表达HBsAg的重组病毒联合免疫和pVR-IL15基因佐剂的免疫效果研究

发布时间：2019-05-27 13:01

【摘要】：乙型肝炎是一个严重威胁人类健康的全球性公共卫生问题。目前,乙肝的防治工作主要分为预防和治疗两个部分,乙肝预防性疫苗已被证实是高免疫原性的,具有保护性的,能够有效的预防乙肝病毒感染。乙肝治疗的最终目的是防止肝纤维化、肝功能衰竭和肝癌的进展,围绕此目的,产生了一些抗乙肝病毒的药物,但这些药物都无法解决所有乙肝患者的肝脏损伤问题,并且耐药性也是目前乙肝防治工作中面临的困难之一。所以亟待发展一种能够彻底治愈乙肝的方法,而治疗性疫苗是目前最具有发展潜力的治疗手段之一。为提高乙肝治疗性疫苗的免疫效力,研究人员进行了大量疫苗构建方法和不同免疫策略的尝试。使用具有免疫增强作用的细胞因子作为疫苗佐剂便是其中一种策略[1]。已有一些研究结果证实IL15能够增强抗原特异性细胞免疫反应、特异性CTL反应以及增加记忆性T淋巴细胞的存活[2]。本研究选取表达乙肝病毒HBsAg基因的重组质粒pVR-S、重组腺病毒rAdv-S和重组痘病毒rMVA-S,组合成不同免疫策略,辅以pVR-IL15基因佐剂免疫,研究其对不同免疫策略诱导的免疫应答水平的影响。实验结果显示,在体液免疫应答中,pVR-IL15基因佐剂能够增强本研究中除了单独免疫重组病毒策略以外的所有免疫策略诱导的体液免疫应答。在单独免疫重组病毒、一次免疫pVR-S和一次免疫重组病毒、两次免疫pVR-S和一次免疫重组病毒的策略中,均显现出rAdv-S诱导的体液免疫应答水平高于rMVA-S,此三种免疫策略诱导的体液免疫效果由强到弱的顺序为两次免疫pVR-S和一次免疫重组病毒策略、一次免疫pVR-S和一次免疫重组病毒策略、单独免疫重组病毒策略。免疫两次重组病毒策略中显示免疫两次rMVA-S诱导的体液免疫应答水平低于交叉免疫rAdv-S和rMVA-S诱导的免疫应答水平。细胞免疫应答检测结果显示,pVR-IL15基因佐剂均能增强实验中所有免疫策略诱导的细胞免疫应答水平。免疫两次重组病毒策略的结果显示,免疫同源载体病毒和异源载体病毒所诱导的细胞免疫水平无差异;一次免疫pVR-S和一次免疫重组病毒策略和免疫两次重组病毒策略所诱导的细胞免疫应答水平无差异;两次免疫pVR-S和两次免疫重组病毒策略所诱导的细胞免疫应答水平在各组免疫策略中效果最好。综上,pVR-IL15基因佐剂能够增强各小鼠免疫方案中重组病毒诱导的体液免疫应答,在细胞免疫应答,pVR-IL15基因佐剂也起到了免疫增强的作用,在免疫两次重组病毒方案中,pVR-IL15基因佐剂最高可将免疫应答水平提高3.6倍。在体液免疫中rAdv-S诱导的免疫效果强于rMVA-S;免疫两次rMVA-S诱导的体液免疫应答水平低于交叉免疫rAdv-S和rMVA-S。在细胞免疫中,免疫同源载体病毒和异源载体病毒所诱导的细胞免疫水平无差异,一次免疫pVR-S和一次免疫重组病毒免疫策略和免疫两次重组病毒策略所诱导的细胞免疫效果相当。无论是体液免疫还是细胞免疫均显示两次重组质粒与两次重组病毒联合免疫所诱导的免疫效果最好。
[Abstract]:Hepatitis B is a global public health problem that is a serious threat to human health. At present, the prevention and treatment of hepatitis B is divided into two parts: prevention and treatment, and the preventive vaccine of hepatitis B has been proved to be highly immunogenic and protective, and can effectively prevent the hepatitis B virus infection. The ultimate goal of the treatment of hepatitis B is to prevent liver fibrosis, liver failure, and progression of liver cancer, around which a number of anti-HBV-resistant drugs are produced, but none of these drugs address the liver injury problems of all hepatitis B patients, And drug resistance is also one of the difficulties in the prevention and treatment of hepatitis B. Therefore, it is urgent to develop a method capable of completely curing hepatitis B, and the therapeutic vaccine is one of the most promising therapeutic methods. In order to improve the immune efficacy of the hepatitis B therapeutic vaccine, the researchers conducted a large number of vaccine construction methods and different immunization strategies. The use of cytokines with immunopotentiation as a vaccine adjuvant is one of the strategies[1]. Some studies have shown that IL15 can enhance antigen-specific cellular immune response, specific CTL responses, and increase the survival of memory T-lymphocytes[2]. The recombinant plasmid pVR-S, the recombinant adenovirus rAdv-S and the recombinant poxvirus rMVA-S, which express the HBsAg gene of the hepatitis B virus, are selected to be combined into different immune strategies, and the recombinant adenovirus rAdv-S and the recombinant poxvirus rMVA-S are combined into different immune strategies, and the pVR-IL15 gene adjuvant is used for immunizing to study the effect of the recombinant adenovirus rMVA-S on the immune response level induced by different immune strategies. The results of the experiments show that in the humoral immune response, the pVR-IL15 gene adjuvant can enhance the humoral immune response induced by all the immune strategies other than the individual immune-recombinant viral strategy in this study. The humoral immune response level induced by rAdv-S is higher than that of rMVA-S in the strategy of single-immune recombinant virus, one-time immune pVR-S and one-time immune recombinant virus, two immunopVR-S and one-time immune recombinant virus. The humoral immune effects induced by the three kinds of immune strategies are two immune pVR-S and one-time immune-recombination virus strategy, one-time immune pVR-S and one-time immune-recombination virus strategy, and the strategy of the recombinant virus is separately immunized. The level of humoral immune response induced by rMVA-S was lower than that of cross-immunized rAdv-S and rMVA-S. The results of cellular immune response indicate that the pVR-IL15 gene adjuvant can enhance the level of immune response induced by all the immune strategies in the experiment. The results of the two-time recombinant virus strategy showed no difference in the cellular immune response induced by the immune-homologous vector virus and the heterologous vector virus, and there was no difference in the level of the cellular immune response induced by the one-time immune pVR-S and one-time immune-recombination virus strategy and the two-time recombinant virus strategy. The level of cellular immune response induced by two immune pVR-S and two immune-recombinant virus strategies is the best in each group of immune strategies. on the whole, the pVR-IL15 gene adjuvant can enhance the humoral immune response induced by the recombinant virus in each mouse immune scheme, The highest immune response level of pVR-IL15 gene adjuvant can be increased by 3.6 times. The immune response induced by rAdv-S in humoral immunity was stronger than that of rMVA-S, and the level of humoral immune response induced by rMVA-S was lower than that of rMVA-S and rMVA-S. In cellular immunity, the cellular immune levels induced by the immune-homologous carrier virus and the heterologous vector virus were not different, and the immune response of the primary immune pVR-S and the one-time immune-recombinant virus and the two-time recombinant viral strategy were comparable. Both humoral and cellular immunity show the best immune response induced by the combination of two recombinant plasmids with two recombinant viruses.
【学位授予单位】：中国疾病预防控制中心
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R512.62

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