三个不同亚型HIV-1调节/辅助基因DNA疫苗的构建和免疫原性研究  

Immunogenicity of DNA vaccines encoding regulatory/accessory proteins derived from three different prevalent strains in China

作　　者：解晓燕[1] 万延民[2] 李斌[4] 史继静[4] 仇超[3] 刘朝奇[4] 徐建青[2] 张焕相[1] 

机构地区：[1]苏州大学基础医学与生物科学学院,215123 [2]上海市(复旦大学附属)公共卫生临床中心,201508 [3]复旦大学生物医学研究院 [4]三峡大学

出　　处：《中华微生物学和免疫学杂志》2011年第2期157-161,共5页Chinese Journal of Microbiology and Immunology

摘　　要：目的 构建表达中国主要流行亚型HIV-1 tat-rev-integrase（c-half）-vif-nef（TRIVN）融合基因的DNA疫苗,并比较其免疫原性.方法 按人源密码子使用频率对HIV-1 CN54（B＇/C重组亚型）与RL42（B＇亚型）的tat、rev、integrase（C端144个氨基酸）、vif和nef基因序列进行优化,构建DNA疫苗.通过Western blot测定上述DNA疫苗与HIV-1 AE2f株来源的tat-rev-integrase（c-half）-vifnef融合基因DNA疫苗的体外表达效率 利用小鼠模型比较3个DNA疫苗单独免疫与混合免疫的免疫原性特征.结果 限制性酶切及DNA测序结果表明两个融合基因重组质粒构建正确 Western blot 检测结果显示：3个DNA疫苗的体外表达效率基本相当.小鼠免疫后ELISPOT检测结果显示：在总T细胞反应强度方面AE2f-TRIVN最强[（948.0±330.0）SFCs/10^6脾细胞],次之为混合DNA免疫组（500.0±155.0 SFCs/10^6脾细胞）,再者为RL42-TRIVN[（195.1±44.0）SFCs/106脾细胞],CN54-TRIVN最弱[（89.5±17.0）SFCs/106脾细胞].T细胞反应分布情况显示：3个DNA疫苗单独免疫时,T细胞反应主要集中在Integrase和Vif蛋白上 而混合免疫可以部分改善针对Nef蛋白的免疫识别.结论 3个亚型TRIVN DNA疫苗中以AE2f-TRIVN的免疫原性最强 DNA疫苗混合免疫倾向于促进特异性T细胞反应在TRIVN融合抗原上的均匀分布.0bjective To determine the immunogenicities of DNA vaccines expressing tat-rev-integrase（c-half）-vif-neffusion genes（TRIVN） derived from prevalent B＇, B＇/C and AE recombinant subtypes of HIV-1 in China. Methods Two DNA vaccines were constructed by inserting the codon optimized tat-revintegrase（c-half）-vif-nef fusion genes derived from B＇ and B＇/C subtype of HIV-1 into mammalian expression vector pSVI. 0. DNA vaccine containing tat-rev-integrase （c-half）-vif-nef fusion gene derived from HIV-1AE2f has been constructed previously. In vitro expression efficiencies of three DNA vaccines were determined by Western blot and their immunogenicities were compared by immunizing female BALB/c mice. IFN-γ ELISPOT assay was used to read out the specific T cell immunity. Results The constructed DNA vaccines were validated by restriction enzyme digestion and DNA sequencing. Western blot assay showed three constructed DNA vaccines could be expressed at a comparable level in vitro. After vaccination, AE-TRIVN mounted T cell immune responses at （948.0 ± 330.0） SFCs/106 splenocytes, followed by the mixed DNA vaccine[ （500.0 ± 155.0） SFCs/10^6 splenocytes ], RL-TRIVN r[ （ 195. 1 ± 44.0） SFCs/10^6 splenocytes ]and CN-TRIVN [ （89.5 ± 17.0） SFCs/10^6 splenocytes]. Interestingly, we observed that single DNA vaccination induced specific T cell responses predominantly targeting Integrase （C-half） and Vif, whereas the mixed DNA could significantly improve T cell responses against Nef. Conclusion AE-TRIVN was the most immunogenic among the three DNA vaccines and the mixed DNA vaccination could change the immunogenic hierarchy of T cell epitopes across the fusion genes vaccine.

关 键 词：艾滋病病毒 疫苗 T细胞 调节/辅助蛋白 亚优势表位 

分 类 号：R392[医药卫生—免疫学]