中药白芨提取物抑制肿瘤血管生成机制的实验研究

目的　探讨中药白芨提取物抑制肿瘤血管生成的机制。方法　应用细胞培养方法 ,比较空白组、对照组及不同浓度白芨胶组之间人肝癌细胞系 (Hep G2 )细胞增殖率、凋亡率、血管内皮生长因子 (VEGF)分泌水平的差异 ,并观察细胞培养上清对人脐静脉内皮细胞系 (ECV 30 4)生长的影响。对Walker 2 5 6移植性肝癌大鼠行经动脉化疗栓塞治疗 ,分为对照组、单纯化疗组、碘油栓塞组及白芨微球栓塞组 ,每组 2 0只大鼠。栓塞术后 2周 ,用免疫组织化学法检测肿瘤细胞Ⅷ因子 用以计算微血管密度 (MVD) ]、VEGF及碱性成纤维细胞生长因子 (b FGF)表达情况。结果　细胞培养各组之间Hep G2细胞增殖率、凋亡率及上清液中VEGF浓度的差异无显著意义 ,但经白芨胶处理后的Hep G2细胞上清液可明显抑制ECV 30 4内皮细胞的增殖 ,当白芨胶浓度为 0 5、1 0、2 0、4 0、8 0 μg/ml时 ,内皮细胞生长抑制率分别为 5 7 6 %、6 6 7%、86 4%、87 5 %、94 8% ,呈剂量依赖性。动脉化疗栓塞实验中 ,白芨微球栓塞组肿瘤MVD(血管计数 /视野 )为 5 9± 34,明显低于对照组 ( 81± 2 4)、单纯化疗组 ( 83±2 0 )及碘油栓塞组 ( 85± 2 4) (F =5 177,P =0 0 0 3) ;VEGF、b FGF的表达各组间差异无显著意义。结论白芨提取物可能通过抑制肿瘤血管内

Mechanism of inhibition of tumor angiogenesis by Bletilla colloid: an experimental study

OBJECTIVE: To study the mechanism of inhibition of tumor angiogenesis by Bletilla colloid. METHODS: Human Hep-G2 hepatocellular carcinoma cells were cultured and treated with Bletilla colloid of different concentrations. Pure culture of Hep-G2 cells was used as control and pure culture medium without Hep-G2 cell was used as blank control. The concentration of vascular endothelial growth factor (VEGF) in the supernatant was detected by ELISA. The apoptosis and proliferation of the Hep-G2 cells were examined by flow cytometry. Cells of human endothelial cell line ECV-304 were cultured in the supernatant of culture media of Hep-G2 treated with Bletilla colloid of different concentrations. MTT method was used to observe the growth of the ECV-304 endothelial cells. Eighty rats were made animal model of transplanted Walker-256 hepatoma and then randomly divided into 4 groups of 20 rats 12 - 14 days after to undergo transarterial chemoembolization (TACE) treated with normal saline, 5-fluouracil (5-Fu), iodized poppy seed oil, and 5-Fu-Blettila microsphere respectively. Two weeks after, the rats were killed and the tumors were taken out. Immunohistological staining was conducted to detect the expression and localization of factor VIII, VEGF, and basic fibroblast growth factor (b-FGF). The microvcascular density (MVD) was calculated by counting the factor VIII positive endothelial cells. RESULTS: There was no statistically significant difference in the apoptosis rate, proliferation rate and supernatant VEGF level between the control group and the Bletilla groups. The inhibition rates of ECV-304 endothelial cell growth were 57.6%, 66.7%, 86.4%, 87.5%, and 94.8% in the groups of Bletilla of the concentrations of 0.5, 1.0, 2.0, 4.0, and 8.0 micro g/ml respectively in a dose-dependent manner. The MVD of tumor was 59 +/- 34 per field in the 5-Fu-Blatilla group,significantly lower than those in the other 3 groups (F = 5.177, P = 0.003). The expression of VEGF and the expression of b-FGF were not significantly different among the 4 TACE groups. CONCLUSION: Bletilla colloid inhibits angiogenesis after TACE, potentially, through inhibition of the binding of vascular endothelial growth factor to its receptor.