柯萨奇病毒B组3型亚单位疫苗与基因靶向疫苗免疫小鼠的效果比较

目的:观察柯萨奇病毒VP1基因靶向核酸疫苗pcDNA3/C3d3-sVP1、重组腺病毒rAd/C3d3-sVP1和亚单位疫苗VP1蛋白的免疫效果。方法:雄性BALB/c小鼠随机分为4组,每组18只,分别肌肉注射PBS、pcDNA3/C3d3-sVP1、rAd/C3d3-sVP1和VP1蛋白,除重组腺病毒rAd/C3d3-sVP1组免疫2次,间隔2周外,其余3组均免疫3次,间隔3周。质粒每次每只接种100μg/100μl,腺病毒每次每只接种1.2×107pfu/100μl,蛋白每次每只接种50μg。分别用ELISA法和微量中和试验法检测血清CVB3 VP1特异性IgG抗体和中和抗体;CCK-8法检测脾淋巴细胞特异性CTL杀伤活性;以致死量的CVB3病毒液感染已免疫小鼠,检测小鼠血中病毒滴度,观察存活情况以评价各种疫苗的免疫保护作用。结果:各实验组小鼠的特异性IgG抗体和中和抗体滴度随免疫次数增加而提高,末次免疫后,VP1蛋白组的特异性IgG抗体和中和抗体滴度均明显高于pcDNA3/C3d3-sVP1组和rAd/C3d3-sVP1组(P<0.05),但rAd/C3d3-sVP1组CTL杀伤活性高于pcDNA3/C3d3-sVP1组和VP1蛋白组;经致死量CVB3感染后,VP1蛋白组血中病毒滴度低于其他实验组,而生存率明显高于其他各组。结论:VP1蛋白疫苗诱导小鼠产生较强的特异性免疫应答,提高了小鼠的生存率,免疫效果优于靶向基因疫苗pcDNA3/C3d3-sVP1和rAd/C3d3-sVP1。

Comparative immunogenicity of subunit vaccine expressing CVB3 VP1 and targeted gene vaccine in mice

Objective:To compare the immunogenicity of targeted gene vaccine pcDNA3/C3d3-sVP1,rAd/C3d3-sVP1 and subunit vaccine expressing coxsackievirus B3(CVB3) VP1 in mice.Methods:Male BALB/c mice were divided into 4 groups randomly,18 for each,and injected imtramuscularly(i.m.) PBS,pcDNA3/C3d3-sVP1,rAd/C3d3-sVP1 and VP1 protein.In addition to twice at two week interval with rAd/C3d3-sVP1 group,the other 3 groups were inoculated once three week for 3 times.The mice were immunized with plasmid pcDNA3/C3d3-sVP1 at a dose of 100 μg /100 μl,rAd/C3d3-sVP1 at a dose of 1.2×107 pfu/100 μl and protein VP1 at a dose of 50 μg.The titers of sera CVB3 VP1 IgG antibody and neutralizing antibody were detected by ELISA and micro-neutralization assay,respectively;the spleen lymphocytes specific CTL cytotoxic activity were detected with CCK-8 assay.The others were challenged with lethal dose of CVB3,the titers of sera virus were titrated and the protection efficacy against CVB3 was observed.Results:The titers of special IgG antibody and neutralizing antibody in sera of mice in every experimental groups increased with the time of inoculation.More specifically,special IgG antibody and neutralizing antibody in VP1 immunized protein were remarkably stronger than others after the last immunization;the specific CTL activity of the spleen lymphocytes of rAd/C3d3-sVP1 group was highest in three groups.After challenged with lethal dose of CVB3,the virus titers of blood in VP1 immunized protein were significantly decreased(P<0.05) while its protection rate was significantly higher than other two groups.Conclusion:Subunit vaccine can induce a stronger immunological effect,result in a higher survival rate,and had better influence than targeted gene vaccine pcDNA3/C3d3-sVP1 group and rAd/C3d3-sVP1 group.