Soares, Sandra Clara Chaves; (2001) Interactions between dendritic cells and mycobacterium tuberculosis. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Mycobacterium tuberculosis (Mtb) is an intracellular pathogen that has a major impact on human health. Control of tuberculosis has proved extremely difficult, particularly in developing and underdeveloped countries; this has been exacerbated by the variable efficacy of BCG, the current vaccine, and by the increasing prevalence of drug resistant strains. Effective immunity against Mtb involves cell-mediated mechanisms. Dendritic cells (DC) are likely to play a critical role in the induction of a cellular response to Mtb since they are the most efficient antigen-presenting cells (APC) for priming naive T cell lymphocytes. In this study we have investigated the interaction between Mtb and DC, and how that interaction might be used to generate protective immune responses. Working with a dendritic cell line and using electron microscopy we have shown that coincubation of DC with M.tuberculosis results in the rapid internalisation of the mycobacteria. Twelve hours after infection the mycobacteria are found within membrane-bound phagosomes. By 96 hours we could see some lysis of the DC although there was no evidence of apoptosis; the presence of Mtb was more difficult to detect by this stage. Over a 5-day period, the viability of M.tuberculosis that had been phagocytosed by DC was found to decline slightly, whereas an identical inoculum was able to replicate in cultured macrophages. We therefore investigated the mechanisms associated with this growth suppression and found that both oxygen and nitrogen radicals where involved. Changes in cytokine production by DC infected with Mtb were observed, with a significant up-regulation of cytokines involved in Th1 and Tc1 responses. Similar results were obtained when primary bone-marrow derived DC were infected with Mtb. Characterisation of surface molecules expressed by DC which had been infected with Mtb confirmed the maturation process of the cells with significant up-regulation of the costimulatory molecules B7-1 and B7-2 and increased expression of MHC class II molecules and ICAM-1. This response was found to be dependent on the rapid activation of the nuclear transcription factor NF-kB, and was independent of TNF-a release. We also demonstrated expression of c-Rel and Rel-B proteins in Mtb-activated DC. In addition to these in vitro studies, we have also demonstrated that Mtb-activated DC are extremely efficient in priming naive murine T cells and that this immune response does not require T cell help. Mtb-activation of DC also results in efficient cross priming of T cells specific for Mtb. These responses enable Mtb-activated DC to confer protection against challenge with viable Mtb, with levels of protection as good or better than those conferred by BCG. The further understanding of the mechanisms involved in the interaction of mycobacteria with DC, and the mechanisms underlying the transfer of protective immunity, should provide important insights for the development of novel approaches to immunotherapy or for the development of new vaccines.

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