AKT2基因沉默抑制卵巢癌细胞侵袭和粘附能力的实验研究

Experimental study on silencing of AKT2 gene by RNAi to inhibit the invasion and adhesion of human ovarian cancer cell line.

目的:探讨AKT2基因对人卵巢癌细胞A2780侵袭和黏附活性的影响。方法:构建AKT2基因的短发卡状RNA质粒转染人卵巢癌细胞A2780,运用RT-PCR、蛋白质免疫印迹法比较转染前后AKT2表达的差异;用transwell小室检测细胞侵袭人工基底膜的能力;MTT法检测卵巢癌细胞与细胞外基质黏附能力。结果:与对照相比,构建shR-NA表达载体可抑制AKT2 mRNA转录和蛋白的表达;AKT2表达下降后,穿透重组基底膜的细胞数明显下降(P<0.05),且可显著抑制细胞与细胞外基质的黏附(P<0.05)。结论:靶向AKT2 shRNA能有效地抑制卵巢癌细胞中AKT2基因的表达,并抑制卵巢癌细胞体外侵袭和黏附能力。AKT2基因有可能成为治疗卵巢癌的新途径。

Objective：To investigate the effect of AKT2 gene expression on the invasion and adhesion of human ovarian cancer cell line 322780 by RNAi. Methods：Vectors containing shRNA targeting AKT2 gene were constructed and transfected into human ovarian carcinoma cell line 322780 ,The RT-PCR and western blot were used to detect the efficiency of gene silencing. The invasion of cells was measured by transwell chambers attached with polycarbonate filters and reconstituted basement membrane （Matrigel）, the adhesion of cells to artifical basement membrane Matrigel was measured by methyl thiazolyl tetrazolium（MTT） ; Empty plasmid- transfected A2780 and normal A2780 were used as control. Results：Western blot and RT-PCR indicated that the expression of AKT2 was suppressed by RNAi ; Compared with control groups, the cell numbers penetrated polycarbonate membrane were decreased and the adhesive inhibi- tion rate was obviously increased （ P 〈 0.05 ）. Conclusion： Transfection of shRNA targeting at AKT2 gene can efficiently inhibit the AKT2 expression in A2780 cells,which can suppress the abilities of adhesion and invasion of ovarian cancer cell. So AKT2 gene may provide a new target and an effective way to treat cvarian cancer.