体内电穿孔对载体表达效率和人类免疫缺陷病毒1型DNA 疫苗免疫反应效果的研究

摘要

This study aims to evaluate the effect of electroporation on the expression of DNA vaccines in vivo and induced adaptive immune responses .The gene expression in vivo following DNA delivery via electropo-ration was determined by assessing reporter gene products of firefly luciferase by using in vivo imaging de-vice ,which was further dissected as the tissue distribution of gene products ,the peak level and the lasting time of expression when compared to the traditional immunization through muscular injection . The data showed that electroporation did not alter the tissue distribution ,but increased the peak level and lowered the plateau dosage of DNA vaccine .Further experiments with DNA vaccine pDRVI1-HIV expressing env from human immunodeficiency virus type 1 (HIV-1) CRF07_BC demonstrated that electroporation was able to enhance cellular and humoral immune responses which were gauged by the level of interferon-γ enzyme-linked immunospot (ELISPOT ) responses and HIV-1 specific binding antibody titers , respectively . Altogether ,electroporation in vivo technology is likely to be a useful tool in the administration of DNA vaccines .%本文旨在分析体内电穿孔（EP）技术对 DNA载体 pDRVI1．0表达效率和人类免疫缺陷病毒1型（HIV-1）DNA疫苗免疫反应的辅助效果，为其在DNA疫苗中的应用提供参考数据。通过构建携带荧光素酶基因的pDRVI1．0-Fluc质粒，利用活体成像技术分析EP接种对荧光素酶蛋白的组织分布、表达水平和持续时间的影响；同时，构建携带我国HIV-1 CRF07＿BC流行毒株 env基因的DNA疫苗pDRVI1．0-HIV ，利用酶联免疫斑点法（ELISPOT）、酶联免疫吸附试验（ELISA）和中和抗体法对 EP辅助免疫反应的特点进行分析。结果显示，EP接种后，pDRVI1．0-Fluc质粒未改变组织分布特点，但其体内表达效率显著提高，载体的饱和接种量降低。同时，EP技术提高了pDRVI1．0-HIV疫苗免疫小鼠后诱导的γ干扰素（IFN-γ）分泌型T细胞反应和Env特异性结合抗体效价。结果提示，EP技术可在DNA疫苗应用方面发挥作用。