CAJ | 학술논문

目的研究登革2型病毒(Dengue virus type 2,DENV-2)病毒样颗粒(virus-Like particles,VLPs)的免疫原性.方法利用已构建的DENV-2 ZS01/01株病毒样颗粒的表达质粒转染293T细胞,对分泌型VLPs进行大量培养并通过蔗糖密度梯度离心法对其进行纯化.纯化的VLPs经Western Blot及透射电镜观察等方法鉴定后免疫BALB/c小鼠.利用ELISA及中和试验等方法对体液免疫反应进行检测,ELISPOT法测定细胞免疫水平.结果登革2型病毒样颗粒表达质粒转染哺乳动物细胞所得上清经蔗糖密度梯度离心后,电镜下可观察到类似于天然登革病毒的大小在45～55nm之间的病毒样颗粒.体液及细胞免疫检测结果显示登革2型VLPs可以刺激小鼠产生较高水平的登革E蛋白特异性抗体及一定水平的中和抗体,免疫小鼠脾淋巴细胞经体外刺激后IFN-γ水平显著升高.结论登革2型病毒病毒样颗粒免疫BALB/c小鼠后可引起一定水平的细胞免疫及体液免疫反应,该研究结果为四价登革病毒样颗粒疫苗的研制奠定了基础.
목적 연구등혁2형병독(Dengue virus type 2,DENV-2)병독양과립(virus-Like particles,VLPs)적면역원성.방법 이용이구건적DENV-2 ZS01/01주병독양과립적표체질립전염293T세포,대분비형VLPs진행대량배양병통과자당밀도제도리심법대기진행순화.순화적VLPs경Western Blot급투사전경관찰등방법감정후면역BALB/c소서.이용ELISA급중화시험등방법대체액면역반응진행검측,ELISPOT법측정세포면역수평.결과 등혁2형병독양과립표체질립전염포유동물세포소득상청경자당밀도제도리심후,전경하가관찰도유사우천연등혁병독적대소재45～55nm지간적병독양과립.체액급세포면역검측결과현시등혁2형VLPs가이자격소서산생교고수평적등혁E단백특이성항체급일정수평적중화항체,면역소서비림파세포경체외자격후IFN-γ수평현저승고.결론 등혁2형병독병독양과립면역BALB/c소서후가인기일정수평적세포면역급체액면역반응,해연구결과위사개등혁병독양과립역묘적연제전정료기출.
Objective To study the immunogenicity of virus-like particles (VLPs) of dengue virus type 2 (DENV-2) ZS01/01 strain.Methods Secreted VLPs of DENV-2 ZS01/01 strain were purified by ultracentrifugation from 293T cells transfected with plasmid which was established previously in our lab.After identified by Western Blot analysis and transmission electron microscope (TEM),the immunogenicity of DENV-2 VLPs was evaluated in BALB/c mice.Humoral immune responses were identified by ELISA and CPE-based neutralization analysis while cellular immune responses were detected by ELISPOT.Results The analysis of humoral immune responses revealed that DENV-2 VLPs induced high levels of dengue rEIII-specific IgG antibodies and elicited homotypic neutralizing antibodies against DENV-2 NGC strain.Furthermore,spleen cells from mice vaccinated with DENV-2 VLPs and virions produced comparable levels of IFN-γ.Conclusion Monovalent DENV-2 VLPs produced by mammalian cells were capable to induce dengue-specific humoral and cellular immune responses in mice,and this may open a promising approach to the development of tetravalent dengue VLPs vaccine.