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目的 比较修饰型及未修饰型 Rel A反义脱氧寡核苷酸 ( AS- ODN)在人白血病细胞 HL- 60中的分布及稳定性。方法 将荧光素 ( FAM)标记的硫代磷酸酯修饰型及未修饰型 AS- ODN经脂质体介导或直接转染人 HL- 60细胞 ,应用荧光显微镜及流式细胞仪观察细胞内荧光的时相分布。结果 发现 1μm ol/ L 修饰型 AS- ODN进入细胞后 ,细胞内荧光强度 6h达到高峰。在有脂质体存在时 ,细胞内荧光明显增强 ,12 h后荧光减弱并逐渐消失 ;而 AS- ODN直接转染细胞 ,细胞内特别是核内荧光强度明显比脂质体组弱。未修饰型 AS- ODN直接或经脂质体介导转染 ,细胞内荧光强度均很弱 ,滞留时间不超过 4h。结论 脂质体可以提高细胞对 AS- ODN的摄取及核聚积 ,而硫代磷酸酯修饰型 AS-ODN具有更好的稳定性
Objective To compare the distribution and stability of modified and unmodified Rel A antisense oligodeoxynucleotides (AS-ODNs) in human leukemia HL-60 cells. Methods Fluorescein (FAM) labeled phosphorothioate-modified and unmodified AS-ODN were transfected into human HL-60 cells by liposomes, and fluorescence microscopy and flow cytometry were used to observe intracellular fluorescence The distribution of time. The results showed that 1μmol / L modified AS-ODN into cells, the intracellular fluorescence intensity peaked at 6h. In the presence of liposomes, intracellular fluorescence was significantly enhanced, after 12 h the fluorescence decreased and gradually disappear; and AS-ODN transfected cells, the intracellular especially nuclear fluorescence intensity was weaker than the liposome group. Unmodified AS-ODN directly or via liposome-mediated transfection, intracellular fluorescence intensity are very weak, residence time does not exceed 4h. Conclusion Liposomes can enhance the cellular uptake of AS-ODN and nuclear accumulation, while the phosphorothioate-modified AS-ODN has better stability