携多药耐药基因1反义RNA重组腺病毒逆转肝癌多药耐药细胞的实验研究

Reversal of adriamycin resistance of hepatocellular carcinoma by targeting it with recombined adenovirus carrying antisense multidrug resistance gene 1 RNA

目的探讨携带多药耐药基因1（multidrug resistance gene 1，MDRI）反义RNA重组腺病毒载体在裸鼠体内逆转肝癌多药耐药细胞HepG2／阿霉素（adriamycin，ADM）的疗效及作用机制。方法构建携带AFP和asmdrl的重组腺病毒载体Adeno-asmdrl，ADM分级诱导肝癌细胞HepG2为多药耐药细胞HepG2／ADM，建立HepG2／ADM裸鼠皮下移植瘤模型，Adeno-asmdrl局部注射，观察移植瘤的体积、透射电镜检测移植瘤组织细胞凋亡、RT-PCR检测MDRI转录水平，评价Adeno-asmdrl的抗肿瘤活性。结果在Adeno—asmdrl＋ADM组，移植瘤体积无增大，而PBS组、ADM组体积明显增大;RT-PCR检测移植瘤细胞1周和4周MDRImRNA水平，ADM组无明显变化，Adeno-amdrl＋ADM组在4周时MDRI转录水平仅为单纯ADM组的20％，经ADM＋Adeno-asmdrl处理组，可见凋亡增加，ADM组和PBS处理组的裸鼠移植瘤组织中出现少量或没有凋亡。结论携带MDRI反义RNA重组腺病毒部分逆转HepG2／ADM的多药耐药，阻止肿瘤生长，下调MDR1转录水平，导致肿瘤细胞凋亡。

Objective To investigate if an adenovirus vector carrying antisence multidrug resistance gene 1 （MDR1） could reverse multidrug resistance （MDR） of HepG2/adriamycin （ADM） cells in tumors transplanted in athymic mice. Methods An adenovirus vector carrying AFP promoter and antisence MDR1 was constructed. HepG2 MDR cells （HepG2/ADM） were induced by graded resistance to ADM and were subcutaneouly inoculated into athymic mice to construct the transplanted tumor. After adeno-asmdrl was injected, the volume of the transplanted tumor and the apoptotic body in the xenograft tumor cells were observed and reverse transcriptase polymerase chain reaction was employed to investigate the expression of the mdrl-mRNA from the mouse transplanted tumor cells. Results Following injection with adeno-asmdrl, the tumor volumes in this mice group did not increase. However the tumor volume in the PBS plus ADM group did increase significantly （P 〈 0.05）. In the tumor xenograft cells, mdrl mRNA in the xenografts was assessed by RT-PCR and found to be reduced at week 1, and at week 4 in the ADM＋asmdrl group, but it was stable in the ADM group. It was only 20% in the ADM＋asmdrl group compared to the ADM group at the 4th week （P 〈 0.05）. Evidence of apoptosis was observed in the tumor xenograft cells treated with adeno-asmdrl, but there was rarely any apoptosis in the group treated with ADM and PBS. Conclusion Adenovirus carrying antisense mdrl RNA can partially reverse the MDR of HepG2/ADM cells and inhibit tumor growth by down-regulating mdrl mRNA resulting in tumor cell apoptosis.