精胺普鲁兰-siRNA复合物转染白血病细胞方法的研究  

The Development of A Methodology to Transfect Leukemia Cells with Spermine-Introduced Pullulan-Sirna Complex

作　　者：万严 刘红[2] 赵昀 马文娟 WAN Yan;LIU Hong;ZHAO Yun;MA Wen-juan(Cyrus Tang Hematology Center,Soochow University,Suzhou,Jiangsu,215123,China;The First Affiliated Hospital of Soochow University,Suzhou,Jiangsu,215006,China)

机构地区：[1]苏州大学唐仲英血液学研究中心,江苏苏州215123 [2]苏州大学附属第一医院,江苏苏州215006

出　　处：《中国血液流变学杂志》2020年第2期139-143,148,F0002,共7页Chinese Journal of Hemorheology

基　　金：国家自然科学基金青年项目(81800151)。

摘　　要：目的 优化以精胺普鲁兰(Ps)为载体,递送siRNA到多种白血病细胞株及患者原代细胞的最佳条件.方法 Ps和siRNA按不同比例形成复合物,分别转染不同类型白血病细胞株,以优化转染条件;以最佳条件转染细胞,研究siRNA对靶基因表达的抑制状况;并研究Ps-siRNA复合物转染处于静息期的K562细胞和白血病患者CD34+细胞的能力.结果 数据显示Ps与siRNA的比例(N/P值)为1.2时,Ps-siRNA复合物对多种白血病/淋巴瘤细胞的转染最优,且细胞毒性较小(<10%);最优条件下Ps-siRNA复合物能有效抑制K562细胞中ID2的转录本和蛋白表达;该复合物也能有效转染患者静息期CD34+细胞(>80%).结论 Ps-siRNA复合物能高效转染多种白血病细胞株并抑制靶基因的表达,且能高效转染患者静息期CD34+细胞.该研究为白血病基础研究提供一种简便而高效的siRNA转染方法.Objective To optimize the delivery of siRNA to various leukemia cells using spennine-introduced pullulan(Ps)as a carrier.Methods Various amounts of Ps and siRNA were mixed to form Ps-siRNA nanoparticles,and then the transfection condition were optimized.The inhibitory effect of siRNA against its target gene was analyzed with the optimal condition.In addition,the abilities of Ps-siRNA nanoparticles to transfect quiescent K562 cells and CD34+ cells from leukemia patients were studied.Results The data showed that Ps-siRNA nanoparticles with a N/P value as 1.2 achieved optimal in vitro transfection of a variety of leukemia/lymphoma cells with low cytotoxicity(<10%).With the optimal condition,the Ps-siRNA nanoparticles effectively inhibited the expression of ID2 transcript and protein expression in K562 cells;the nanoparticles effectively transfected quiescent K562 cells as well as quiescent CD34 cells from leukemia patients(>80%).Conclusion We validated that Ps-siRNA nanoparticles were capable to transfect various leukemic cell lines and inhibit the expression of target gene,and they allowed effective transfection of quiescent CD34 cells from leukemia patients as well.Thus,we have developed a robust delivery tool for siRNA with modified pullulan,which will facilitate leukemia research ultimately.

关 键 词：精胺普鲁兰(Ps) SIRNA 白血病细胞 静息期细胞 

分 类 号：R733.7[医药卫生—肿瘤]