| Tuberculosis is a most serious disease of humans in the world. It is estimated that one third of the world’s population have been infected with Mycobacterium tuberculosis(M.tb). Although most of infection is asymptomatic, approximately8.7million new cases in the world occur annually. M.tb could disseminate through and survive for a long time in the aerosol. It’s an intracellular pathogen that can be phagocytosed by antigen presenting cells(APC), such as macrophages and dendritic cells (DCs). However, this bacterium can evade immune response and survive in APC by inhibition of antigen presentation. Furthermore, the bacteria will spread to other places in the body with APC migration and thereby lead to and pulmonary or extra-pulmonary tuberculosis. The molecular mechanism that M.tb causes persistence and immune inhibition is largely unclear.Mycobacterium bovis(M.bovis) is the main pathogen for bovine tuberculosis, and shares>99.95%identity to M.tb at the genome level. Besides, it can also infect humans and a wide variety of wildlife animals. It has become more difficult to control and prevent bovine tuberculosis once the wildlife animals become the carrier and source of this patheogen. M.tb and M.bovis share similar immune response and pathogenesis in vivo.M.bovis bacillus Calmette-Gue’rin (BCG) is an attenuated strain of virulent M. bovis which lost its virulence after230passages in vitro and widely used as a unique vaccine to protect children against tuberculosis.DCs are professional APCs and work as the bridge linking innate immunity to acquired immunity of the hosts. Therefore, we used M.bovis and the isogenic attenuated strain BCG as the reference strains and DCs as the model cells to research the different immune response in DCs after infection, and thereafter the differntiaton of CD4+T cells driven by infected DCs. The results were expected to explain the molecular mechanism of immune response evasion and persistent infection. The main findings are as follows:1. We analysed cytokines and chemokines secretion by DCs infected with M.bovis and BCG. As a result, BCG induced significantly more concentrations of IL-12,TNF-a,RANTES and MCP-1;but M.bovis promoted secretion of IL-1β and IL-23. It suggests that BCG induced more Thl type cytokines and chemokines, in favor of development of a protective immunity; while M.bovis induced more inflammation cytokines, and would be responsible for inflammatory damage.2. Detection of the cytokines and chemokines after DCs infected with live or heat killed BCG/M.bovis revealed that live BCG/M.bovis induced more concentrations of cytokines and chemokines than the heat killed BCG/M.bovis. These findings demonstrated that to be alive for BCG/M.bovis are necessary to induce an effect DCs response.3. The NF-κB inhibitor PDTC could inhibit NF-κB signal activation in DCs after infected with BCG/M.bovis. This suggests that NF-κB regulated the secretion of cytokines and chemokines in DCs after mycobacterium infection.4. The different parts of DCs culture after infection with BCG/M.bovis including infected DCs alone, the supernatant of infected DCs culture, the whole culture(infected DCs+supernatant with soluble cytokines), the live and heat killed BCG/M.bovis, were incubated with naive CD4+T cells, and the cytokines and the proportions of CD4+T subsets were detected. As a result, DCs culture infected with BCG could induce most production of IFN-y suggesting the development of Th1subset. However DCs culture infected with M.bovis induced highest levels of IL-17indicating the development of Th17subset which may relate to inflammation damage. In addition, DCs culture infected with M.bovis induced most levels of IL-10and highest proportion of CD25+FoxP3+CD4+T cells detected by flow cytometry, a typical phenotype of Treg cells which would be responsible for suppression of immune response. In a word, the results of this study might be important in explaining the molecular mechanism of M.bovis persistent infection and immune response suppression in vivo. |
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