GENETICALLY ENGINEERED ORAL VACCINES AND CARIES IMMUNITY

基因工程口服疫苗和龋齿免疫

• 批准号: 6150519
• 负责人: Suzanne M. Michalek
• 金额: $ 24.23万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-02-01 至 2001-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6150519
• 关键词: Peyer's patches; Salmonella; Salmonella vaccines; Streptococcus mutans; bacterial antigens; biotechnology; cholera toxin; dental caries vaccine; drug administration rate /duration; electron microscopy; enzyme linked immunosorbent assay; genetic promoter element; gram negative bacteria; immunofluorescence technique; immunogenetics; immunomodulators; laboratory rat; lymphocyte; molecular cloning; mucosal immunity; nonhuman therapy evaluation; oral administration; recombinant DNA; transfection /expression vector; vaccine development; vector vaccine

Dental caries is an infectious disease in which the major etiologic agents in humans are Streptococcus mutans and Streptococcus sobrinus. Studies aimed at inducing immunity against infectious diseases, including dental caries, have provided valuable information on microbial antigens important in inducing protective responses; on the role of IgA antibodies in defense against infections which invade or colonize surfaces bathed by external secretions; and on the mechanisms involved in inducing immune responses. Concurrent with these studies, the advancements in recombinant DNA technology and gene cloning have led to additional information on microbial gene products important in virulence and to the development of novel vaccines. The overall goal of this project is to define mechanisms by which mucosal vaccines consisting of recombinant, avirulent Salmonella strains expressing cloned genes of mutans streptococci, with and without adjuvant, induce specific immune responses. Specifically, we will: l) Determine the effectiveness of Salmonella expressed cloned B subunit of cholera toxin (CTB) in promoting immune responses to co-expressed cloned antigens of mutans streptococci. The levels and isotype of antibodies to the cloned antigens in serum and external secretions of mice and rats immunized orally with Salmonella expressing chimeric proteins will be measured by ELISA to determine the immunogenic and adjuvant properties of the different types of fusions between antigen and CTB in vivo. Analysis of caries activity in a rat model will assess the protective effect of the responses. These studies will determine which structural type of chimeric protein, when expressed by Salmonella in vivo, is most effective in promoting salivary IgA antibody responses protective against mutans streptococci. 2) Determine the effect of the Salmonella vaccine strain, vector, and promoter on the expression of cloned antigen of mutans streptococci and on the induction of immune responses. The amount of cloned antigen of mutans streptococci produced by Salmonella mutant strains containing expression vectors with different promoters and replicons will be measured by ELISA. Stability of the expression vectors will be assessed by microbiologic detection of Salmonella in tissues of mice orally-immunized with the Salmonella vaccine. The level and isotype of antibodies to the cloned antigen in serum and external secretions of animals orally immunized with a Salmonella strain containing different vectors will be measured to determine if differences exist in the magnitude and duration of the mucosal immune responses and in protection against infection by mutans streptococci. These studies will determine the effect of the vector replicon on the in vivo expression of cloned protein and on the induction of immune responses, and whether immune responses induced by the in vivo expression of cloned protein under the control of a constitutive promoter (trc) versus one induced in an anaerobic environment (nirB) are of greater magnitude and longer duration. 3) Determine the effect of route of immunization with a recombinant Salmonella vaccine on the magnitude and protective capability of the salivary IgA antibody response. The level of IgA antibodies to the cloned antigen in saliva in animals immunized by the oral or intranasal route will be measured by ELISA to determine which route of immunization induces the highest salivary IgA antibody response protective against infection by mutans streptococci. The results of these studies will be relevant in establishing the practicability of Salmonella vaccine delivery systems for the induction of protective immune responses against mucosal pathogens including those associated with the oral cavity.

龋病是一种感染性疾病，其主要病原为 人类感染的是变形链球菌和远缘链球菌。研究 旨在诱导对包括牙科在内的传染病的免疫力 龋病，提供了关于微生物抗原的有价值的信息 在诱导保护性反应中；关于IgA抗体在防御中的作用 防止感染入侵或侵占沐浴在 分泌物；以及诱导免疫反应的机制。 在这些研究的同时，重组DNA的进展 技术和基因克隆带来了更多关于 微生物基因产物对致病力和发展至关重要 新型疫苗。该项目的总体目标是定义机制 由重组无毒沙门氏菌组成的黏膜疫苗 表达变形链球菌克隆化基因的菌株 佐剂，诱导特异性免疫反应。具体来说，我们将：L) 沙门氏菌表达的克隆化B亚基的效力测定 霍乱毒素(CTB)促进共表达克隆的免疫应答 变形链球菌的抗原。日本血吸虫病患者的抗体水平和亚型 小鼠和大鼠血清和外分泌物中的克隆抗原 表达嵌合蛋白的沙门氏菌口服免疫将是 用ELISA法测定其免疫原性和佐剂特性 体内抗原与CTB的不同融合类型。分析 对大鼠龋齿活动的研究将评估黄连素的保护作用 回应。这些研究将确定嵌合体的结构类型 当沙门氏菌在体内表达蛋白质时，对 促进唾液免疫球蛋白A抗体应答对变种的保护作用 链球菌。2)确定沙门氏菌疫苗株的效果， 变形杆菌克隆抗原表达载体和启动子的研究 对链球菌和免疫反应的诱导作用。金额的多少 沙门氏菌突变株产生的变形链球菌抗原的克隆 含有不同启动子和不同启动子表达载体的菌株 复制子将通过酶联免疫吸附试验进行测量。表达载体的稳定性 将通过沙门氏菌在组织中的微生物检测来评估 用沙门氏菌疫苗口服免疫的小鼠。水平和同种类型 血清和体外分泌物中抗克隆抗原抗体的含量 口服免疫沙门氏菌的动物，沙门氏菌含有不同的 将测量向量，以确定在 粘膜免疫反应的大小和持续时间及其保护作用 抗变形链球菌感染。这些研究将确定 载体复制子对克隆蛋白体内表达的影响 以及免疫反应的诱导，以及免疫反应是否 控制下的克隆蛋白在体内表达诱导 构成启动子(TRC)与厌氧条件下诱导的启动子 环境(NIRB)的规模更大，持续时间更长。3) 确定重组疫苗免疫途径的效果 沙门氏菌疫苗对沙门氏菌数量和保护能力的影响 唾液免疫球蛋白A抗体反应。克隆体免疫球蛋白A抗体水平 经口服或鼻腔免疫动物唾液中的抗原 将通过酶联免疫吸附试验进行测量，以确定哪种免疫途径可诱导 唾液中最高的IgA抗体应答，可预防感染 变形链球菌。这些研究的结果将与 建立沙门氏菌疫苗投放系统的可行性 诱导对粘膜病原体的保护性免疫反应 包括与口腔相关的那些。