Interferon-gamma independent formation of pulmonary granuloma in mice by injections with trehalose dimycolate (cord factor), lipoarabinomannan and phosphatidylinositol mannosides isolated from Mycobacterium tuberculosis.
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The mechanisms by which pulmonary granuloma formation is caused by administration of mycobacterial glycolipids such as trehalose dimycolate (TDM), lipoarabinomannan (LAM) and phosphatidylinositol mannosides (PIM) were investigated. When peritoneal and alveolar macrophages were stimulated with TDM, LAM and PIM in vitro, TDM exhibited the strongest tumour necrosis factor (TNF)-inducing activity. Responsiveness of macrophages from mice defected Toll-like receptor 4 (TLR4) was much higher than that of the wild-type mice. Although PIM and LAM also had a significant activity, LAM rather than PIM stimulated higher TNF-alpha production by alveolar macrophage. When mycobacterial glycolipids were injected as water-in-oil-in-water emulsion into mice via the tail vein, development of pulmonary granuloma in response to glycolipids were related closely to their TNF-inducing activity and TDM exhibited the strongest activity. Granuloma formation was observed not only in mice lacking interleukin (IL)-12 signalling but also interferon (IFN)-gamma knock-out mice. Granuloma formation caused by glycolipids correlated with TNF-alpha levels in lungs. Administration of anti-TNF-alpha monoclonal antibody into TDM-injected IFN-gamma knock-out mice decreased in granuloma formation, suggesting that development of pulmonary granuloma by mycobacterial glycolipids such as TDM is due to IFN-gamma-independent and TNF-alpha-dependent pathway. (+info)
Trehalose 6,6'-dimycolate and lipid in the pathogenesis of caseating granulomas of tuberculosis in mice.
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Trehalose 6,6'-dimycolate (TDM) is the most abundant, most granulomagenic, and most toxic lipid extractable from the surface of virulent Mycobacterium tuberculosis (MTB). We further examined its toxicity, which requires activation by oily surfaces. Injections of MTB and/or TDM into sensitized mice induced caseating granulomas that centered on oil droplets. If large doses of MTB were injected in saline, caseating granulomas developed in adipose tissue, but MTB with surface TDM removed induced only acute inflammation that did not persist. Variations in protocols produced several variants of caseating granulomas, each with characteristics of human tuberculosis. In each instance, MTB were localized in fat cells or oil drops during initiation of caseating granulomas suggesting that necrosis was caused by activation of the toxicity of TDM toxicity. Evidence extending these findings to the lung was derived from the observation that in sensitized mice, as in humans, tuberculosis development stimulates accumulation of lipid selectively in alveoli. MTB preferentially associated with lipid droplets in developing necrotic foci in late-stage murine tuberculosis. This supports the hypothesis that pulmonary tuberculosis sequesters MTB in a protected environment that accumulates lipid until it is able to activate the toxicity of TDM and initiate necrosis that results in caseating granulomas. (+info)
Of mice, men, and elephants: Mycobacterium tuberculosis cell envelope lipids and pathogenesis.
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Mycolic acids and structures attached to them constitute a major part of the protective envelope of Mycobacterium tuberculosis, and for this reason, their role in tuberculosis pathogenesis has been extensively studied. In this issue of the JCI, Rao et al. examine the effect of trans-cyclopropanation of oxygenated mycolic acids attached to trehalose dimycolate (TDM) on the murine immune response to infection (see the related article beginning on page 1660). Surprisingly, they found that an M. tuberculosis mutant lacking trans-cyclopropane rings was hypervirulent in mice. The recent recognition of a hypervirulence phenotype in mice associated with laboratory and clinical M. tuberculosis strains with altered cell wall components has provided new insights into how M. tuberculosis may establish persistent infection. However, to date, characterization of these bioactive products in pathogenesis has been largely reductionistic; the relationship of their effects observed in mice to the persistent infection and tuberculosis caused by M. tuberculosis observed in humans remains obscure. (+info)
Trans-cyclopropanation of mycolic acids on trehalose dimycolate suppresses Mycobacterium tuberculosis -induced inflammation and virulence.
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Recent studies have shown that fine structural modifications of Mycobacterium tuberculosis cell envelope lipids mediate host cell immune activation during infection. One such alteration in lipid structure is cis-cyclopropane modification of the mycolic acids on trehalose dimycolate (TDM) mediated by proximal cyclopropane synthase of alpha mycolates (pcaA), a proinflammatory lipid modification during early infection. Here we examine the pathogenetic role and immunomodulatory function of mycolic acid cyclopropane stereochemistry by characterizing an M. tuberculosis cyclopropane-mycolic acid synthase 2 (cmaA2) null mutant (Delta cmaA2) that lacks trans-cyclopropanation of mycolic acids. Although titers of WT and Delta cmaA2 organisms were identical during mouse infection, Delta cmaA2 bacteria were hypervirulent while inducing larger granulomas than WT M. tuberculosis. The hypervirulence of the Delta cmaA2 strain depended on host TNF-alpha and IFN-gamma. Loss of trans-cyclopropanation enhanced M. tuberculosis-induced macrophage inflammatory responses, a phenotype that was transferable with petroleum ether extractable lipids. Finally, purified TDM lacking trans-cyclopropane rings was 5-fold more potent in stimulating macrophages. These results establish cmaA2-dependent trans-cyclopropanation of TDM as a suppressor of M. tuberculosis-induced inflammation and virulence. In addition, cyclopropane stereochemistries on mycolic acids interact directly with host cells to both positively and negatively influence host innate immune activation. (+info)
CD3+ cells transfer the hypersensitive granulomatous response to mycobacterial glycolipid trehalose 6,6'-dimycolate in mice.
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The granulomatous response is the characteristic histological feature of Mycobacterium tuberculosis infection that is essential for organism containment. Trehalose 6,6-dimycolate (TDM), a cell-wall glycolipid present on most mycobacterial species, has been implicated in the pathogenesis of M. tuberculosis infection. TDM has potent immunoregulatory and inflammatory properties, and can be used to model granulomatous reactions that mimic, in part, pathology caused during active infection. This study examined the hypersensitive granulomatous response, focusing on cellular responses specific to TDM. Lungs from mice immunized with TDM emulsion demonstrated exacerbated histological damage, inflammation, and lymphocytic infiltration upon subsequent challenge with TDM. Splenocytes recovered from these mice demonstrated significant interferon (IFN)-gamma production during recall response to TDM, as well as increased production of proinflammatory mediators (tumour necrosis factor-alpha, interleukin-6 and macrophage inflammatory protein-1alpha). The exacerbated response could be adoptively transferred to naive mice. Administration of non-adherent lymphocytes or purified CD3(+) cells from TDM-immunized mice led to increased inflammation, lymphocytic infiltration, and vascular endothelial cell damage upon challenge with TDM. Recipient mice that received immunized CD3(+) lymphocytes demonstrated significant increases in Th1-type cytokines and proinflammatory mediators in lung tissue following TDM challenge. When CD1d(-/-) mice were immunized with TDM, they failed to generate a specific IFN-gamma response, suggesting a role for this molecule in the generation of hypersensitivity. These experiments provide further evidence for the involvement of TDM-specific CD3(+) T cells in pathological damage elicited during M. tuberculosis infection. (+info)
Adjuvant-enhanced antibody responses in the absence of toll-like receptor signaling.
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Innate immune signals mediated by Toll-like receptors (TLRs) have been thought to contribute considerably to the antibody-enhancing effects of vaccine adjuvants. However, we report here that mice deficient in the critical signaling components for TLR mount robust antibody responses to T cell-dependent antigen given in four typical adjuvants: alum, Freund's complete adjuvant, Freund's incomplete adjuvant, and monophosphoryl-lipid A/trehalose dicorynomycolate adjuvant. We conclude that TLR signaling does not account for the action of classical adjuvants and does not fully explain the action of a strong adjuvant containing a TLR ligand. This may have important implications in the use and development of vaccine adjuvants. (+info)
Identification of trehalose dimycolate (cord factor) in Mycobacterium leprae.
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Glycolipids of Mycobacterium leprae obtained from armadillo tissue nodules infected with the bacteria were analyzed. Mass spectrometric analysis of the glycolipids indicated the presence of trehalose 6,6'-dimycolate (TDM) together with trehalose 6-monomycolate (TMM) and phenolic glycolipid-I (PGL-I). The analysis showed that M. leprae-derived TDM and TMM possessed both alpha- and keto-mycolates centering at C78 in the former and at C81 or 83 in the latter subclasses, respectively. For the first time, MALDI-TOF mass analyses showed the presence of TDM in M. leprae. (+info)
Control of cell wall assembly by a histone-like protein in Mycobacteria.
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Bacteria coordinate assembly of the cell wall as well as synthesis of cellular components depending on the growth state. The mycobacterial cell wall is dominated by mycolic acids covalently linked to sugars, such as trehalose and arabinose, and is critical for pathogenesis of mycobacteria. Transfer of mycolic acids to sugars is necessary for cell wall biogenesis and is mediated by mycolyltransferases, which have been previously identified as three antigen 85 (Ag85) complex proteins. However, the regulation mechanism which links cell wall biogenesis and the growth state has not been elucidated. Here we found that a histone-like protein has a dual concentration-dependent regulatory effect on mycolyltransferase functions of the Ag85 complex through direct binding to both the Ag85 complex and the substrate, trehalose-6-monomycolate, in the cell wall. A histone-like protein-deficient Mycobacterium smegmatis strain has an unusual crenellated cell wall structure and exhibits impaired cessation of glycolipid biosynthesis in the growth-retarded phase. Furthermore, we found that artificial alteration of the amount of the extracellular histone-like protein and the Ag85 complex changes the growth rate of mycobacteria, perhaps due to impaired down-regulation of glycolipid biosynthesis. Our results demonstrate novel regulation of cell wall assembly which has an impact on bacterial growth. (+info)