【摘要】 目的研究FOXP3shRNA对肝癌细胞株SMMC-7721和MHCC-97H的增殖、凋亡和侵袭功能的影响。方法设计3种编码FOXP3shRNA的FOXP3干扰慢病毒:sh-FOXP3-1-pGreenPuro、sh-FOXP3-2-pGreenPuro和shFOXP3-3-pgreenpuro,并分别转染SMMC-7721和MHCC-97H2个肝癌细胞培养体系,检测转染后细胞培养体系中FOXP3的mRNA和蛋白质表达水平,以评估3个慢病毒的干扰效果。采用干扰效果最好的慢病毒转染上述细胞,以CCK8检测细胞增殖、TUNEL检测细胞凋亡、Transwell检测细胞侵袭功能,并比较转染前后上述指标差异。结果经菌落PCR和测序验证,3个FOXP3干扰慢病毒载体构建正确;其中sh-FOXP3-1干扰效果最明显,后期实验均使用sh-FOXP3-1转染。与对照组相比:2种肝癌细胞转染后OD值均明显下降,细胞增殖能力显著降低;凋亡显著升高;细胞侵袭功能显著下降,以上差异均有统计学意义(P值均<0.05)。结论实验条件下,干扰FOXP3的表达可抑制肝癌细胞增殖、促进肝癌细胞凋亡并降低肝癌细胞侵袭能力。未来应进一步研究其作用机理,以优化肝癌的临床治疗与预防策略。更多还原

【Abstract】 Objective To study the effects of FOXP3 shRNA on proliferation,apoptosis and invasion of liver carcinoma cells.Methods Three FOXP3 interfering lentivirus encoding FOXP3 shRNA were designed:sh-FOXP3-1-pGreenPuro,shFOXP3-2-pGreenPuro and sh-FOXP3-3-pGreenPuro,which were transfected to 2 liver carcinoma cell lines of SMMC-7721 and MHCC-97 H.The interference efficiency were evaluated for 3 interfering letivirus,the letivirus with the highest interference efficiency was selected to transfect liver carcinoma cell cultures for proliferation,apoptosis and invasion analysis.Cell counting kit CCK-8,terminal dexynucleotidyl transferase(TdT)-mediated dUTP nick end labeling and transwell assay were used to evaluate cell proliferation,apoptosis and invasion capacity,the results were compared before and after lentivirus transfection.Results Three FOXP3 interfering lentivirus vectors were constructed successfully and validated by colonial PCR and sequencing analysis.sh-FOXP3-1 was the most effective in reducing both mRNA and protein expression of FOXP3,which was selected for transfection studies.Compared with the normal control group,the OD values of the 2 carcinoma cells decreased significantly after transfection,the cell proliferation ability decreased significantly,the apoptosis increased significantly,and the cell invasion function decreased significantly(all P<0.05).Conclusion Under experimental conditions,FOXP3-shRNA transfection could inhibit cell proliferation,induce apoptosis and inhibit invasion of liver carcinoma cells.Further study should be conducted to understand its mechanism in order to optimize the clinical treatment and prevention strategy for liver cancer.更多还原