摘要
目的探讨Livin基因沉默对视网膜母细胞瘤细胞侵袭性的影响及其机制。方法体外培养视网膜母细胞瘤细胞Y79和视网膜色素上皮细胞ARPE-19,采用实时荧光定量PCR(RT-qPCR)和Western blot分别检测细胞中Livin的mRNA和蛋白表达水平。将Y79细胞分为对照组(细胞正常培养)、si-Livin组(转染Livin小干扰RNA)和si-NC组(转染乱序无意义阴性序列),CCK-8法检测各组细胞存活率,采用Transwell实验检测各组细胞迁移和侵袭能力,Western blot检测各组细胞中血管内皮生长因子(VEGF)、血管生成素-2(Ang-2)、基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶-9(MMP-9)的蛋白表达。结果与ARPE-19细胞中Livin的mRNA(1.01±0.06)和蛋白(0.18±0.07)表达比较,Y79细胞中Livin的mRNA(1.81±0.10)和蛋白(0.51±0.10)表达水平均显著升高(均为P<0.05)。与对照组细胞存活率(99.87±1.36)%、细胞迁移能力(126.89±8.54)个、细胞侵袭能力(100.89±7.92)个或si-NC组细胞存活率(98.43±1.22)%、细胞迁移能力(120.44±7.51)个、细胞侵袭能力(99.56±7.23)个比较,si-Livin组Y79细胞存活率(46.27±1.73)%、细胞迁移能力(58.00±5.68)个和细胞侵袭能力(37.56±4.30)个及VEGF、Ang-2、MMP-2和MMP-9的蛋白表达水平均显著降低(均为P<0.05)。结论Livin基因沉默可抑制视网膜母细胞瘤细胞的侵袭性,作用机制可能与下调血管形成相关因子VEGF、Ang-2、MMP-2和MMP-9的表达有关。
Objective To investigate the effect of silencing Livin gene on the invasiveness of retinoblastoma and its possible molecular mechanism.Methods The retinoblastoma cells Y79 and retinal pigmented epithelial cells ARPE-19 were cultured in vitro.The mRNA and protein expression levels of Livin in the cells were detected by real-time fluorescent quantitative PCR(RT-qPCR)and Western blot,respectively.Y79 cells were divided into control group(cells were cultured normally),si-Livin group(cells were transfected with Livin small interfering RNA),and si-NC group(cells were transfected with non-sequential negative sequence).Cell counting kit-8(CCK-8)was used to detect the cell survival rate of each group,Transwell was used to detect the cell migration and invasion ability of each group,and Western blot was used to detect the protein expression levels of vascular endothelial growth factor(VEGF),angiopoietin-2(Ang-2),matrix metalloproteinase 2(MMP-2)and matrix metalloproteinase 9(MMP-9).Results Compared with the expression of Livin’s mRNA(1.01±0.06)and protein(0.18±0.07)in ARPE-19 cells,the expression level of Livin’s mRNA(1.81±0.10)and protein(0.51±0.10)increased significantly(both P<0.05).Compared with the control group cell survival rate(99.87±1.36)%,cell migration ability(126.89±8.54)cells,cell invasion ability(100.89±7.92)cells or si-NC group cell survival rate(98.43±1.22)%,cell migration ability(120.44±7.51)cells,cell invasion ability(99.56±7.23)cells,the survival rate of Y79 cells(46.27±1.73)%,cell migration ability(58.00±5.68)cells and cell invasion ability(37.56±4.30)cells and the protein expression levels of VEGF,Ang-2,MMP-2 and MMP-9 were significantly reduced(all P<0.05).Conclusion Silencing Livin can inhibit the invasiveness of omentalblastoma cells,which may be related to down-regulating the expression of angiogenesis-related factors VEGF,Ang-2,MMP-2 and MMP-9.
作者
邓姿峰
罗瑜琳
汤文权
DENG Zifeng;LUO Yulin;TANG Wenquan(Department of Ophthalmology,Hunan Children’s Hospital,Changsha 410007,Hunan Province,China)
出处
《眼科新进展》
CAS
北大核心
2020年第12期1130-1134,共5页
Recent Advances in Ophthalmology
