脂質体转染反义脱氧寡核苷酸对K562细胞α珠蛋白基因表达及细胞增殖的影响.doc

摘要】 本研究查明脂质体转染反义脱氧寡核苷酸（ASON）对K562细胞α珠蛋白基因表达的抑制作用及对细胞增殖的影响，探讨基因调控治疗β地中海贫血的新思路。 设计合成靶向α珠蛋白基因的ASON，并与正义寡核苷酸（SON）和空白对照进行比较。采用脂质体转染技术将ASON、SON与K562细胞共同培养，用荧光显微术、流式细胞术检测脂质体转染效率，用实时荧光定量RTPCR法检测K562细胞中α珠蛋白基因表达情况，并通过Cell Counting Kit8（CCK8）法观察脂质体转染ASON技术对细胞增殖的影响。结果表明：脂质体转染效率在ASON作用24h达到最高，脂质体转染ASON组的α珠蛋白基因表达水平显著低于SON和空白对照组（p0.01），并对细胞的增殖有明显的抑制作用，上述效应呈浓度依赖性。结论：脂质体转染ASON能特异性的抑制K562细胞中α珠蛋白基因表达，可能会成为β地中海贫血基因治疗的一个新靶点。 【关键词】 脂质体转染 Effect of Liposomal Transfection of Antisense Oligodeoxynucleotide on Alphaglobin Gene Expression and Proliferation of K562 Cells LIU RongRong,LAI YongRong,MA Jie Department of Hematology, The First Affiliated Hospital, Guangxi Medical University,Nanning 530021, China Abstract The objective of study was to investigate the effect of liposomal transfection of antisense oligodeoxynucleotide (ASON) on alphaglobin gene expression and proliferation of K562 cells, to explore the new way of gene therapy in βthalassemia. Targeted ASON of alphaglobin was designed and synthesized, and compared with positive control ［sense oligodeoxynucleotide (SON) group］ and blank control. By liposomal transfection, ASON, SON was cocultured with K562. The efficiency of trasfection was assayed by fluorescence microscopy and flow cytometry(FCM), the alphaglobin gene expression of K562 was measured by realtime PCR, and the proliferation of K562 was determined by Cell Count Kit8 assay. The results indicated that the highest efficiency was at 24 hours after liposomal transfection, the gene expression level of alphaglobin in ASON group was significantly lower than that in SON group and blank control (p 0.01). The proliferation of K562 cells was obviously inhibited, meanwhile the above effect showed the dosedependent manner. It is concluded that the liposomal transfection of ASON inhibits the alphaglobin gene expression of K562 cells, which may be the new target for gene therapy in βthalassemia. Key words alphaglobin g