不同疫苗佐剂对OVA诱生CTL免疫应答的影响

发布时间：2018-02-14 15:44

本文关键词： 佐剂 CpG-ODN Th1型免疫应答细胞毒性T淋巴细胞细胞免疫　出处：《西北农林科技大学》2011年硕士论文　论文类型：学位论文

【摘要】：CD8~+ CTL反应在肿瘤和慢性感染等疾病的治疗中具有重要作用,然而在研究中多使用DNA疫苗和载体活疫苗等通过内源性抗原加工提呈途径诱生细胞免疫,往往具有安全性差、免疫效果不理想和抗载体效应等缺点。蛋白和多肽等外源性抗原能够通过交叉提呈和交叉致敏诱生CD8~+ CTL反应,但需要有效的疫苗佐剂来增强抗原提呈和共刺激信号并提供Th1类细胞因子环境。本论文初步评价了新型疫苗佐剂CpG-ODN与Al(OH)3或Montanide ISA 720等组成的复合佐剂在小鼠体内促进蛋白抗原通过交叉提呈和交叉致敏诱生CD8~+ CTL反应的能力,从而为设计基于蛋白和多肽抗原的新型治疗性疫苗提供理论基础。在本研究中,首先以鸡卵清蛋白(OVA)为模型抗原,分别以CpG-ODN、Al(OH)3、Montanide ISA 720、CpG-ODN + Al(OH)3和CpG-ODN + Montanide ISA 720为疫苗佐剂,肌肉注射免疫C57BL/6小鼠,以胞内细胞因子染色和体内CTL杀伤等方法评价细胞免疫效果。在此基础上,进一步通过ELISPOT、胞内细胞因子染色、体内CTL杀伤和李斯特菌攻击等方法评价了A、B和C三种具有不同化学结构和生物学活性的CpG-ODN与Al(OH)3组成的复合佐剂对OVA抗原的细胞免疫佐剂效应。两针免疫后结果表明,与无佐剂对照组相比,Al(OH)3本身并不能有效诱生细胞免疫,CpG-ODN或Montanide ISA 720单独使用能够在一定程度上增强抗原特异性CD8~+ T细胞的IFN-γ分泌和CTL活性,但不增强抗原特异性CD4~+ T细胞反应。两种复合佐剂具有比单佐剂更强的细胞免疫佐剂效应,其中CpG-ODN + Montanide ISA 720只能增强抗原特异性CD4~+和CD8~+ T细胞的IFN-γ分泌,而CpG-ODN + Al(OH)3不但能够增强抗原特异性CD4~+和CD8~+ T细胞的IFN-γ反应,还能够增强CD8~+ CTL反应。三种不同类型CpG-ODN的比较研究结果表明,B和C型具有相近的细胞免疫佐剂效应,不但能够促进抗原特异性CD4~+和CD8~+ T细胞的IFN-γ分泌,而且能够诱生抗原特异性CD8~+ CTL反应。与此不同,尽管A型CpG-ODN能够在一定程度上促进抗原特异性CD8~+ CTL反应,但并不增强抗原特异性CD4~+和CD8~+ T细胞的IFN-γ分泌。与此相一致,B和C型CpG-ODN在小鼠李斯特菌攻击试验中也显示了比A型CpG-ODN更强的抗原特异性免疫保护作用。综上所述,在三种类型CpG-ODN中,B或C型CpG-ODN与Al(OH)3组成的复合佐剂在小鼠体内具有更强的促进蛋白抗原通过交叉提呈和交叉致敏诱生CD8~+ CTL反应的能力。
[Abstract]:CD8 ~ CTL reaction plays an important role in the treatment of diseases such as tumor and chronic infection. However, the use of DNA vaccine and vector live vaccine to induce cellular immunity through endogenous antigen processing is often of poor safety. The immune effect is not ideal and the anti-carrier effect is not good. Exogenous antigens such as protein and polypeptide can induce CD8 ~ CTL reaction by cross-presentation and cross-sensitization. But effective vaccine adjuvant is needed to enhance antigen presentation and costimulatory signal and to provide Th1 cytokine environment. In this paper, we preliminarily evaluate the effect of a new vaccine adjuvant, CpG-ODN and Al(OH)3 or Montanide ISA 720, in mice. The ability of protein entry antigen to induce CD8 ~ CTL reaction by cross presentation and cross sensitization, This provides a theoretical basis for the design of novel therapeutic vaccines based on protein and polypeptide antigens. In this study, chicken ovalbumin (ovalbumin) was first used as model antigen. CpG-ODN ISA 720CpG-ODN Al(OH)3 and CpG-ODN Montanide ISA 720 were used as vaccine adjuvants to immunize C57BL / 6 mice intramuscularly. The cellular immune effect was evaluated by intracellular cytokine staining and CTL killing in vivo. In vivo CTL killing and listeria attack methods were used to evaluate the cellular immune adjuvant effect of three kinds of complex adjuvants of CpG-ODN and Al(OH)3 with different chemical structure and biological activity on OVA antigen. The results of two-dose immunization showed that the CpG-ODN or Montanide ISA 720 alone could not effectively induce CpG-ODN or Montanide ISA 720 to enhance the IFN- 纬 secretion and CTL activity of antigen-specific CD8T cells compared with the control group without adjuvant. But the antigen-specific CD4T cell response was not enhanced. The two kinds of complex adjuvants had stronger cellular immune adjuvant effect than single adjuvant, and CpG-ODN Montanide ISA 720 could only enhance IFN- 纬 secretion of antigen-specific CD4- and CD8T cells. CpG-ODN Al(OH)3 could not only enhance IFN- 纬 reaction of antigen-specific CD4 ~ and CD8T cells, but also enhance CD8 ~ CTL reaction. It can not only promote IFN- 纬 secretion of antigen-specific CD4- and CD8T cells, but also induce antigen-specific CD8- CTL reaction, although type A CpG-ODN can promote antigen-specific CD8- CTL reaction to some extent. However, the IFN- 纬 secretion of antigen-specific CD4- and CD8T cells was not enhanced. In accordance with this, B and C CpG-ODN also showed stronger antigen-specific immune protection than type A CpG-ODN in Listeria mutans attack test. In conclusion, the complex adjuvant composed of B or C type CpG-ODN and Al(OH)3 in three types of CpG-ODN has a stronger ability to induce CD8 ~ CTL reaction by cross-presentation and cross-sensitization of protein antigen in mice.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2011
【分类号】：R392

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