活性氧响应的新型阳离子共聚物构建及其基因递送性能研究  

Construction of a Novel Reactive Oxygen Species-responsive Cationic Copolymer and Its Performance in Gene Delivery

作　　者：韩旭[1] 张留伟 张强[1] 睢晞航 钱明 陈麒先 王静云[1] Xu Han;Liuwei Zhang;Qiang Zhang;Xihang Sui;Ming Qian;Qixian Chen;Jingyun Wang(School of Bioengineering,Dalian University of Technology,Dalian 116024,China)

机构地区：[1]大连理工大学生物工程学院,大连116024

出　　处：《化学学报》2021年第6期794-802,共9页Acta Chimica Sinica

基　　金：国家自然科学基金(Nos.21878041,22078050);中央高校基本科研业务费专项资金(Nos.DUT17RC(3)059,DUT20YG126);大连市科技创新基金(Nos.2020JJ26SN050,2020JJ26GX025)资助.

摘　　要：引入快速、主动释放基因的机制是提升非病毒基因递送效率的关键.本研究以2-甲基丙烯酰氧乙基磷酰胆碱(MPC)与(2-丙烯酰基)乙基(硼酸苄基)二乙基溴化铵(BD)为单体,基于可逆加成-断裂链转移聚合(RAFT)反应制备具有活性氧响应性质的聚阳离子嵌段共聚物pM-pBD.通过静电作用,阳离子共聚物pM-pBD能够与带负电荷的DNA以分子自组装的方式形成纳米复合物.其中,阳离子pBD片段具有活性氧触发电荷反转的特性,因此,有助于获得活性氧触发的静电组装复合体结构解离,从而实现可控基因释放的功能.理化表征结果表明,pM-pBD与质粒DNA静电复合后能够形成粒径约为99.1 nm,ζ电位约为+13.8 mV,显微形貌近似球形的纳米复合物.在加入促过氧化氢产生的抗坏血酸后,上述pM-pBD基因递送系统的转染效率得到了显著的提升.因此,本研究创制的pM-pBD为基因递送系统的可控释放提供了新的解决方案.The development of rapid and active gene release function is the key to improve the efficiency of non-viral gene delivery system.Herein,a novel reactive oxygen-responsive cationic block copolymer non-viral gene delivery vector(termed as pM-pBD)consisting of biocompatible poly(2-methacryloyloxyethyl phosphorylcholine)(pMPC)segment and charge re versible poly[(2-acryloyl)-ethyl-(boronic acid benzyl)-diethylammonium bromide]segment(pBD)was synthesized via re versible addition-fragmentation chain transfer polymerization(RAFT).Dynamic light scattering(DLS)assay andζpotential measurements,transmission electron microscope(TEM)and gel electrophoresis were performed to characterize pM-pBD/(plasmid DNA,pDNA)complexes.Gel electrophoresis retardation assay displayed that pM-pBD can firmly bind pDNA through electrostatic interaction even in the presence of high concentration of heparin,but capable of releasing pDNA in response to reactive oxygen(such as H2O2).The pM-pBD/pDNA complexes show low cytotoxicity against HeLa cells,even at high weight ratio of pM-pBD and pDNA(N/P)(up to 16)demonstrated by MTT(methylthiazolyldiphenyl tetrazolium bromide)assay.The pM-pBD/pDNA complexes demonstrated appreciable colloidal stability in the presence of 10%fetal bovine serum.The pM-pBD/pDNA complexes at N/P ratio of 3 displayed spherical morphologies with average diameter of approximate 99.1 nm andζpotential of approximate+13.8 mV.However,once upon incubation in presence of 1 mmol/L H2O2,the diameter of pM-pBD/pDNA complexes at N/P ratio of 3 was enlarged to 330 nm andζpotential was reversed to-3.91 mV due to charge reversal of BD responsive to reactive oxygen.Flow cytometry revealed uptake effi-ciency(88.9%)and the highest transfection efficiency(31.2%)for pM-pBD at N/P ratio of 3 and 16,respectively.The trans fection efficiency of pM-pBD/pDNA against HeLa cells was observed to be significantly augmented(1.5-fold)after the addi tion of ascorbic acid,which could stimulate the production of hydrogen peroxide.Therefore,pM-pBD represented

关 键 词：聚阳离子 电荷反转 基因递送 活性氧响应 抗坏血酸 

分 类 号：TQ317[化学工程—高聚物工业] R450[医药卫生—治疗学]