Identification and functional analysis of non-antigenic factors of intracellular bacteria essential for the induction of cell-mediated protective immunity

本研究探讨了T细胞介导的保护性免疫中细胞内细菌抗原以外的其他因子的作用和机制。主要研究结果如下：1.γ-干扰素-γ（IFN-γ）的内源性产生对于诱导针对细胞内细菌的TH 1依赖性免疫是必需的。在此过程中，巨噬细胞产生的多种细胞因子中，IL-12起重要作用.单核细胞增生李斯特菌LLO是一种非抗原性因子，具有高度诱导细胞因子的能力。通过在体内使用脂质体包封的LLO，可以将内源性细胞因子诱导至与用活细菌免疫（实际感染）诱导的水平相当的水平。将脂质体包裹的LLO作为一种佐剂，LLO与灭活菌或LLO阴性菌株联合免疫小鼠后，均能诱导出较强的保护性免疫力，单独使用这两种菌株均不能诱导保护性免疫. TH-1依赖性保护作用主要是由IFN-γ诱导的巨噬细胞活化所致。活化的巨噬细胞中增强的细菌杀伤的表达是由于活性氧中间体或活性氮中间体，这取决于活化的时间和巨噬细胞水平的细菌感染。申克孢子丝菌（一种可能的细胞内寄生真菌）的感染似乎涉及与上述类似的机制。在LLO诱导IFN-γ的过程中，IL-12和IL-18在LLO刺激巨噬细胞的初始阶段是非常必要的。结果表明，膜上15，000 kDa的蛋白质参与了细胞的初始降解.在LLO分子的4个结构域中，第4结构域是膜损伤表达所必需的。相反，结构域1至3似乎是必不可少的表达的甜菜碱诱导能力。

This study investigated the mechanism and the role of factors other than antigens from intracellular bacteria in the induction of T cell-mediated protective immunity. Results summarized as follows have been obtained.1. The endogenous production of γ-interferon-γ (IFN-γ) was essential for the induction of TH1-dependent immunity against intracellular bacteria. In this process, IL-12 was important among various cytokines produced by macrophages.2. One of the non-antigenic factor, LLO from Listeria monocytogenes, was highly capable of inducing cytokines. By using the liposome-encapsulated LLO in vivo, it was possible to induce the endogenous cytokines to the level comparable to that induced by immunization with viable bacteria (actual infection).3. Application of liposome-encapsulated LLO as a sort of adjuvant, a strong protective immunity could be induced after immunization of mice with LLO plus killed bacteria or LLO-negative strain, both of which are incapable of inducing protection if used alone.4. TH-1 dependent protection was ascribed to the activation of macrophages mainly induced by IFN-γ. The expression of enhanced bacterial killing in activated macrophages was due either to reactive oxygen intermediates or to reactive nitrogen intermediates, depending on the timing of activation and bacterial infection in macrophage level.5. A mechanism similar to the above appeared to be involved in the infection by Sporothrix schenkii, a possible intracellular parasitic fungus.6. In the induction of IFN-γ by LLO, both IL-12 and IL-18 were highly essential at the initial stage of macrophage stimulation by LLO. It was suggested that 15,000 kDa protein of the membrane was involved in the initial triggering.7. Among 4 domains of LLO molecule, 4th domain was indispensable for the expression of membrane damage. In contrast, domains 1 through 3 seemed to be essential for the expression of cytokine-inducing ability.