基于生殖支原体黏附蛋白模拟表位的多抗原肽免疫效果的观察  

Observation on the immunological competence of multiple antigen peptides based on the mimic epitope of the adhesion protein of Mycoplasma genitalium

作　　者：曾焱华[1] 游晓星[1] 唐双阳[1] 朱翠明[1] 余敏君[1] 吴移谋[1] 何军[2] 

机构地区：[1]南华大学病原生物学研究所,衡阳421001 [2]南华大学附属南华医院

出　　处：《中华微生物学和免疫学杂志》2013年第4期287-292,共6页Chinese Journal of Microbiology and Immunology

基　　金：国家自然科学基金项目（30801064）;湖南省高校科技创新团队（湘教通2010-212号）;湖南省研究生科研创新项目（CX20108381）

摘　　要：目的制备基于生殖支原体黏附蛋白（MgPa）模拟表位的多抗原肽（MAP），检测其诱导小鼠产生特异性体液免疫和细胞免疫应答的水平，为研制安全、有效的基于模拟表位的Mg表位肽疫苗奠定实验基础。方法以多聚赖氨酸为核心基质，人工合成3种含MgPa模拟表位的八分枝MAP，用反相高效液相色谱（RP—HPLC）和质谱分析对其进行纯化与鉴定。将其单独或混合免疫BALB／c小鼠。间接ELISA测定小鼠血清中的特异性IgG抗体及其亚类的水平；MTT比色法检测MAP刺激免疫小鼠脾淋巴细胞的增殖反应，并用ELISA检测其脾细胞培养上清中的IL-4和WN-γ水平。结果成功合成了较高纯度的含MgPa模拟表位的3种MAP，这3种MAP均能刺激小鼠产生IgG抗体，且以Thl型免疫应答产生的IgG2a型抗体为主；与单个MAP组相比，混合MAP免疫组小鼠产生了更多的IgG、IgGl和IgG2a抗体（P均〈0．01）；3种MAP均能特异性诱导免疫小鼠的脾淋巴细胞增殖，且能刺激其产生较多的IL-4和IFN-1，混合MAP免疫组小鼠产生的IL4和IFN-γ，与单个MAP组相比差异具有统计学意义（P均〈0．01）。结论基于MgPa模拟表位的3种MAP均能刺激小鼠产生较强的体液免疫和细胞免疫应答，且混合免疫组诱导的免疫应答效果比单个MAP组更好。Objective To provide experimental evidence for the development of multi-epitope-baeded marker vaccines through investigating the humoral and cellular immune responses in BALB/c mice induced by the multiple antigen peptides （MAPs） with the mimic epitope. Methods Three types of MAPs in eight branched forms containing the mimic epitope of Mycoplasma genitalium adhesion protein （MgPa） were prepared using poly-lysine as the core matrix. The purity of MAPs was analyzed by reverse phase high performance liquid chromatography （RP-HPLC）. The molecular weights of MAPs were characterized by Mass Spectrometry. The BALB/c mice were immunized intramuscularly for four times with single or mixed MAPs. The specific IgG antibody and the subtype of IgG antibody in serum of the immunized mice were detected by indirect ELISA. The proliferative responses of the spleen lymphocytes were detected using M＇Fr assay. The ELISA were used to detect IFN-γ and IL-4 levels in the cultured supernatant of spleen lymphocytes. Results The three types of MAPs containing the mimic epitopes were successfully prepared with high purity. They, could stimulate mice to produce specific IgG antibodies, of which, the major antibody isotype was Thl immune response-associated IgG2a. Compared with the single MAP immunization group, the mixed-MAPs immunized mice produced more IgG, IgG1 and IgG2a antibody （P〈0.05）. Furthermore, these MAPs could enhance the specific proliferation of spleen lymphocytes in immunized mice and induce the production of IFN-γ and IL-4. The levels of IFN-γ and IL-4 in mixed-MAPs group were significantly higher than those of the single MAPs group （P〈0.01）. Conclusion The three types of MAPs could induce strong specific cellular and humoral immune responses. The immunological competence of the mixed-MAPs was stronger than those of the single MAP.

关 键 词：生殖支原体 黏附蛋白 模拟表位 多抗原肽 免疫原性 

分 类 号：R3[医药卫生—基础医学]