘤乏氧的影响

目的 观察肝动脉结扎(HAL)对转移性人肝癌裸鼠原位移植瘤乏氧的影响.方法 采用24只BALB/C-nu/nu裸鼠,建立转移性人肝癌裸鼠原位移植模型;随机分为2组,A组于移植瘤术后2周进行HAL(n=12),B组假手术(Sham)作为对照(n=12).分别于干预术后2 d和4周各随机处死每组中6只荷瘤裸鼠,利用免疫组织化学显色(SP法)和Western blot检测移植瘤内哌莫硝唑(Pimonidazole)和缺氧诱导因子(HIF)-1α的阳性表达和染色强度,以及血管内皮生长因子(VEGF)蛋白水平和肿瘤组织微血管密度(MVD),并用肺连续切片法计数4周时荷瘤裸鼠肺转移灶.结果 与Sham组比较,HAL组移植瘤内Pimonidazole阳性细胞及HIF-1α表达水平在术后2 d显著增加(P<0.05),并且肿瘤组织和血清内VEGF水平[(922.5±59.3)比(349.6±46.5)ng/L,P<0.01]明显增加.术后4周,荷瘤裸鼠肺转移率较对照组增加(6/6比1/6,P<0.05),但此时乏氧细胞,VEGF表达和MVD则与对照组比较差异无统计学意义(P>0.05).结论 HAL促进转移性人肝癌裸鼠原位移植瘤短期乏氧及VEGF表达,但不影响移植瘤长期乏氧效应.

The hypoxic effects of hepatic artery Hgation on metastatic human orthotopic hepatoma in nude mice model

Objective To investigate the hypoxic effects of hepatic artery ligation on metastatic human orthotopic nude mouse model of hepatocellular carcinoma (HCC) and further analyze the changes of angiogenesis in tumor tissues.Methods Human HCC cell line MHCC97L was inoculated into 24 BALB/C nude mice.All these cases were divided equally into HAL and sham-operation subgroups at the 14th day after orthotopie implantation.Six randomly selected nude mice of each subgroup were sacrificed at 2nd and/or 24th day after the second operation respectively, and immunostaining and Western blot were carried out to detect the expression of Pimonidazole and hypoxia inducible factor (HIF)-1α in tumor tissues.The vascular endothelial growth factor (VEGF) level in tumor tissues and serum was determined by Western blot and enzyme-linked immunosorbent assay (ELISA) ,respectively.The mean micrevessel density (MVD) of intratumoral tissues was analyzed by immunostaining.At the 6th week after orthotopic implantation, the pulmonary metastasis in nude mice bearing HCC was evaluated.Results HAL significantly increased the intensity of Pimonidazole (9.33 ± 2.34 vs 5.00 ± 2.10, P < 0.05) and the expression level of HIF-1α (+++- ++++ vs + - ++ ,P < 0.05) in xenografts at the 2nd day after occlusion of hepatic arterial blood, as compared with sham operation group, but could not augment long-term intratumoral hypoxia.In line with hypoxia, VEGF level in tumor tissues and serum [(922.5 ± 59.3) vs (349.6 ± 46.5) ng/L, P <0.01] was significantly higher at the 2nd day after HAL than the controls,and no significant difference was found after 24 days.There was no significant difference in intratumoral MVD between HAL group and sham-operation group (P < 0.05).Conclusion HAL could induce dramatic short-term hypoxia in xenograft HCC and overexpression of VEGF, but had no effects on hypoxia and angiogenesis of tumor for a long time.