IRAK-4基因沉默对人成骨样细胞凋亡的影响

【目的】研究siRNA沉默白细胞介素-1受体相关激酶-4(IRAK-4)基因对人成骨样细胞株MG63凋亡及凋亡相关基因Bcl-2、Bax表达的影响。【方法】 以Lipofectamine 2000为载体将IRAK-4-siRNA转染入MG63细胞(CON组:不加入任何转染试剂;SC组:以scrambled siRNA序列进行转染;KD组:以特异性IRAK-4-siRNA序列进行转染),应用流式细胞术及TUNEL法检测细胞凋亡水平,western blotting检测Bcl-2、Bax基因的表达。【结果】 与对照组相比,靶基因沉默组(KD组)细胞的IRAK-4 mRNA及蛋白表达水平显著降低;转染48h后,KD组细胞增殖变缓,凋亡增加,Bcl-2蛋白的表达明显下调且Bcl-2/BaxSC>Bcl-2/BaxKD; Spearman等级相关分析显示MG63细胞凋亡比例与Bcl-2/Bax的比值间有显著的相关性。 【结论】 siRNA沉默IRAK-4基因能够诱导人成骨样细胞株MG63凋亡,且靶细胞凋亡水平与Bcl-2、Bax基因表达水平的比值相关.

Effects of IRAK-4 Gene Silencing on Apoptosis in Human Osteoblast-like Cell

【Objective】 To explore the effects of siRNA-induced interleukin-1 receptor-associated kinase-4 (IRAK-4) gene silence on apoptosis of MG63 cells and the expression levels of related genes Bcl-2 and Bax.【Methods】 The gene transfection (control group = MG63 cells; SC group = MG63 cells transfected with scrambled IRAK-4 siRNA; KD group = MG63 cells transfected with IRAK-4-specific siRNA) was performed using Lipofectamine 2000. The apoptosis of MG63 cells were tested by FCM and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) methods. The protein level of Bcl-2 and Bax were detected by Western blotting. 【Result】 The expression of IRAK-4 mRNA and protein in the KD group were significantly decreased compared with other two groups. Compared with the control groups, 48 hours after the transfection, IRAK-4 gene silencing in MG63 cells caused morphological changes, inhibited growth, increased apoptosis (P < 0.05) and decreased protein expression of Bcl-2 and the ratio of Bcl-2/Bax (P <0.05). There was a significant correlation between the percentages of apoptotic MG63 cells and Bcl-2/Bax protein expression (P =0.011). 【Conclusion】 The results indicated that IRAK-4 gene silencing in MG63 cells increase apoptosis, which may be related to the decreased Bcl-2/Bax ratio.