CAJ | 학술논문

目的利用小干扰RNA (siRNA)技术沉默Nestin基因,观察其逆转胶质瘤细胞恶性表型的效果.方法设计靶向Nestin基因的siRNA片段,利用脂质体转染人U251胶质瘤细胞株,逆转录-聚合酶链反应(RT-PCR)和免疫组织化学法检测Nestin基因和蛋白的表达,噻唑蓝(MTT)比色法检测细胞增殖,Transwell实验检测细胞的迁移能力,软琼脂实验检测细胞的恶性增殖能力.结果转染siRNA基因片段的U251胶质瘤细胞株基因表达水平(U251细胞为0.83±0.16、U251/vect细胞为0.81 ±0.23、U251/S3为0.20±0.11)和蛋白表达水平(U251细胞为157.73±4.12、U251/vect细胞为153.34±5.27、U251/S3为112.20±3.16)显著下降(P＜0.05),MTT实验显示siRNA干扰后U251胶质瘤细胞株体外增殖速度减慢、细胞侵袭能力及软琼脂集落形成能力均显著降低(P＜0.01).结论 Nestin基因表达下调可抑制胶质瘤细胞株U251的侵袭及增殖能力.
목적 이용소간우RNA (siRNA)기술침묵Nestin기인,관찰기역전효질류세포악성표형적효과.방법 설계파향Nestin기인적siRNA편단,이용지질체전염인U251효질류세포주,역전록-취합매련반응(RT-PCR)화면역조직화학법검측Nestin기인화단백적표체,새서람(MTT)비색법검측세포증식,Transwell실험검측세포적천이능력,연경지실험검측세포적악성증식능력.결과 전염siRNA기인편단적U251효질류세포주기인표체수평(U251세포위0.83±0.16、U251/vect세포위0.81 ±0.23、U251/S3위0.20±0.11)화단백표체수평(U251세포위157.73±4.12、U251/vect세포위153.34±5.27、U251/S3위112.20±3.16)현저하강(P＜0.05),MTT실험현시siRNA간우후U251효질류세포주체외증식속도감만、세포침습능력급연경지집락형성능력균현저강저(P＜0.01).결론 Nestin기인표체하조가억제효질류세포주U251적침습급증식능력.
Objective To investigate the therapeutic efficacy of siRNA fragments silencing Nestin,which may inhibits the growth and invasion of glioma cells.Methods The siRNA sequence fragments targeting Nestin were designed and transferred into human glioma cell line U251.Reverse transcription-polymerase chain reaction (RT-PCR) and immunochemistry method were used to explore the expression of Nestin.The tumor-suppressing effect of silencing Nestin on human glioma cells was assessed by methyl thiazol tetrazolium (MTT) and transwell chamber assays.Soft agar clone formation was adopted to identify oncogenicity.Results Nestin mRNA expression levels in U251 cells,U251/vect cells,U251/S3 cells were 0.83 ±0.16,0.81 ± 0.23,0.20 ± 0.11.Nestin protein expression levels in U251 cells,U251/vect cells,U251/S3 cells were 157.73 ± 4.12,153.34 ± 5.27,112.20 ± 3.16.MTT assays showed that the cell growth rate of U251 was significantly inhibited after transferred siRNA sequence fragments.The cellular adhesion ability and oncogenicity were weakly relevant.Conclusion Downregulation expression of Nestin can inhibits the growth of the human glioma cell line U251.