两性葡聚糖免疫应答与免疫调节机制研究

发布时间：2018-04-02 20:40

本文选题：中性糖　切入点：两性葡聚糖　出处：《福州大学》2006年硕士论文

【摘要】： 多糖属于作为T细胞非依赖性抗原,主要通过增强机体的非特异性免疫功能以抵御细菌、病毒和寄生虫等病原微生物的侵袭,其没有免疫记忆效应,无法直接诱导机体产生高效价特异性IgG抗体,因而在疫苗的制备上通常需将多糖连接于一定得载体上形成偶联物,刺激机体产生高亲和力IgG抗体。而两性葡聚糖与一般多糖不同,它能与抗原递呈细胞进而激活T细胞的免疫调节机制,之后诱导机体产生IgG亚类抗体。本课题着重讨论了两性葡聚糖体液免疫应答和调节效应的特异性及其产生机制。分别以中性糖、两性葡聚糖、糖蛋白为抗原,比较它们在抗体水平和细胞水平上的体液免疫应答和免疫调节效应,从而探寻两性葡聚糖在免疫行为上的特性。本课题采用硫酸酯化法修饰葡糖胺制备带有阴性基团和阳性基团的两性葡聚糖,采用双环氧试剂(1,4-二丁醇二缩水甘油醚)交联蛋白载体制备糖蛋白。相关检测结果显示,两性葡聚糖硫酸根浓度为36. 14 ug/ml,取代度为0.90,而氨基氮含量为35.04ug/ml,取代度为0.91,制得带有正、负电荷的基团的两性葡聚糖。糖蛋白交联产物电泳及HPLC检测结果表明,经双环氧试剂交联以及分离纯化等步骤获得了纯度较高的糖蛋白分子。采用中性葡聚糖(分子量分别为2万,27万)、两性葡聚糖、糖蛋白三种抗原体外刺激的细胞增殖以及细胞因子表达的结果表明,两性葡聚糖能显著促进不同糖免疫后的细胞增殖,并刺激细胞表达TLR4、TLR9、TNFα等细胞因子。中性糖、糖蛋白、两性葡聚糖刺激机体形成抗体生成细胞的实验结果显示,两性葡聚糖除了能刺激细胞表达IgM亚类抗体外还能刺激其表达IgG亚类抗体,而中性多糖则只能表达IgM亚类抗体。在抗体水平上通过ELISA检测中性糖、两性葡聚糖、糖蛋白抗体效价以及相对亲和常数,结果显示两性葡聚糖与糖蛋白有相似的免疫应答效应,都能刺激机体表达糖特异性的IgG类抗体,且两性葡聚糖抗原与其他糖刺激的抗体表现出较强的交叉反应。实验结果说明两性葡聚糖具有特异的免疫应答和调节效应,为多糖疫苗的设计与开发提供一个新的方向。
[Abstract]:Polysaccharides, as T-cell independent antigens, can resist the invasion of bacteria, viruses, parasites and other pathogenic microorganisms by enhancing the body's non-specific immune function, and it has no immune memory effect.It is impossible to directly induce the production of high titer specific IgG antibodies, so it is usually necessary to attach polysaccharides to a certain carrier to form conjugates in order to stimulate the production of high affinity IgG antibodies.The amphoteric dextran is different from other polysaccharides in that it can activate the immune regulation mechanism of T cells with antigen-presenting cells and then induce the production of IgG subclass antibodies.The purpose of this paper is to discuss the specificity and mechanism of humoral immune response and regulation of amphoteric dextran.Neutral sugar, amphoteric dextran and glycoprotein were used as antigens to compare their humoral immune responses and immunomodulatory effects at the antibody level and cell level, respectively, in order to explore the immunological behavior of amphoteric dextran.In this study, glucosamine was modified by sulfate to prepare amphoteric dextran with negative and positive groups.Correlation analysis showed that the concentration of amphoteric dextran sulfate was 36. 5%.The degree of substitution is 0.90, while the content of amino nitrogen is 35.04 渭 / ml, and the degree of substitution is 0.91.Amphoteric dextran with positive and negative charge groups is prepared.The results of electrophoretic analysis and HPLC analysis showed that high purity glycoprotein molecules were obtained by double epoxide crosslinking and separation and purification.The results of cell proliferation and cytokine expression stimulated by neutral dextran (molecular weight 20, 270000), amphoteric dextran and glycoprotein in vitro showed that amphoteric dextran could significantly promote cell proliferation after different glucose immunization.TLR4, TLR9, TNF 伪 and other cytokines were also stimulated.Neutral sugar, glycoprotein and amphoteric dextran stimulated the formation of antibody producing cells. The results showed that amphoteric dextran not only stimulated the expression of IgM subclass antibody, but also stimulated the expression of IgG subclass antibody.Neutral polysaccharides can only express IgM subclass antibodies.The antibody titers of neutral sugar, amphoteric dextran, glycoprotein antibody and relative affinity constant were detected by ELISA at the antibody level. The results showed that amphoteric dextran and glycoprotein had similar immune response effects.Amphoteric dextran antigen showed strong cross reaction with other sugar stimulated antibodies.The results show that amphoteric dextran has a specific immune response and regulatory effect, which provides a new direction for the design and development of polysaccharide vaccine.
【学位授予单位】：福州大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392.1

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