心脏基因工程研究对一级康复的干预作用:构建柯萨奇B2病毒毒粒蛋白候选基因疫苗诱导体液免疫的评估  

Role of cardiac gene engineering in the primary rehabilitation induction of humoral immunity by constructing virion protein 1 candidate gene vaccine against coxsackievirus B2

作　　者：郭宏[1] 田野[1] 曲秀芬[1] 李为民[1] 孟繁超[1] 

机构地区：[1]哈尔滨医科大学第一临床医院心内科,黑龙江省哈尔滨市150001

出　　处：《中国临床康复》2005年第27期48-50,F0003,共4页Chinese Journal of Clinical Rehabilitation

基　　金：国家自然科学基金项目(30270553)~~

摘　　要：目的:探讨心脏基因工程研究中构建柯萨奇B2病毒毒粒蛋白侯选基因疫苗,并评价其诱导体液免疫的效果。方法:实验于2003-05/12在中国农业科学院国家生物重点实验室哈尔滨兽医研究所生物二室完成。采用反转录聚合酶链反应技术扩增柯萨奇B2病毒的主要中和抗原毒粒蛋白基因,通过分子克隆构建pcDNA3-柯萨奇B2病毒毒粒蛋白基因免疫质粒。选择3周龄BALB/c雄性小鼠30只,随机分成3组,pcDNA3-柯萨奇B2病毒毒粒蛋白组:对小鼠进行pcDNA3-柯萨奇B2病毒毒粒蛋白免疫;空载体对照组对小鼠进行pcDNA3免疫;磷酸盐缓冲液组:对小鼠进行磷酸盐缓冲液免疫,每组10只。每组免疫时间和接种量、接种方法相同。采用酶联免疫吸附法测定pcDNA3-柯萨奇B2病毒毒粒蛋白、pcDNA3、磷酸盐缓冲液免疫小鼠后2,4,6,8周柯萨奇B2病毒特异性抗体免疫球蛋白G水平。结果:①获得pcDNA3-柯萨奇B2病毒毒粒蛋白基因免疫质粒,可在HeLa细胞中表达。②免疫后6,8周pcDNA3-柯萨奇B2病毒毒粒蛋白组柯萨奇B2病毒特异性抗体免疫球蛋白G水平明显高于免疫后4周(A450:0.153±0.013,0.150±0.011,0.142±0.014,P<0.01,0.05);免疫后4周pcDNA3-柯萨奇B2病毒毒粒蛋白组柯萨奇B2病毒特异性抗体免疫球蛋白G水平明显高于空载体对照组和磷酸盐缓冲液组(0.120±0.007,0.119±0.009,P<0.01)。结论:pcDNA3-柯萨奇B2病毒毒粒蛋白可诱导小鼠产生体液免疫,做为候选基因疫苗,以基因免疫的角度参与心脏基因工程研究,有望为该病毒性心肌炎的发生起到一级预防作用。AIM：To construct a novel virion protein 1 （VPI） candidate gene vaccine against coxsackievirus B2 （CVB2） in cardiac gene engineering, and toevaluate the outcomes of humoral immunity induced by it. METHODS：The experiment was conducted in the Second Department of Biology,Harbin Veterinary Research Institute,State Key Laboratory for Biotechnology of The Chinese Academy of Agricultural Sciences from May to December 2003.Tbe major neutral antigen virion protein genes of CVB2 were proliferated by using reverse transcription-polymerase chain reaction （RT-PCR） technology.pcDNA3-CVB2 VPI was constructed by molecular cloning.Thirty BALB/c male mice at the age of 3 weeks were randomly and equally divided into 3 groups：pcDNA3-CVB2 VPI group,blank vector control group and phosphate buffer saline（PBS） group.pcDNA3-CVB2 VPI, pcDNA3 and PBS were used in the above 3 groups respectively. The immunity time,inoculation quantity and methods were all same in the three groups.The level of CVB2 specific immunoglobulin （IgG） of rats in the three groups was detected by enzyme-linked immunoadsordent assay （ELISA） at 2,4,6 and 8 weeks after corresponding intramuscular inoculation. RESULTS： ①A gene vaccine plasmid pcDNA3-CVB2 VPI was obtained, unich could be expressed in HeLa cells. ②The level of CVB2-specific antibody IgG detected at 6 and 8 weeks was higher than that at 4 weeks after first inoculation in pcDNA3-CVB2 VPI group （A450 0.153±0.013,0.150±0.011,0.142±0.014,P 〈 0.01,0.05）.At 4 weeks after first inoculation,the level of CVB2-specific antibody IgG in the pcDNA3-CVB2 VPI group was significantly higher than that in the blank vector control group （0.120±0.007） and PBS group（O.119＋0.009）（P 〈 0.01）. CONCLUSION：Humoral immunity will be induced by the constructed VPI gene vaccine against CVB2 in mice.As the candidate gene vaccine, pcDNA3-CVB2 VPI is expected to primarily prevent viral myocarditis in the research of cardiac gene engineering from the view of gene immune.

关 键 词：基因 疫苗 柯萨奇病毒感染 

分 类 号：R349.8[医药卫生—基础医学]