CAJ | 학술논문

目的探讨受体相互作用蛋白(RIP)140基因敲低对小鼠小胶质瘤细胞(BV-2)增殖的影响.方法用脂质体转染方法将RIP140-短发夹RNA(shRNA)质粒导入BV-2细胞,用嘌呤霉素筛选稳定表达RIP140-shRNA的细胞系,用实时定量PCR和Western印迹法检测RIP140敲低的效果,用四甲基偶氮唑蓝(MTT)法检测RIP140敲低对细胞增殖的影响.结果成功构建稳定表达RIP140-shRNA的BV-2-71674和BV-2-71080;与BV-2细胞相比,BV-2-71674和BV-2-71080细胞中RIP140的表达无论mRNA水平还是蛋白质水平均明显低,下调率(mRNA)分别为73%和75%(均P<0.01);MTT法检测显示,BV-2细胞在24、48、72、96 h 4个时间点增殖率分别为3.03±0.01、7.36±0.08、14.15±0.25、23.35±0.09,BV-2-71674细胞则分别为2.15±0.03、6.43±0.08、11.77±0.31、18.47±0.04,BV-2-71080细胞分别为1.77±0.03、4.74±0.25、10.03±0.02、17.66±0.21;BV-2-71674和BV-2-71080细胞增殖率均明显低于BV-2细胞,差异均有统计学意义(均P<0.01).结论 RIP140敲低可以抑制BV-2细胞的增殖.
목적 탐토수체상호작용단백(RIP)140기인고저대소서소효질류세포(BV-2)증식적영향.방법 용지질체전염방법장RIP140-단발협RNA(shRNA)질립도입BV-2세포,용표령매소사선은정표체RIP140-shRNA적세포계,용실시정량PCR화Western인적법검측RIP140고저적효과,용사갑기우담서람(MTT)법검측RIP140고저대세포증식적영향.결과 성공구건은정표체RIP140-shRNA적BV-2-71674화BV-2-71080;여BV-2세포상비,BV-2-71674화BV-2-71080세포중RIP140적표체무론mRNA수평환시단백질수평균명현저,하조솔(mRNA)분별위73%화75%(균P<0.01);MTT법검측현시,BV-2세포재24、48、72、96 h 4개시간점증식솔분별위3.03±0.01、7.36±0.08、14.15±0.25、23.35±0.09,BV-2-71674세포칙분별위2.15±0.03、6.43±0.08、11.77±0.31、18.47±0.04,BV-2-71080세포분별위1.77±0.03、4.74±0.25、10.03±0.02、17.66±0.21;BV-2-71674화BV-2-71080세포증식솔균명현저우BV-2세포,차이균유통계학의의(균P<0.01).결론 RIP140고저가이억제BV-2세포적증식.
Objective To investigate the effects of receptor-interacting protein (RIP) 140 gene knockdown on the proliferation of microglioma cells. Methods Mouse microglioma cells of the line BV-2 were cultured and transfected with 2 kinds of recombinant RIP140-shRNA plasmids (V2MM-71674 and V2MM-71080) or blank plasmid MSCV-EGFP. Real-time PCR and Western blotting were used to detect the mRNA and protein expression of RIP140; and the cell proliferation was detected by MTT assay. Results There were not significant differences in the RIP140 mRNA and protein expression between the BV-2 and BV-2-MGCV-EGFP groups. Compared to those of the BV-2 group, the RIP140 mRNA expression levels of the BV-2-71674 and BV-2-71080 groups were lower by 73% and 75% respectively. The protein expression levels of the BV-2-71674 and BV-2-71080 groups were remarkably lower than those of the BV-2 and BV-MSGV-EGFP groups. MTT assay showed that there were not significant differences in the proliferation rates at different time points between the BV-2 and BV-2-MSCV-EGFP groups, however, the proliferation rates at the time points of 24, 48, 72, and 96 h of the BV-2-71674 and BV-2-71080 groups were significandy lower than those of the BV-2 group (all P < 0. 01). Conclusion RIP140 gene knockdown effectively inhibits the proliferation of microglioma cells.