鳖甲煎丸对HUVEC增殖及HepG2中VEGF表达的影响

目的:通过研究鳖甲煎丸对人脐静脉血管内皮细胞(human umbilical vascular endothelial cell,HUVEC)增殖及肝癌细胞系HepG2中血管内皮生长因子(vascular endothelial growth factor,VEGF)表达的影响,探讨其抗肝细胞癌血管生成的机制。方法:将24只6周龄Wistar大鼠随机分为3组(8只/组),分别以临床剂量20,10倍的鳖甲煎丸水溶液及生理盐水ig 3d,3 d后采血,离心,获取血清。用含有不同浓度HepG2培养上清的条件培养基培养HUVEC,48 h后用MTT比色法检测HUVEC的增殖情况并确定条件培养基的最适浓度。用该浓度的条件培养基培养HUVEC,分别于48,72 h后采用MTT比色法检测药物血清对HUVEC增殖的影响。用含药血清培养HepG2,48 h后采用ELISA方法检测培养液上清中VEGF的浓度,采用qRT-PCR法检测VEGF mRNA的表达情况。结果:鳖甲煎丸高、中剂量组10%药物血清能够显著抑制HUVEC的增殖,这种抑制作用具有浓度和时间依赖性。鳖甲煎丸高、中剂量10%药物血清组中VEGF浓度与VEGF mRNA表达水平均明显下降,并且下降程度与药物浓度有关。结论:鳖甲煎丸能够抑制HUVEC的快速增殖并且显著地降低HepG2中VEGF的表达水平。研究表明鳖甲煎丸在抗肝细胞癌血管生成方面具有一定作用。

Effect of Biejiajian Pills on HUVEC Proliferation and VEGF Expression in HepG2

Objective: To investigate the effect of Biejiajian pills (BP)on human umbilical vascular endothelial cell(HUVEC) proliferation and vascular endothelial growth factor(VEGF) expression in HepG2 and to explore the mechanism of BP to suppress angiogenesis of hepatocellular carcinoma(HCC, HepG2). Method: Twenty-four Wister rats were randomized equally into 3 groups for gavage of BP at 20-fold and 10-fold clinical doses and normal saline for 3 days. Blood samples were then collected from the rats, and the serum was separated and added in HepG2 cell cultures. HUVEC was cultured in conditioned medium (CM) supplemented with culture medium supernatant of HepG2 for 48 h. Then the proliferation of HUVEC was measured by MTT colorimetry. The effect of drug serum on HUVEC proliferation stimulated by CM was investigated by MTT colorimetry. After cultured in the medium containing drug serum for 48 h, the concentration of VEGF in HepG2 culture medium was assayed by ELISA and the expression of VEGF mRNA was investigated by qRT-PCR. Result: High and middle dose BP inhibited the proliferation of HUVEC in a concentration-and-time depending manner. High and middle dose BP decreased the secretion of VEGF as well as the expression of VEGF mRNA significantly, which is also correlated with the concentration of BP. Conclusion: BP can effectively inhibit the proliferation of HUVEC and lower the expression of VEGF. It may serve as a potential anti-angiogenesis agent in HCC.