摘要
目的构建HIV-1C亚型gp120负载人树突状细胞(dentriti ccell,DC)疫苗,并对其体外功能进行初步检测。方法利用Amaxa细胞核转染技术将pcDNA3.1-gp120质粒转染至人成熟DC,以Western blot检测gp120的表达。通过流式细胞仪检测DC表面共刺激分子的变化、混合淋巴细胞反应、CD8+T细胞表面活化分子CD25的表达及其分泌IFN-γ的变化。结果通过Western blot检测,gp120在DC中得到了正确表达。经流式细胞仪检测,DC表面分子CD80表达率由刺激前的33.34%上升至43.20%,CD86表达率由刺激前的60.08%上升至90.34%;负载gp120DC刺激淋巴细胞增殖率为86.72%;CD8+T细胞表面分子CD25表达率由刺激前的5.27%上升至74.21%,IFN-γ的表达率达37%。结论负载了HIV-1gp120的人树突状细胞能够显著刺激淋巴细胞的增殖、增强CD8+T细胞表面活化分子CD25表达以及促进CD8+T细胞分泌IFN-γ,为下一步DC治疗性疫苗的体内研究奠定基础。
This study is aimed to construct a therapeutic vaccine containing HIV-1 subtype C gp120 and test the function in vitro. Mature DCs were transfected by recombined expression plasmid pcDNA3.1-gp120. Then the expressions of gp120 were identified by Western blot; flow cytometry was applied to analyze the expression levels of costimulatory molecules CD80 and CD86,the mixed lymphocyte reaction,the CD25 production and IFN-γ secretion. We found that gp120 was correctly expressed in DCs; CD80 and CD86 increased from 33.34% to 43.20% and 60.08% to 90.34%,respectively; T cell proliferation rate was 86.72%. The expression rate of CD25 increased from 5.27% to 74.21%. IFN-γ secreted by CD8+ T cells reached a high level of 37%. We conclude that T cell proliferation is promoted by gp120-loaded DCs,and The CD25 production and IFN-γ secretion are significantly up-regulated,which provides a base for studying the function of the vaccine in vivo.
出处
《免疫学杂志》
CAS
CSCD
北大核心
2010年第9期741-746,共6页
Immunological Journal
基金
国家自然科学基金(30471605)
中国科学院知识创新工程重要方向(KSCX1-YW-R-15,KSCX2-YW-R-092)
云南省科技基础条件平台计划(2006PT08)