Mfn-2沉默对肝癌细胞侵袭及PITPNM3表达的影响

Influence of Mfn-2 silence on hepatoma cell invasion and PITPNM3 expression

目的探究Mfn-2沉默对肝癌细胞侵袭的影响及其潜在的分子机制。方法用Mfn-2-siRNA和siRNA转染SMMC-7721细胞,为Mfn-2-siRNA组和siRNA组,以未经转染的细胞作为对照组。转染48 h后,采用荧光倒置显微镜检测转染效率,实时荧光定量PCR(qRT-PCR)检测SMMC-7721细胞中Mfn-2的表达情况,Transwell小室检测SMMC-7721细胞侵袭能力,免疫组化和Western blot检测SMMC-7721细胞中基质金属蛋白酶-2(MMP-2)和磷脂酰肌醇转移蛋白3(PITPNM3)蛋白表达情况。结果荧光倒置显微镜检测结果显示,转染效率约85%;siRNA组SMMC-7721细胞中Mfn-2 mRNA的表达水平与对照组相比无显著差异(P>0.05),Mfn-2-siRNA组SMMC-7721细胞中Mfn-2 mRNA的表达水平显著低于对照组(P<0.05);siRNA组SMMC-7721细胞侵袭能力、以及MMP-2和PITPNM3的表达水平与对照组相比无显著差异(P>0.05),Mfn-2-siRNA组SMMC-7721细胞侵袭能力、以及MMP-2和PITPNM3的表达水平显著高于对照组(P<0.05)。结论 Mfn-2沉默能够增强肝癌细胞SMMC-7721的侵袭能力,推测这一作用是通过上调MMP-2和PITPNM3的表达水平实现的,上述结果为肝癌的治疗和靶向药物的开发提供了一定理论基础。

Objective To explore the influence of Mfn-2 silence on hepatoma cell invasion and its potential molecular mechanism.Methods SMMC-7721 cells were transfected by using Mfn-2-siRNA and siRNA,and named for Mfn-2-siRNA and siRNA group,and those not transfected were as the control group.After transfection of 48 h,transfection efficiency was tested by using fluorescence inverted microscope,and Mfn-2 expression in SMMC-7721 cells was tested by real time quantitative PCR(qRT-PCR).SMMC-7721 cell invasion was detected by boyden chamber transwell.MMP-2 and PITPNM3 protein expression in SMMC-7721 cells was tested by Immunohistochemistry and Western blot.Results The result of fluorescence inverted microscope showed that transfection efficiency was about 85%.There was no significant difference in SMMC-7721 cell Mfn-2 mRNA expression level between siRNA group and control group(P>0.05).SMMC-7721 cell Mfn-2 and mRNA expression level in the siRNA group were significantly lower than those of the control group(P<0.05).There was no significant difference in SMMC-7721 cell invasive ability,and MMP-2 and PITPNM3 expression levels between the siRNA group and the control group(P>0.05).SMMC-7721 cell invasion ability,MMP-2 and PITPNM3 expression levels in the Mfn-2-siRNA group were significantly higher than those in the control group(P<0.05).Conclusion Mfn-2 silence can enhance the invasive ability of hepatoma cell SMMC-7721.It is speculated that the effect is achieved by up-regulating MMP-2 and PITPNM3 expression levels,which provides a theoretical basis for the treatment of hepatoma and the development of targeted drugs.