柯萨奇病毒B3的VP1 DNA疫苗与蛋白或重组腺病毒疫苗不同组合免疫方式的免疫效果观察  

Generation of protective immune responses against coxsackievirus B3 challenged by DNA priming VP1 protein or recombinant adenovirus boosted vaccination

作　　者：蓝佳明[1] 高志云[1] 金玉怀[1] 李剑[1] 刘贵霞[1] 羡鲜[1] 张永红[1] 王永祥[1] 

机构地区：[1]河北医科大学病原生物教研室,石家庄050017

出　　处：《中国人兽共患病学报》2012年第1期41-45,50,共6页Chinese Journal of Zoonoses

基　　金：河北省医学科学研究重点课题资助项目(20090054;20090049)

摘　　要：目的观察柯萨奇病毒B3(Coxsackievirus B3,CVB3)衣壳蛋白VP1DNA疫苗初免后VP1蛋白或重组腺病毒rAd/VP1加强的prime-boost策略的免疫效果。方法用CVB3VP1的真核表达质粒pcDNA3/VP1初次免疫小鼠后,分别用VP1蛋白或重组腺病毒rAd/VP1加强免疫2次。检测免疫小鼠血清特异性IgG抗体、中和抗体滴度以及脾脏细胞毒性T淋巴细胞(cytotoxic lymphocytes,CTLs)杀伤活性;致死量CVB3攻击后,检测小鼠血中病毒滴度并观察动物的存活情况。结果 pcDNA3/VP1+VP1蛋白组小鼠血清IgG抗体、中和抗体滴度以及动物生存率明显高于pcDNA3/VP1+rAd/VP1组和pcDNA3/VP1质粒组(P<0.05);pcDNA3/VP1+rAd/VP1组和pcDNA3/VP1+VP1蛋白组小鼠脾脏CTLs杀伤活性明显高于pcDNA3/VP1质粒组(P<0.05)。结论质粒pcDNA3/VP1初次免疫后,VP1蛋白或重组腺病毒rAd/VP1加强的prime-boost策略有较好的免疫效果,两者比较pcDNA3/VP1+VP1蛋白prime-boost免疫策略的效果更为明显。In order to compare the immune effects of 2 different prime-boost strategies,BALB/c mice were primed immunization with eukaryotic expression plasmid pcDNA3/VP1 contained the VP1 capsid gene of coxsackievirus B3（CVB3）,following by two boosts with the VP1 protein from procaryotic cells or recombinant adenovirus rAd/VP1.Mice were bled from the retroorbital sinus plexus every 2 weeks after each immunization.Enzyme-linked immunosorbent assay and micro-neutralization test were used to detect levels of CVB3-specific IgG antibody and neutralizing antibody titers in the sera of immunized mice.Three weeks after the last immunization,the Cytotoxic T lymphocytes（CTLs） killing activity of spleen lymphocytes was detected with Cytotoxicity Counting Kit-8 assay.Followed the 3 immunization,mice were challenged with CVB3 infection.Virus titers in the sera of immunized mice were determined by the 50% cell culture infective dose assay on HeLa cell monolayers.Percentage of animals surviving was observed after lethal CVB3 attacking over a period of 21 days.The results showed the titers of specific IgG antibody and neutralizing antibody in sera of the pcDNA3/VP1 and VP1 protein regime immunized mice were higher（P〈0.05） than the other 2 groups including pcDNA3/VP1 and pcDNA3/VP1＋rAd/VP1 groups.CTLs killing activity of spleen lymphocytes of the pcDNA3/VP1＋rAd/VP1 immunized mice were higher（P〈0.05） than the pcDNA3/VP1 or pcDNA3/VP1＋VP1 protein group.In addition,the pcDNA3/VP1＋VP1 protein vaccine resulted to the strongest protection of mice from lethal CVB3 challenge and the lowest viral load in sera of immunized mice after acute CVB3 infection.It has been concluded pcDNA3/VP1 priming and VP1 protein or rAd/VP1 boosting immunization had obvious immune responses against CVB3 infection in mice.The pcDNA3/VP1 priming and VP1 protein boosting regimen surpassed plasmid pcDNA3/VP1 and recombinant adenovirus rAd/VP1 regime and might be a promising vaccine candidate against CVB3 infection.

关 键 词：柯萨奇病毒B3 PRIME-BOOST DNA 蛋白 重组腺病毒 

分 类 号：Q373.2[生物学—遗传学]