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目的:探讨siRNA靶向沉默CDX2基因联合长春新碱(vincristine,VCR)对白血病K562细胞增殖、凋亡的影响。方法:设计靶向沉默CDX2基因表达的特异性siRNA序列CDX2-siRNA-921和相应的阴性对照序列CDX2-siRNA-NC,分别转染K562细胞,同时设正常细胞组做对照。分别应用Real-time PCR和Western blotting法检测转染CDX2-siRNA-921对细胞内CDX2、BCR-ABL mRNA和蛋白表达的影响;siRNA和VCR单独或联合作用于K562细胞后,应用MTT、流式细胞术分别检测细胞增殖抑制率、凋亡率的变化。结果:与正常细胞组相比,CDX2-siRNA-921能够显著降低目的基因CDX2和BCR-ABL mRNA与蛋白的表达(均P<0.05),而CDX2-siRNA-NC组CDX2和BCR-ABL mRNA与蛋白表达量无明显变化;与VCR组、CDX2-siRNA-NC组和VCR+CDX2-siRNA-NC组相比,VCR+CDX2-siRNA-921组细胞增殖抑制率降低,凋亡率明显增加(均P<0.05)。结论:CDX2-siRN-921能显著降低K562细胞内CDX2 mRNA和蛋白的表达,并能够下调BCR-ABL融合基因表达水平,同时增强VCR对K562细胞增殖抑制、凋亡诱导作用。
OBJECTIVE: To investigate the effects of siRNA targeted silencing CDX2 gene combined with vincristine (VCR) on the proliferation and apoptosis of leukemia K562 cells. Methods: CDX2-siRNA-921 and CDX2-siRNA-NC were designed to target specific silencing CDX2 gene and transfected into K562 cells respectively. The effect of CDX2-siRNA-921 transfection on the expression of CDX2 and BCR-ABL mRNA and protein were detected by Real-time PCR and Western blotting respectively. After treated with siRNA or VCR alone or in combination with K562 cells, Cytometry were detected cell proliferation inhibition rate, apoptosis rate changes. Results: CDX2-siRNA-921 significantly decreased the mRNA and protein expression of CDX2 and BCR-ABL (all P <0.05) compared with normal cells, while CDX2 and BCR-ABL mRNA and CDX2- Compared with VCR group, CDX2-siRNA-NC group and VCR + CDX2-siRNA-NC group, the cell proliferation inhibition rate and the apoptosis rate of VCR + CDX2-siRNA-921 group were significantly increased P <0.05). CONCLUSION: CDX2-siRN-921 can significantly decrease the expression of CDX2 mRNA and protein in K562 cells and down-regulate the expression of BCR-ABL fusion gene. Meanwhile, CDX2-siRN-921 can inhibit the proliferation and induce the apoptosis of K562 cells.