SiRNA沉默β-catenin对人牙周膜干细胞增殖能力的影响

Effects of siRNA-mediated β-catenin silence on proliferation in human periodontal ligament stem cells

目的:研究小干扰RNA(siRNA)干扰β-catenin基因表达对人牙周膜干细胞增殖能力的影响。方法:分离培养人牙周膜干细胞(periodontal ligament tissue-derived mesenchymal stem cells,PDLSCs)。应用β-catenin基因的siRNA转染PDLSCs,荧光显微镜下观察转染后细胞形态,流式细胞仪检测转染后GFP细胞表达率。流式细胞仪分析转染后细胞周期,用实时定量PCR技术检测转染后β-catenin、LEF-1、Cyclin D1 mRNA表达。结果:RNA干扰β-catenin处理PDLSCs 24小时后可特异并有效地抑制β-catenin基因的表达。siRNA抑制β-catenin使PDLSCs"S"期细胞减少,"G0,G1"期细胞增多;LEF-1及Cyclin D1 mRNA表达水平显著降低。结论:β-catenin si RNA可特异性抑制PDLSCs细胞中β-catenin基因表达,并明显抑制细胞增殖。

Objective： To investigate the effects of siRNA-induced specific silence of β-catenin on the proliferation of human periodontal ligament stem cells. Methods： Periodontal ligament stem cells were obtained from human healthy individuals（PDLSCs）. The PDLSCs were transfected with β-catenin siRNA. Cell morphology was observed under fluorescent microscope after transfection of PDLSCs. After transfection GFP expression rate were tested by flow cytometry of PDLSCs.After transfection we used flow cytometric analysis the cell cycle and tested the mRNA expression of β-catenin, LEF-1 and Cyclin D1 by Real time PCR. Results： β-catenin gene expression in PDLSCs was inhibited specifically and efficiently by the treatment of β-catenin-si RNA for 24 h and we found cell proliferation was significantly inhibited, with a reduction of S phase cells and a concomitant increase of G0 G1 phase cells; Real-time PCR results showed that the mRNA expression of LEF-1 and Cyclin D1 were significant reduced after transfection for 24 h in PDLSCs. Conclusion： β-catenin-si RNA could be employed to specifically inhibit the β-catenin gene express ion in PDLSCs, leading to significant inhibition of cell proliferation.