虫草素抑制肝癌转移及分子机制研究

Antimetastatic effects of cordycepin and its molecular mechanism in human hepatoma MHCC97H cells in vitro and in vivo

探讨虫草素抑制肝癌细胞MHCC97H转移及其分子机制。采用MTT法检测细胞增殖;应用细胞划痕实验检测细胞迁移能力;Transwell检测细胞侵袭及迁移能力;Western blotting检测蛋白的表达水平;应用皮下移植及尾静脉注射法检测肿瘤原位生长及肺转移。实验结果表明虫草素剂量依赖性地抑制MHCC97H细胞的生长、迁移及侵袭能力,这与其降低AKT、p-AKT、p-GSK-3β、β-catenin、N-cadherin、MMP-7和MMP-9蛋白表达,上调E-cadherin的蛋白表达等有关。动物实验表明虫草素剂量依赖性地抑制MHCC97H细胞的原位生长及肺转移,虫草素高剂量组(40 mg·kg^(-1))、中剂量组(20 mg·kg^(-1))、低剂量组(10 mg·kg^(-1))及5-氟尿嘧啶组的原位移植瘤重分别为0.38±0.04、0.61±0.08、1.13±0.36和0.65±0.07 g,与对照组瘤重(1.52±0.46 g)相比,虫草素高、中剂量组及5-氟尿嘧啶组具有显著性差异(P<0.01),但虫草素低剂量组无统计学差异(P>0.05);在肺转移模型中,以上各组的肺转移结节数分别为48.9±7.2、67.2±9.4、106.4±11.3和73.6±8.6,与对照组肺转移结节数(123.5±14.5)相比,虫草素高、中剂量组及5-氟尿嘧啶组具有显著性差异(P<0.01),虫草素低剂量组无统计学差异(P>0.05)且肺转移结节中相关蛋白表达的变化与细胞实验一致。故虫草素通过调控AKT信号通路抑制肝癌细胞MHCC97H生长及转移。

The objective of this study was to examine the antimetastatic effects of cordycepin and elucidate its molecular mechanism using MHCC97H cells in vitro and in vivo. Cellular proliferation was detected with MTT assay. The migration and metastatic potential were measured with scratch wound healing as well as transwell migration assays in vitro. Protein expression was detected by Western blotting. Antitumor and antimetastatic effects of cordycepin were evaluated by subcutaneous xenograft and lung metastatic model in vivo. The results demonstrated that cordycepin significantly inhibited MHCC97H cells proliferation and metastasis which was due to the down-regulation of AKT, p-AKT, p-GSK-3β, β-catenin, N-cadherin, MMP-7, MMP-9 and up-regulation the expression of E-cadherin. Furthermore, cordycepin inhibited tumor growth and metastasis in a dose-dependent manner in vivo. Cordycepin（40 and 20 mg·kg^（-1）） and 5-fluorouracil group significantly inhibited the tumor weights to 0.38 ± 0.04, 0.61 ± 0.08 and 0.65 ± 0.07 g, respectively, comparing with the control group（1.52 ± 0.46 g）（P 0.01）, but not 10 mg·kg^（-1） cordycepin group（1.13 ± 0.36 g）（P 0.05）; the lung metastasis nodus numbers showed the same results, which in all group above（48.9 ± 7.2, 67.2 ± 9.4, 73.6 ± 8.6, respectively） were fewer than the control group（123.5 ± 14.5）（P 0.01）, except 10 mg·kg^（-1）cordycepin（106.4 ± 11.3）（P 0.05）. Collectively, cordycepin inhibited MHCC97H cell proliferation and metastasis in vivo and in vitro.