黄芪多糖作为佐剂对乙型肝炎疫苗免疫反应影响的研究
发布时间：2018-05-16 07:01
本文选题：黄芪多糖 + 乙型肝炎 ； 参考：《四川农业大学》2010年硕士论文
【摘要】： 乙型肝炎是由乙型肝炎病毒(hepatitis B virus, HBV)引起的一种影响人类健康的世界性传染病。接种乙肝疫苗是目前控制乙型肝炎感染的最有效的措施,但是传统的乙型肝炎亚单位疫苗是由乙型肝炎病毒的表面抗原和铝佐剂组成,铝佐剂疫苗仅能引起很好的体液免疫反应而不能引起有效的细胞免疫反应。 佐剂可以增强细胞免疫和体液免疫,活化T细胞,上调细胞因子的表达,增强免疫记忆,并能改变免疫反应类型。因此改良佐剂可能增强传统乙型肝炎亚单位疫苗的免疫反应。黄芪多糖(Astragalus polysaccharides,APS)是中药黄芪的提取物,具有广泛的药理作用,也具有很强的免疫调节作用。APS可从多个方面发挥免疫调节作用：直接影响免疫细胞内的物质代谢,诱导产生相关的免疫调节因子；不仅能够增强机体的特异性免疫反应,还可以增强非特异性免疫反应；既可增强正常机体的免疫功能,又可调节异常机体的免疫功能。 本研究将APS作为乙型肝炎亚单位疫苗的佐剂与疫苗一起免疫昆明白小鼠,研究其对乙肝疫苗免疫反应的影响,尤其是对细胞免疫反应的影响。实验选取健康雌性6-8周龄昆明白小鼠35只,体重20-24 g,随机分为5组,每组7只,分别为rHBsAg组、rHBsAg+APS组、rHBsAg+aulm组、APS组和对照组(不进行免疫)。初次免疫设为0天,第14天加强免疫一次,在第21天进行免疫学指标的检测。主要结果如下： 实验使用定量ELISA法检测IgG水平,用MTT法检测T细胞增殖反应,用流式细胞仪检测体内CTL反应,结果发现APS显著增强乙肝疫苗诱导产生较高的IgG抗体水平,较强的T淋巴细胞增殖反应和体内CTL杀伤反应,表明APS能够增强乙肝疫苗的免疫反应。 为进一步研究APS作为乙型肝炎亚单位疫苗的佐剂对免疫反应的影响,对有关免疫学指标作了进一步检测。用RT-PCR法检测PFP、GraB、Fas L和Fas的表达以及用流式细胞仪检测体内细胞因子的表达水平,结果表明APS能够增强乙肝疫苗上调PFP、GraB、Fas L、Fas和CD8+T细胞IFN-γ的表达水平,与体内CTL反应结果一致；APS能够增强乙肝疫苗诱导较高的CD4+T细胞IFN-γ、IL-2和IL-4的表达水平,表明APS作为乙肝疫苗的佐剂能够促进机体Thl型和Th2型免疫反应。 为进一步探讨APS作为佐剂增强免疫反应的作用机理,用半定量PT-PCR法对TLR2、TLR4、TGF-β1和Foxp3的表达水平进行检测,结果表明APS能够很好地增强乙肝疫苗诱导TLR2和TLR4表达,通过上调TLR2和TLR4的表达激发协同刺激分子和相关细胞因子的表达参与适应性免疫应答；APS作为乙肝疫苗的佐剂能够明显降低机体TGF-β1和Foxp3表达水平,通过抑制TGF-β1和Foxp3的表达促进机体免疫反应。 以上结果表明,APS作为乙型肝炎亚单位疫苗的佐剂能增强乙肝疫苗的免疫反应。本研究初步证实了APS是一种良好的免疫佐剂,能够显著地提高乙型肝炎亚单位疫苗的体液免疫水平和细胞免疫水平,为乙型肝炎疫苗的研发提供了有意义的基础研究数据资料。
[Abstract]:Hepatitis B is a worldwide infectious disease affecting human health caused by hepatitis B virus (HBV). Vaccination of hepatitis B is the most effective measure to control hepatitis B infection. However, the traditional hepatitis B subunit vaccine is composed of the surface antigen of hepatitis B virus and aluminum adjuvant, aluminum adjuvant. The vaccine can only cause good humoral immune response without inducing effective cellular immune response.
The adjuvant can enhance cell immunity and humoral immunity, activate T cells, increase the expression of cytokines, enhance the immune memory, and change the type of immune response. Therefore, the modified adjuvant may enhance the immune response of the traditional hepatitis B subunit vaccine. The Astragalus Polysaccharide (Astragalus polysaccharides, APS) is the extract of Astragalus membranaceus. Extensive pharmacological action and strong immunoregulation effect.APS can play an immunomodulatory role in many aspects: directly affecting the metabolism of substances in the immune cells and inducing related immunoregulatory factors; not only can enhance the specific immune response of the body, but also enhance the non specific immune response; it can enhance the normal function. The immune function of the body can also regulate the immune function of the abnormal body.
In this study, we immunized Kunming white mice with the adjuvant of APS as a subunit vaccine of hepatitis B, and studied the effect of the vaccine on the immune response of the hepatitis B vaccine, especially on the immune response of the hepatitis B vaccine. The experiment selected 35 healthy female 6-8 weeks old mice and weighed 20-24 G, randomly divided into 5 groups, each group of 7, respectively, group rHBsAg, rHBs Group Ag+APS, group rHBsAg+aulm, group APS and control group (no immunization). The first immunization was set for 0 days, the immunization was strengthened on the fourteenth day, and the immunological index was tested on the twenty-first day. The main results were as follows:
The experiment used quantitative ELISA to detect the IgG level, detect the proliferation reaction of T cells by MTT, and detect the CTL reaction in vivo by flow cytometry. The results showed that APS significantly enhanced the level of IgG antibody induced by hepatitis B vaccine, strong T lymphocyte proliferation reaction and in vivo CTL killing reaction, indicating that APS can enhance the immune reaction of hepatitis B vaccine. It should.
In order to further study the effect of APS as a adjuvant on hepatitis B subunit vaccine, the immune response was further examined. The expression of PFP, GraB, Fas L and Fas and the expression of cytokines in the body were detected by RT-PCR and the expression of cytokines in the body was detected by flow cytometry. The results showed that APS enhanced the hepatitis B vaccine to increase PFP and GraB, The expression level of IFN- gamma in Fas L, Fas and CD8+T cells is consistent with the results of CTL reaction in the body. APS can enhance the expression level of IFN- gamma, IL-2 and IL-4 of the hepatitis B vaccine, which indicates that the adjuvant of APS as a hepatitis B vaccine can promote the body type and the immune response of CD4+T.
In order to further explore the mechanism of APS as an adjuvant to enhance immune response, the expression level of TLR2, TLR4, TGF- beta 1 and Foxp3 was detected by semi quantitative PT-PCR method. The results showed that APS could enhance the expression of TLR2 and TLR4 induced by hepatitis B vaccine, and up up the table of TLR2 and TLR4 up to stimulate the table of synergistic stimulants and related cytokines. As an adjuvant of hepatitis B vaccine, APS can significantly reduce the expression level of TGF- beta 1 and Foxp3, and promote the immune response by inhibiting the expression of TGF- beta 1 and Foxp3.
The above results show that APS as a adjuvant of hepatitis B subunit vaccine can enhance the immune response of hepatitis B vaccine. This study preliminarily confirmed that APS is a good immune adjuvant. It can significantly improve the level of humoral immunity and cellular immunity of HBV subunit vaccine, which provides significance for the research and development of hepatitis B vaccine. Basic research data.
【学位授予单位】：四川农业大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R392
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