裸鼠人肝癌原位移植多药耐药模型的建立及其耐药机制的探讨

目的　在体内建立人肝癌裸小鼠原位移植多药耐药模型，并研究其耐药机理。方法采用人肝癌（BEL-7402）裸小鼠（4～6周龄）原位移植，给予阿霉素(1.5mg/kg体重，每周1次)腹腔注射诱导耐药（共8周），经四唑蓝(MTT)快速比色法检测原代培养的耐药细胞对抗癌药的敏感性，以流式细胞仪检测癌细胞表面mdr1基因产物P-糖蛋白（P-gp）的表达，并用罗丹明试验观察该蛋白功能。结果　移植瘤组织形态及生物学方面符合人肝癌特征，实验组肝癌细胞表面P-gp表达为(75.45±5.67)%，而对照组表达仅(4.25±1.28)%，差异有非常显著性（P＜0.01），对阿霉素的耐药倍数提高了16.67倍，对羟基喜树碱具有交叉耐受性（13.67倍）。耐药细胞表面P-gp有较强的药物外排功能。结论　阿霉素较易诱导原位移植于裸小鼠的人肝癌多药耐药性的产生，其耐药倍数与临床肝细胞癌相似。该模型的建立对研究肝癌多药耐药的产生机制以及逆转其多药耐药性具有重要价值。

Establishment of multidrug resistance in a model of orthotopic tr ansplantation of human hepatocellular carcinoma in nude mice and study on the me chanism of multidrug resistance

Objective　To establish a multidrug resist ance (MDR) model of orthotopic transplantation of human hepatocellular carcinoma (HCC) in nude mice (BALB/C nu/nu) and explore the mechanism of multidrug resist ance. Methods　Four to six week-old nude mice underwent o r thotopic transplantation of human HCC tissue (BEL-7402) and treated by injection of adriamycin (ADM 1.5 mg/kg, once a week for 8 weeks) to abdominal cavity to i nduce drug resistance. The primary culture of nude mice liver tumor was done 8 w eeks after HCC tissue transplantation. The cytotoxicity to cancer cells was dete rmined by MTT assay. The pump-efflux activity and the expression of mdr1 gene -coded p-glycoportein (P-gp) were examined by flow cytometric assay and by rhodamine123 (R123) test. Results　The typical behaviors of HCC were observed in the model. P-gp level in the ADM-treated group was (75.45±5.67)% , while (4.25±1.28)% in the control group (P＜0.01). Hepatocellular carcino ma cells with positive mdr1 expression showed cross drug-resistance to ADM, hyd roxycamptothecin by 16.67 and 13.67 times of inhibiting concentration(IC 50) as compared with negative mdr1 expression respecti vely. The drug-pump-efflux function of P-gp on the cytomembrane of hepatocell ular carcinoma cells with positive mdr1 gene expression was markedly enhanced. Conclusion　The multidrug resistance model of orthotopic t ransplatation of human HCC was easy to establishment and it could provide a reli a ble model for exploring the mechanism of multidrug resistance and reversing mult idrug resistance in HCC.