天然抗体IgM 3B4体外抑制白念珠菌与宿主细胞相互作用的实验研究

发布时间：2018-04-02 12:26

本文选题：天然抗体　切入点：白念珠菌　出处：《第四军医大学》2008年硕士论文

【摘要】： 天然抗体(natural antibodies, NAbs)是指正常机体在没有任何抗原刺激的情况下由胚系基因编码的,正常机体产生的针对一种或多种自身和(或)外源性抗原的抗体,具有多反应性和低亲和力的特征。自NAbs被发现以来,人们对其可能具有的功能进行了一系列的深入研究。研究结果表明,NAbs是构成机体天然免疫系统的重要组成部分,在肿瘤监视、自身免疫、异体排斥、抗感染等方面均发挥着重要的作用。近年来,随着抗生素、皮质类固醇激素、免疫抑制剂等的广泛应用,以及HIV感染者增加、异种移植术的开展、肿瘤发病率的上升,真菌感染呈明显上升趋势,特别是白念珠菌引起的感染明显上升,越来越引起人们的重视。人们逐渐认识到体液免疫在抗真菌感染中起着重要作用,而天然体液免疫又构成了机体抵御外来感染的第一道防线,天然抗体的作用逐渐引起人们的重视。虽然有部分事实表明天然抗体可能具有抗真菌免疫的作用,但还缺乏直接的证据,为此倍受医学界学者的广泛关注。本课题组在国家自然科学基金(30330510、30300318, 30330550, 30425015)的支持下,应用未免疫小鼠B细胞与骨髓瘤细胞融合,成功筛选到了3株分泌IgM型天然抗体的杂交瘤细胞,它们的可变区基因与胚系基因高度同源,其中以天然抗体IgM 3B4与抗原的结合活性最好,并且已经证实在体外天然抗体IgM 3B4可以调理吞噬细胞对金黄色葡萄球菌的吞噬作用和天然抗体IgM 3B4可以与白念珠菌孢壁的糖蛋白抗原结合。本研究在前期工作的基础上利用纯化得到的天然抗体IgM 3B4,在体外以白念珠菌为对象,初步探讨天然抗体IgM 3B4抗真菌感染的作用机制。本研究利用纯化得到的天然抗体IgM 3B4在体外证实:1在体外,不同浓度的天然抗体IgM 3B4可以抑制白念珠菌芽管生成,2在体外,天然抗体IgM 3B4可以抑制白念珠菌与宿主细胞之间的粘附;3采用ELISA和RT-PCR方法检测了天然抗体IgM 3B4在体外对感染了白念珠菌的脐静脉血管内皮细胞IL-6、IL-8、ICAM-1和VCAM-1表达和分泌的影响,证实天然抗体IgM 3B4明显促进了脐静脉血管内皮细胞上述细胞因子的表达和分泌。从而证实了天然抗体IgM 3B4抵抗白念珠菌感染的能力与其抑制白念珠菌芽管生成、抑制白念珠菌与宿主细胞之间的粘附以及促进脐静脉血管内皮细胞IL-6、IL-8、ICAM-1和VCAM-1的表达和分泌有关。
[Abstract]:Natural antigens (NAbs) are antibodies that are encoded by embryogenic genes in normal organisms without any antigenic stimulation, and are produced against one or more of their own and / or exogenous antigens. Since the discovery of NAbs, a series of in-depth studies have been carried out on its possible functions. The results show that NAbs is an important part of the innate immune system. It plays an important role in tumor surveillance, autoimmunity, allograft rejection and anti-infection. In recent years, with the widespread use of antibiotics, corticosteroids, immunosuppressants, and the increase of HIV infection, xenotransplantation, the incidence of tumor and fungal infection have increased significantly. In particular, the infection caused by Candida albicans has increased significantly and attracted more and more attention. People have come to realize that humoral immunity plays an important role in antifungal infections. And natural humoral immunity constitutes the first line of defense against foreign infections, and the role of natural antibodies has gradually attracted people's attention. Although some facts have shown that natural antibodies may have the effect of antifungal immunity, However, the lack of direct evidence has attracted wide attention from medical scholars. With the support of the National Natural Science Foundation of China, 30330510C30300318,30330550,30425015, three hybridoma cells secreting IgM type natural antibodies were successfully screened by fusion of unimmunized B cells with myeloma cells. Their variable region genes are highly homologous to embryogenic genes, and the natural antibody IgM 3B4 has the best binding activity to antigens. It has been proved that natural antibody IgM 3B4 can regulate phagocytosis of phagocytes to Staphylococcus aureus in vitro and that natural antibody IgM 3B4 can bind to glycoprotein antigen of Candida albicans spore wall. Based on the purified natural antibody IgM 3B4, Candida albicans was used in vitro. To explore the mechanism of natural antibody IgM 3B4 in antifungal infection. In this study, the purified natural antibody IgM 3B4 was used to confirm that IgM 3B4 of different concentrations could inhibit the formation of Candida albicans bud tube in vitro. Natural antibody IgM 3B4 could inhibit adhesion between Candida albicans and host cells. The effect of natural antibody IgM 3B4 on the expression and secretion of IL-8 ICAM-1 and VCAM-1 in umbilical vein endothelial cells infected with Candida albicans was detected by ELISA and RT-PCR in vitro. It was proved that the natural antibody IgM 3B4 significantly promoted the expression and secretion of the above cytokines in the endothelial cells of the umbilical vein, thus confirming the ability of the natural antibody IgM 3B4 to resist the infection of Candida albicans and to inhibit the formation of the germ tubes of Candida albicans. The inhibition of the adhesion between Candida albicans and host cells and the promotion of the expression and secretion of ICAM-1 and VCAM-1 in endothelial cells of umbilical vein (HUVEC) were related to the inhibition of the adhesion between Candida albicans and host cells.
【学位授予单位】：第四军医大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392

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