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本实验将基于埃博拉病毒表面糖蛋白抗原GP基因序列,按照哺乳动物密码子使用频率优化,并与人IgG恒定区构建融合蛋白GP-Fc基因序列,重组蛋白序列插入基因疫苗载体pVR中,构建重组蛋白基因疫苗pVRmodGP-Fc。通过基因疫苗导入系统免疫小鼠,用间接ELISA和间接免疫荧光对抗体效价进行评估。实验结果显示,重组蛋白GP-Fc的基因疫苗可以很好的刺激小鼠产生特异性抗体,免疫周期结束后,小鼠血清中结合效价终点达到高剂量组1∶300 000及低剂量组1∶180 000,同时血清中的抗体可以很好地结合表达GP抗原的细胞表面,表现出很强的荧光信号。通过该研究我们验证重组蛋白基因疫苗pVR-modGP-Fc可以很好地诱导BALB/c小鼠产生较高滴度的抗原特异性IgG,具有较好的免疫原性。
Abstract：We constructed the recombination protein gene vaccine pVR-modGP-Fc.The gene code of modGP-Fc(in which the Ebola glycoprotein(GP)extracellular domain is modified by the codon frequency of humans and fused with the human IgG Fc code at the 3′terminal)was synthesized and inserted into a pVR vector.BALB/c mice were vaccinated thrice(0,3,and 6weeks)at two doses.During the immunization period,serum antibody titers were evaluated by an indirect enzyme-linked immunoassay(ELISA).Immunofluorescences were applied to manifest the bond efficiency of serum antibodies to cell-expressed Ebola-virus GPs.Specific antibodies were found 1week after the first dose in each vaccination group.At the end of the immunization period,serum ELISA titers increased to 1∶300 000 at a high dose and 1∶180 000 at a low dose.Specific serum antibodies were well bonded to the surface of GP-expressed HEK 293 Tcells,and fluorescence was not detected in controls.These data suggest that the DNA vaccine pVR-modGP-Fc can induce BALB/c mice to generate high-efficiency combination IgG,and that it shows good immunogenicity.Our study could be the basis for development of a vaccine against the Ebola virus.
[1]Baize S,Pannetier D,Oestereich L,et al.Emergence of Zaire Ebola virus disease in Guinea[J].N Engl J Med,2014,371(15):1418-1425.
[2]Muyembe-Tamfum J J,Mulangu S,Masumu J,et al.Ebola virus outbreaks in Africa:Past and present[J].Onderstepoort J Vet Res,2012,79(2):451.
[3]Falasca L,Agrati C,Petrosillo N,et al.Molecular mechanisms of Ebola virus pathogenesis:focus on cell death[J].Cell Death Differ,2015,22(8):1250-1259.
[4]Mohamadzadeh M,Chen L,Schmaljohn A L.How Ebola and Marburg viruses battle the immune system[J].Nat Rev Immunol,2007,7(7):556-567.
[5]Kaner J,Schaack S.Understanding Ebola:the 2014epidemic[J].Globalization and Health,2016,12(1):53.
[6]Tong Y,Shi W,Liu D,et al.Genetic diversity and evolutionary dynamics of Ebola virus in Sierra Leone[J].Nature,2015,524(7563):93-96.
[7]Sullivan N J,Martin J E,Graham B S,et al.Correlates of protective immunity for Ebola vaccines:implications for regulatory approval by the animal rule[J].Nat Rev Microbiol,2009,7(5):393-400.
[8]Ye L,Yang C.Development of vaccines for prevention of Ebola virus infection[J].Microbes and Infection,2015,17(2):98-108.
[9]Fausther-Bovendo H,Mulangu S,Sullivan N J.Ebolavirus vaccines for humans and apes[J].Current Opinion in Virology,2012,2(3):324-329.
[10]Wang D,Raja N U,Trubey C M,et al.Development of a cAdVax-based bivalent ebola virus vaccine that induces immune responses against both the Sudan and Zaire species of Ebola virus[J].J Virol,2006,80(6):2738-2746.
[11]Henao-Restrepo A M,Longini I M,Egger M,et al.Efficacy and effectiveness of an rVSV-vectored vaccine expressing Ebola surface glycoprotein:interim results from the Guinea ring vaccination cluster-randomised trial[J].Lancet,2015,386(9996):857-866.
[12]Jones S M,Feldmann H,Stroher U,et al.Live attenuated recombinant vaccine protects nonhuman primates against Ebola and Marburg viruses[J].Nat Med,2005,11(7):786-790.
[13]Kutzler M A,Weiner D B.DNA vaccines:ready for prime time?[J].Nature Reviews Genetics,2008,9(10):776-788.
[14]Gurunathan S,Klinman D M,Seder R A.DNA vaccines:immunology,application,and optimization[J].Annu Rev Immunol,2000,18:927-974.
[15]Donnelly J J,Wahren B,Liu M A.DNA Vaccines:Progress and Challenges[J].J Immunol,2005,175(2):633-639.
[16]张晓光,杨韧,王娇,等.埃博拉病毒重组膜蛋白Gp-Fc表达及免疫原性研究[J].病毒学报,2016,32(1):1-5.
[17]Sasaki S.Adjuvant formulations and delivery systems for DNA vaccines[J].Methods,2003,31(3):243-254.
[18]Li L,Petrovsky N.Molecular mechanisms for enhanced DNA vaccine immunogenicity[J].Expert Rev Vaccines,2016,15(3):313-329.
[19]O'Hagan D T,Singh M,Ulmer J B.Microparticles for the delivery of DNA vaccines[J].Immunol Rev,2004,199(1):191-200.
基本信息:
DOI：10.13242/j.cnki.bingduxuebao.003120
中图分类号:R392
引用信息:
[1]杨韧,张晓光,王娇,等.扎伊尔型埃博拉重组糖蛋白GP-Fc基因疫苗构建及免疫原性研究[J].病毒学报,2017,33(02):186-191.DOI:10.13242/j.cnki.bingduxuebao.003120.
基金信息:
埃博拉出血热防控应急研究(项目号:1061400100275)
2017-03-29
2017-03-29
2017-03-29