盐酸千金藤碱抑制肝癌肾脏、睾丸转移及其分子机制

Cepharanthine hydrochloride inhibits kidney and testicular metastases from hepatocellular carcinoma in xenograft tumor models in nude mice

目的通过裸鼠肝原位移植荧光人肝癌细胞,观察肝癌细胞肾脏、睾丸转移情况,并探讨盐酸千金藤碱抑制肝癌细胞转移及其分子机制。方法采用慢病毒转染构建稳定表达荧光素酶基因的人肝癌细胞SMCC-7721-Luc。BALB/c裸鼠皮下接种SMCC-7721-Luc细胞,待肿瘤长至100 mm3左右,取皮下瘤植入裸鼠肝脏包膜,建立裸鼠原位肝癌模型。裸鼠腹腔注射盐酸千金藤碱,用小动物活体成像系统观察肝原位肿瘤肾脏、睾丸转移情况。给药21 d后,解剖取肿瘤组织,Western blot检测相关蛋白表达,同时取肝脏、肾脏、睾丸等组织,用10%中性甲醛固定,进行病理分析。结果构建的SMCC-7721-Luc细胞能稳定表达Luc基因,裸鼠肝原位接种该细胞后产生较强、稳定的荧光。活体成像系统显示盐酸千金藤碱能抑制裸鼠原位肝癌向肾脏、睾丸转移,与其降低MMP-7和N-cadherin蛋白表达,上调E-cadherin蛋白表达有关。结论盐酸千金藤碱通过调控MMP-7、N-cadherin和E-cadherin蛋白抑制原位肝癌向肾脏、睾丸转移。

Objective To observe the kidney and testicular metastases from hepatocellular carcinoma in nude mouse model by orthotopic transplantation of human liver cancer cells stably expressing reporter gene luciferasc, and investigate the anti-metastatic effects of cepharanthine hydrochloride （CEH） and its molecular mechanism. Methods SMMC-7721 cells were transfected with luciferase gene mediated by lentiviral vector to prepare SMMC-7721-Luc cells. Then the obtained cells were inoculated subcutaneously into the back of BALB/c nude mice to establish xenograft tumor model of nude mice. Till the masses grow into a size of 100 mm3 , the masses were collected and cut into blocks of 2 mm × 1 mm × 1 mm, and then transplanted into the liver to establish orthotopic tumor model of liver cancer. The tumor growth and kidney and testicular metastases were observed in nude mice after intraperitoneal injection of CEH by bioluminescence imaging system. In 21 d after CEH injection, the tumor tissues were harvested for expression of related proteins by Western blotting, and the liver, kidneys and testis were collected and fixed with 10% neutral formaldehyde for pathological observation. Results The established SMMC-7721-Luc cells stably expressed luciferase gene, and produced strong and stable fluorescence after orthotopic transplantation in nude mice. The bioluminescence imaging system displayed that CEH significantly inhibited the kidney and testicular metastases of orthotopic liver tumor, which was associated with the down-regulation of MMP-7 and N-cadherin, and up-regulation of E- cadherin. Conclusion CEH significantly inhibits the kidney and testicular metastases from hepatocellular carcinoma by regulating MMP-7, N-cadherin and E-cadherin.