【摘要】 目的制备包载经聚乙二醇(PEG)修饰的重组腺病毒Ad-EGFP/HIF-1α高分子超声造影剂(PLGA),以改善腺病毒载体(Adv)在体内的安全性,提高Adv基因在细胞的转染效率。方法使用活化的甲氧基PEG对重组Adv衣壳蛋白质进行修饰得到PEG化Adv(PEG-Adv),用双乳化法制备载PEG-Adv高分子超声造影剂(PLGA),观测其粒径、包封率、释放规律及释放病毒的活性,并检测其在巨噬细胞中的免疫反应程度。结果 PLGA分布均匀,平均粒径为6.23μm;包封率为17.23%;PEG-Adv和单纯Adv从PLGA释放规律相似,但从PLGA释放出来的PEG-Adv的转染效率比从PLGA释放出来的单纯Adv高;包载PEG-Adv的PLGA产生的免疫反应强度也较低。结论用PLGA微球包载PEG化的重组腺病毒Ad-EGFP/HIF-1α,可改善Adv在体内安全性差的缺点,释放出来的病毒稳定性好,转染效率较高,作为心血管疾病的基因治疗载体具有较好的研究前景。更多还原

【Abstract】 Objective To prepare PEGylated recombinant adenovirus for microencapsulation within poly lactic-co-glycolic acid(PLGA)ultrasound contrast agents,in order to alleviate the safety problems and increasing gene transfection efficiency of adenovirus vector(Adv)at a target tissue site.Methods PEGylated Adv(PEG-Adv)was constructed by PEGylated adenoviral capsid proteins with activated methoxypolyethylene glycols.Then the recombinant adenovirus was encapsulated in PLGA using a water-in-oil-in-water(W/O/W)double emulsion and solvent evaporation method.The diameter,capsulated efficiency,release test and the bioactivity of viruses incorporated in vitro were studied,and the relative extent of the immune response for Adv and PEG-Adv encapsulated within PLGA microspheres was analyzed using macrophage cells.Results The mean diameter of PLGA carrying PEG-Adv was 6.23 μm.More than 17.23% of adenovirus was encapsulated in PLGA.Adv and PEG-Adv were both released from PLGA similarly in a sustained fashion.However,PEG-Adv microspheres demonstrated a higher GFP gene transfection efficiency than Adv microspheres released from PLGA.The PEG-Adv micropheres also exhibited a reduced extent of innate immune response for macrophage cells.Conclusion PEG-Adv can be formulated within PLGA increasing gene transfection efficiency and safety,which can be ideal gene therapy vectors for cardiovascular disease in future.更多还原