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目的:探讨RNA干扰技术沉默CDX2基因对人急性髓细胞白血病细胞系THP1增殖影响的意义。方法:针对CDX2mRNA序列设计合成2对编码小干扰RNA(siRNA)的DNA模板,构建pGenSil-CDX2siRNA重组质粒,转染THP1细胞。采用蛋白质印迹法检测重组质粒对CDX2蛋白表达的影响,MTT法观察THP1细胞增殖情况,Annexin-V-PI双染法流式细胞术检测转染重组质粒后细胞凋亡状况。结果:成功构建pGenSil-CDX2 siRNA重组质粒,成功转染THP1细胞,并能特异地抑制CDX2基因的表达;转染重组质粒后,THP1的活力分别降低为(39.8±5.0)%,与空质粒组组间差异有统计学意义,P=0.016。重组质粒组的THP1细胞凋亡率为37.3%~42.1%。结论:pGenSil-CDX2 siRNA重组质粒明显下调CDX2在急性髓细胞白血病细胞中的表达,并抑制肿瘤细胞生长,促进其凋亡。
Objective: To investigate the effect of silencing CDX2 gene on the proliferation of human acute myeloid leukemia cell line THP1 by RNA interference. METHODS: Two pairs of DNA templates encoding small interfering RNA (siRNA) were designed and synthesized according to the CDX2 mRNA sequence. The recombinant plasmids pGenSil-CDX2siRNA were constructed and transfected into THP1 cells. The expression of CDX2 protein was detected by Western blotting. The proliferation of THP1 cells was observed by MTT assay. The apoptosis of THP1 cells was detected by Annexin-V-PI double staining and flow cytometry. Results: The recombinant plasmid pGenSil-CDX2 was successfully constructed and successfully transfected into THP1 cells and specifically inhibited the expression of CDX2 gene. After transfection with recombinant plasmid, THP1 activity was reduced to (39.8 ± 5.0)%, The difference between the groups was statistically significant, P = 0.016. The apoptosis rate of THP1 cells in recombinant plasmid group was 37.3% ~ 42.1%. Conclusion: pGenSil-CDX2 siRNA recombinant plasmid can significantly down-regulate the expression of CDX2 in acute myeloid leukemia cells and inhibit the growth of tumor cells and promote their apoptosis.