Biomedical subjects

W J Britton

Publications and source records attributed to W J Britton.

At least 19 recordsLinked to original sources

PubMed2006

Effect of phthiocerol dimycocerosate deficiency on the transcriptional response of human macrophages to Mycobacterium tuberculosis.

The control of mycobacterial infections is dependent on the finely tuned synergism between the innate and adaptive immune responses. The macrophage is the major host cell for Mycobacterium tuberculosis and the degree of virulence of mycobacteria may influence the initial macrophage response to infection. The cell wall molecule, phthiocerol dimycocerosate (DIM), is an important virulence factor that influences the early growth of M. tuberculosis in the lungs. To explore the basis for this effect we have compared the early gene response of human THP-1 macrophages to infection with virulent M. tuberculosis and the DIM-deficient DeltafadD26 M. tuberculosis strain using microarrays. Detailed analysis revealed a common core of macrophage genes, which were rapidly induced following infection with both strains, and deficiency of DIM had no significant effect on this initial macrophage transcriptional responses. In addition to chemokines and pro-inflammatory cytokines, the early response genes included components of the Toll-like receptor signalling, antigen presentation and apoptotic pathways, interferon response genes, cell surface receptors and their ligands, including TNF-related apoptosis inducing ligand (TRAIL) and CD40, and other novel genes. Therefore, although fadD26 deficiency is responsible for the early attenuation of the growth of M. tuberculosis in vivo, this effect is not associated with differences in the initial macrophage transcriptional response.

Animals↗

PubMed2005

Sensitivity of human gamma interferon assay and tuberculin skin testing for detecting infection with Mycobacterium tuberculosis in patients with culture positive tuberculosis.

SETTING: Nine university-affiliated chest clinics in Australia. OBJECTIVE: To evaluate the sensitivity of a whole blood human gamma-interferon assay (HGIA, QuantiFERON-TB) for specific T lymphocyte responses and Tuberculin skin testing (TST) for the detection of Mycobacterium tuberculosis infection in subjects with culture-proven M. tuberculosis disease (TBCP). DESIGN: Prospective testing of 129 patients with recent TBCP and 100 patients with non-tuberculosis lung disease (NTBLD). RESULTS: Using a defined level of specific IFN-gamma production and TST 10mm as positive cut-offs, the sensitivity of HGIA was 81% compared to 89% for TST (p=0.06). When positive responses in both TST and HGIA were combined, 96% of TB patients were detected. For the NTBLD group, 43% of whom were born overseas, 73% were negative for both the HGIA and TST. Prior immunization with M. bovis (bacille Calmette-Guerin) (BCG) or the type of TB had no effect on the sensitivities of the assays. For those treated for <2 months, the sensitivities for both assays were 84%, but for those treated for >2 months the sensitivity of TST (90%) tended to be higher than for HGIA (81%) (p=0.07). The distribution of TST results in TB patients showed a broad peak between 10 and 25 mm, while the results in the HGIA were bimodal in both TB and NTBLD patients. CONCLUSION: HGIA may prove an alternative to skin testing for detecting M. tuberculosis infection in certain settings.

Adult↗

PubMed2005

Independent protective effects for tumor necrosis factor and lymphotoxin alpha in the host response to Listeria monocytogenes infection.

Although the essential role of tumor necrosis factor (TNF) in resistance to Listeria monocytogenes infection is well established, the roles of the related cytokines lymphotoxin alpha (LTalpha) and lymphotoxin beta (LTbeta) are unknown. Using C57BL/6 mice in which the genes for these cytokines were disrupted, we examined the contributions of TNF, LTalpha, and LTbeta in the host response to Listeria. To overcome the lack of peripheral lymph nodes in LTalpha(-/-) and LTbeta(-/-) mice, bone marrow chimeras were constructed. TNF(-/-) and LTalpha(-/-) chimeras that lacked both secreted LTalpha(3) and membrane-bound LTalpha(1)beta(2) and LTalpha(2)beta(1) were highly susceptible and succumbed 4.5 and 6 days, respectively, after a low-dose infection (200 CFU). LTbeta(-/-) chimeras, which lacked only membrane-bound LT, controlled the infection in a manner comparable to wild-type (WT) chimeras. The Listeria-specific proliferative and gamma interferon T-cell responses were equivalent in all five groups of infected mice (LTalpha(-/-) and LTbeta(-/-) chimeras, WT chimeras, and TNF(-/-) and WT mice). TNF(-/-) mice and LTalpha(-/-) chimeras, however, failed to generate the discrete foci of lymphocytes and macrophages that are essential for bacterial elimination. Rather, aberrant necrotic lesions comprised predominantly of neutrophils with relatively few lymphocytes and macrophages were observed in the livers and spleens of TNF(-/-) and LTalpha(-/-) chimeras. Therefore, in addition to TNF, soluble LTalpha(3) plays a separate essential role in control of listerial infection through control of leukocyte accumulation and organization in infected organs.

Animals↗

PubMed2004

T cell-derived tumour necrosis factor is essential, but not sufficient, for protection against Mycobacterium tuberculosis infection.

Tumour necrosis factor (TNF) is critical for sustained protective immunity against Mycobacterium tuberculosis infection. To investigate the relative contributions of macrophage- and T cell-derived TNF towards this immunity T cells from wild-type (WT) or TNF-/- mice were transferred into RAG-/- or TNF-/- mice which were then infected with M. tuberculosis. Infected RAG-/- mice and RAG-/- recipients of TNF deficient T cells developed overwhelming infection, with extensive pulmonary and hepatic necrosis and succumbed with a median of only 16 days infection. By contrast, RAG-/- recipients of WT T cells showed a significant increase in survival with a median of 32 days. Although initial bacterial growth was similar in all groups of RAG-/- mice, the transfer of WT, but not TNF-/-, T cells led to the formation of discrete foci of leucocytes and macrophages and delayed the development of necrotizing pathology. To determine requirements for macrophage-derived TNF, WT or TNF-/- T cells were transferred into TNF-/- mice at the time of M. tuberculosis infection. Transfer of WT T cells significantly prolonged survival and reduced the early tissue necrosis evident in the TNF-/- mice, however, these mice eventually succumbed indicating that T cell-derived TNF alone is insufficient to control the infection. Therefore, both T cell- and macrophage-derived TNF play distinct roles in orchestrating the protective inflammatory response and enhancing survival during M. tuberculosis infection.

Animals↗

PubMed2002

Cord blood mononuclear cell cytokine responses in relation to maternal house dust mite allergen exposure.

BACKGROUND: Cord blood mononuclear cells have demonstrated specific immune responses to environmental allergens. OBJECTIVE: To establish whether the nature of this response is related to the level of maternal antenatal exposure to house dust mite (HDM) allergen and, hence, whether antenatal allergen avoidance may have a role in the prevention of allergic sensitization in children. METHODS: Children with a family history of asthma were recruited antenatally as subjects in a randomised controlled trial: the Childhood Asthma Prevention Study. HDM allergen (Der p 1) concentrations were measured in dust collected from the maternal bed at 36 weeks gestation. Cord blood mononuclear cells were stimulated in culture, separately, with phytohaemaglutinin (PHA) and HDM extract. Cytokine IL-4, IL-5, IL-10 and IFN-gamma concentrations in supernatant were measured by ELISA. mRNA signals for these cytokines were measured using RT-PCR. RESULTS: The median concentration of HDM allergen was 18.4 microg/g (interquartile range 7.3-35.3 microg/g). Median concentrations of IL-4, IL-5, IL-10 and IFN-gamma, after PHA stimulation were 4, 19, 401 and 1781 pg/mL, respectively. After HDM allergen stimulation the median concentrations were 0, 0, 20 and 14 pg/mL, respectively. The distribution of mRNA cytokine signals was similar. Neither cytokine protein concentrations nor cytokine mRNA signal levels were correlated with the concentration of HDM allergen in the mothers' beds at 36 weeks gestation. CONCLUSION: These findings do not support the view that the prevention of allergic disease in children requires the institution of HDM avoidance interventions during pregnancy.

Air Pollution, Indoor↗

PubMed2002

Evidence for the genetic control of immunoglobulin E reactivity to the allergens of Alternaria alternata.

BACKGROUND: The fungus Alternaria alternata contains potent allergens, and sensitization to these allergens is associated with a high risk of respiratory disease. The influence of genetic regulation on sensitization to Alternaria is unknown. OBJECTIVE: To determine the influence of genetic factors on IgE responses to specific allergens of Alternaria. METHODS: The concordance of skin prick test (SPT), radioallergosorbent test (RAST) and IgE-binding profiles of sera were examined from a large cohort of monozygotic and dizygotic twins. RESULTS: Casewise concordance for a positive SPT response was monozygous (MZ) 66%: dizygous (DZ) 40% (P = 0.002). Logistic regression confirmed that casewise concordance was significantly stronger between MZ than DZ pairs. Immunoblotting against an Alternaria extract revealed 19 allergenic bands. The differences in concordance between the different bands were not significant for either the MZ (P = 0.97) or DZ (P = 0.84) groups. The pooled MZ : DZ difference in concordance was just significant (P = 0.049), suggesting an overall genetic effect on the response to Alternaria. This was reinforced by the comparison of the MZ and DZ correlations for total number of bands recognized (MZ r = 0.65; DZ r = 0.37, P = 0.015). Overall, there was a moderate correlation between the individual SPT weal size and RAST score (r(2) = 0.41) and a substantial correlation between the number of immunoblotted bands and RAST scores (r(2) = 0.79). CONCLUSION: There is a strong genetic influence on IgE response to the mixture of Alternaria allergens and a lesser effect on IgE response to individual allergens.

Allergens↗

PubMed2001

Secreted lymphotoxin-alpha is essential for the control of an intracellular bacterial infection.

Although the essential role of tumor necrosis factor (TNF) in the control of intracellular bacterial infection is well established, it is uncertain whether the related cytokines lymphotoxin-alpha (LTalpha3) and lymphotoxin-beta (LTbeta) have independent roles in this process. Using C57Bl/6 mice in which the genes for these cytokines have been disrupted, we have examined the relative contribution of secreted LTalpha3 and membrane-bound LTbeta in the host response to aerosol Mycobacterium tuberculosis infection. To overcome the lack of peripheral lymph nodes in LTalpha-/- and LTbeta-/- mice, bone marrow chimeric mice were constructed. LT-/- chimeras, which lack both secreted LTalpha3 and membrane-bound LTbeta (LT1beta2 and LT2beta1), were highly susceptible and succumbed 5 wk after infection. LTbeta-/- chimeras, which lack only the membrane-bound LTbeta, controlled the infection in a comparable manner to wild-type (WT) chimeric mice. T cell responses to mycobacterial antigens and macrophage responses in LTalpha-/- chimeras were equivalent to those of WT chimeras, but in LTalpha-/- chimeras, granuloma formation was abnormal. LTalpha-/- chimeras recruited normal numbers of T cells into their lungs, but the lymphocytes were restricted to perivascular and peribronchial areas and were not colocated with macrophages in granulomas. Therefore, LTalpha3is essential for the control of pulmonary tuberculosis, and its critical role lies not in the activation of T cells and macrophages per se but in the local organization of the granulomatous response.

Animals↗

PubMed2001

DNA encoding a single mycobacterial antigen protects against leprosy infection.

The continuing incidence of leprosy infection around the world and the inability of Mycobacterium bovis bacille Calmette-Guérin (BCG) to protect certain populations clearly indicates that an improved vaccine against leprosy is needed. The immuno dominant 35 kDa protein, shared by Mycobacterium leprae and Mycobacterium avium, but not Mycobacterium tuberculosis or BCG, is recognised by >90% of leprosy patients, making it an ideal candidate antigen for a subunit vaccine. Immunization of outbred Swiss Albino mice with a DNA-35 vaccine stimulated specific T cell activation and IFN-gamma production. DNA-35 immunization induced significant levels of protection against M. leprae footpad infection, comparable to that produced by BCG. Therefore, DNA immunization with the 35 kDa antigen is effective against M. leprae infection and genetic immunization with a combination of antigens holds the potential for an improved vaccine against leprosy.

Animals↗

PubMed2001

Comparative protective effects of recombinant DNA and Mycobacterium bovis bacille Calmette-Guérin vaccines against M. avium infection.

A range of strategies are being explored to develop more effective vaccines against mycobacterial infection, including immunization with DNA plasmids encoding single mycobacterial bacterial genes and the use of recombinant live vectors based on the current vaccine, Mycobacterium bovis bacille Calmette-Guérin (BCG). We have compared these two approaches using a model of virulent M. avium infection, and the gene for the immunodominant 35 kDa protein which is shared by M. avium and M. leprae, but absent from BCG. Recombinant BCG over-expressing the M. avium 35 kDa protein (BCG-35) induced strong antigen-specific proliferative and interferon-gamma (IFN-gamma)-secreting T cell responses. These were comparable to those induced by a single immunization with a plasmid expressing the same antigen (DNA-35); however, repeat DNA-35 immunization evoked the strongest IFN-gamma release. Immunization with BCG-35 significantly reduced the growth of virulent M. avium, although this effect was similar to that induced by wild-type BCG. Immunization with DNA-35 resulted in significantly greater (2 x log(10)) reduction in the growth of M. avium. Prime-boost strategies combining DNA-35 and BCG-35 increased the protective effect above that achieved by BCG-35, but they were not more protective than DNA-35 alone. Therefore, recombinant BCG-35 and BCG induced similar levels of protection in this model, and maximal protection against M. avium infection was attained by immunization with DNA encoding the 35 kDa protein.

Animals↗

PubMed2001

Induction of CD8+ T-lymphocyte responses to a secreted antigen of Mycobacterium tuberculosis by an attenuated vaccinia virus.

Protective immunity against Mycobacterium tuberculosis infection requires the activation of mycobacterium-specific CD8+ T cells, as well as CD4+ T cells. Therefore, optimizing strategies that stimulate CD8+ T cells recognizing dominant mycobacterial antigens, including secreted proteins, may lead to the development of more effective vaccines against tuberculosis. To generate a viral vaccine that is safe in humans, the early secreted protein, MPT64, was expressed in the attenuated vaccinia virus (VV) strain, modified vaccinia virus Ankara (MVA-64). The immunogenicity of MVA-64 was compared with that of the Western Reserve strain of VV (VVWR-64). The replication-defective MVA-64 was as efficient as VVWR-64 in inducing specific antibodies and cytolytic T-cell responses to a defined H-2-Db-restricted epitope on MTP-64. In addition, priming with MPT64-expressing plasmid DNA (DNA-64), and boosting with either MVA-64 or VVWR-64, markedly enhanced MPT64-specific cytolytic and IFN-gamma-producing CD8+ T-cell responses. These findings suggest that MVA may be a suitable vaccine carrier for stimulating mycobacterium-specific CD8+ T-cell responses and may be particularly relevant for developing vaccines for use in regions endemic for tuberculosis and HIV infection.

AIDS-Related Opportunistic Infections↗

PubMed2001

Dendritic cells infected with Mycobacterium bovis bacillus Calmette Guerin activate CD8(+) T cells with specificity for a novel mycobacterial epitope.

Although CD4(+) T cells are essential for protective immunity against Mycobacterium tuberculosis infection, recent reports indicate that CD8(+) T cells may also play a critical role in the control of this infection. However, the epitope specificity and the mechanisms of activation of mycobacteria-reactive CD8(+) T cells are poorly characterized. In order to study the CD8(+) T cell responses to the model mycobacterial antigen, MPT64, we used recombinant vaccinia virus expressing MPT64 (VVWR-64) and a panel of MPT64-derived peptides to establish that the peptide MPT64(190-198) contains an H-2D(b)-restricted CD8(+) T cell epitope. A cytotoxic T lymphocyte response to this peptide could be demonstrated in M. bovis bacillus Calmette Guerin (BCG)-infected mice following repeated in vitro stimulation. When bone marrow-derived dendritic cells (DC) were infected with BCG, the expression of MHC class I molecules by DC was up-regulated in parallel with MHC class II and B7-2, whereas CD1d expression level was not modified. Moreover, BCG-infected DC activated MPT64(190-198)-specific CD8(+) T cells to secrete IFN-gamma, although with a lower efficacy than VVWR-64-infected DC. The production of IFN-gamma by MPT64(190-198)-specific CD8(+) T cells was inhibited by antibodies to MHC class I, but not to CD1d. These data suggest that mycobacteria-specific CD8(+) T cells are primed during infection. Therefore, anti-mycobacterial vaccine strategies targeting the activation of specific CD8(+) T cells by DC may have improved protective efficacy.

Animals↗

PubMed2001

Stimulation of dendritic cells via CD40 enhances immune responses to Mycobacterium tuberculosis infection.

The resolution of pulmonary tuberculosis (TB) critically depends on the development of the Th1 type of immune responses, as exemplified by the exacerbation of TB in IL-12-deficient mice. Therefore, vaccination strategies optimizing IL-12 production by antigen-presenting cells (APC) in response to mycobacteria may have enhanced protective efficacy. Since dendritic cells (DC) are the critical APC for activation of CD4+ and CD8+ T cells, we examined whether stimulation of Mycobacterium bovis bacillus Calmette Guérin (BCG)-infected DC via CD40 increased their ability to generate Th1-oriented cellular immune responses. Incubation of DC with an agonistic anti-CD40 antibody activated CD40 signaling in DC, as shown by increased expression of major histocompatibility complex class II and costimulatory molecules, mRNA production for proinflammatory cytokines and interleukin 12 (IL-12) p40. This activation pattern was maintained when DC were stimulated with anti-CD40 antibody and infected with BCG. Importantly, CD40-stimulated BCG-infected DC displayed increased capacity to release bioactive IL-12 and to activate gamma interferon (IFN-gamma) producing T cells in vitro. Moreover, when C57BL/6 mice were immunized with these DC and challenged with aerosol Mycobacterium tuberculosis, increased levels of mRNA for IL-12 p40, IL-18, and IFN-gamma were present in the draining mediastinal lymph nodes. However, the mycobacterial burden in the lungs was not reduced compared to that in mice immunized with BCG-infected non-CD40-stimulated DC. Therefore, although the manipulation of DC via CD40 is effective for enhancing immune responses to mycobacteria in vivo, additional strategies are required to increase protection against virulent M. tuberculosis infection.

Animals↗

PubMed2001

Priming by DNA immunization augments protective efficacy of Mycobacterium bovis Bacille Calmette-Guerin against tuberculosis.

Sequential immunization with mycobacterial antigen Ag85B-expressing DNA and Mycobacterium bovis bacille Calmette-Guerin (BCG) was more effective than BCG immunization in protecting against Mycobacterium tuberculosis infection. Depletion of the CD8(+) T cells in the immunized mice impaired protection in their spleens, indicating that this improved efficacy was partially mediated by CD8(+) T cells.

Acyltransferases↗

PubMed2001

Vaccination with DNA of the Mycobacterium tuberculosis 85B antigen protects mouse foot pad against infection with M. leprae.

A DNA vaccine composed of the gene for the common mycobacterial secreted protein antigen 85B was demonstrated to protect the mouse foot pad against infection with Mycobacterium leprae. The protective effect was demonstrated by a 61%-88% reduction in the bacterial number, a protective effect less than that of BCG. The same DNA vaccine has been shown to protect mice against M. tuberculosis infection, and the importance of testing other candidate tuberculosis vaccines for their potential to protect against leprosy is discussed.

Acyltransferases↗

PubMed2000

CD4+ and CD8+ T cells mediate adoptive immunity to aerosol infection of Mycobacterium bovis bacillus Calmette-Guérin.

An adoptive-transfer model using recombinase activation gene-deficient (RAG-1-/-) mice was developed to evaluate CD4+ and CD8+ T cell responses to infection with Mycobacterium bovis bacillus Calmette-Guérin (BCG). After receiving immune, unfractionated T cells or T cell subsets isolated by fluorescence-activated cell sorter, the RAG-1-/- mice were exposed to aerosol BCG, and the bacteria load in the infected organs was examined 4 weeks later. Adoptive immunity was expressed more effectively in the spleens than in the lungs. Although CD4+ or unfractionated T cells protected both lungs and spleens, CD8+ T cells conferred significant protection only in the spleens and not in the lungs. The results confirm that in addition to CD4+, CD8+ T cells also play a role in the prevention of bacterial dissemination. This transfer model may be useful for dissecting T cell responses to mycobacterial infection.

Adoptive Transfer↗

PubMed2000

Up-regulation of VCAM-1 and differential expansion of beta integrin-expressing T lymphocytes are associated with immunity to pulmonary Mycobacterium tuberculosis infection.

Immune responses rely on an intricate system of adhesion molecules to coordinate the homing and retention of lymphocytes in both secondary lymphoid tissues and at sites of infection. To define the events associated with pulmonary immune responses, the expression of endothelial addressins and integrins on T cells was analyzed during Mycobacterium tuberculosis infection. In infected lung, expression of endothelial VCAM-1, but not mucosal addressin cell adhesion molecule-1, was up-regulated from 4 wk postinfection and persisted to at least 12 wk. Subsequent analysis of the corresponding integrins expressed on lung CD4+ and CD8+ T cells revealed an accumulation of beta1high/beta7-/low, and to a lesser extent beta7high, integrin-expressing T cells during infection. Examination of integrin heterodimers showed that while alpha4 integrin was predominantly expressed on beta1high/beta7-/low cells, alphaE integrin was primarily associated with beta7high. The majority of activated/memory T cells recruited during infection expressed high levels of beta1 integrin and undetectable or low levels of beta7 integrin. These T cells were capable of producing IFN-gamma, a cytokine crucial for controlling M. tuberculosis infection. Rapid expansion of beta1high, beta7-, and beta7high T cell populations in the lung upon secondary mycobacterial infection indicates the participation of these populations in the acquired immune response to the infection. Furthermore, treatment of infected mice with mAb to alpha4 or alpha4beta7 integrin led to a reduction in lymphocytes and increase in granulocytes in the pulmonary infiltrate. These results reveal a crucial role for adhesion molecules in the generation of an effective pulmonary immune response to M. tuberculosis infection.

Animals↗

PubMed2000

Protective effect of DNA immunization against mycobacterial infection is associated with the early emergence of interferon-gamma (IFN-gamma)-secreting lymphocytes.

The development of more effective anti-tuberculosis (TB) vaccines would contribute to the global control of TB. Understanding the activated/memory T cell response to mycobacterial infection and identifying immunological correlates of protective immunity will facilitate the design and assessment of new candidate vaccines. Therefore, we investigated the kinetics of the CD4+ T cell response and IFN-gamma production in an intravenous challenge model of Mycobacterium bovis bacille Calmette-Guérin (BCG) before and after DNA immunization. Activated/memory CD4+ T cells, defined as CD44hiCD45RBlo, expanded following infection, peaking at 3-4 weeks, and decreased as the bacterial load fell. Activated/memory CD4+ T cells were the major source of IFN-gamma and the level of antigen-specific IFN-gamma-secreting lymphocytes, detected by ELISPOT, paralleled the changes in bacterial load. To examine the effects of a DNA vaccine, we immunized mice with a plasmid expressing the mycobacterial secreted antigen 85B (Ag85B). This led to a significant reduction in mycobacteria in the liver, spleen and lung. This protective effect was associated with the rapid emergence of antigen-specific IFN-gamma-secreting lymphocytes which were detected earlier, at day 4, and at higher levels than in infected animals immunized with a control vector. This early and increased response of IFN-gamma-secreting T cells may serve as a correlate of protective immunity for anti-TB vaccines.

Adjuvants, Immunologic↗

PubMed2000

Interaction of dendritic cells with mycobacteria: where the action starts.

Dendritic cells (DC) are the major antigen-presenting cells in the induction of cellular responses to intracellular pathogens, such as mycobacteria. Recent studies have shown that they also play a critical role in the regulation of immune responses. The interaction of DC with microbial antigens may be the controlling factor in the development of a Th1-orientated protective immunity. Analysis of the innate response of DC to mycobacteria and the involvement of the DC receptors in antigen recognition have highlighted the pivotal role of these cells in T-cell activation. Mycobacteria-infected DC have an enhanced capacity to release pro-inflammatory cytokines and chemokines and are potent inducers of interferon-gamma-producing cells in vivo. Therefore, DC manipulation for maximal antigen presentation and Th1 cytokine production may form the basis of a new generation of vaccines, with improved efficacy against mycobacterial infections.

Animals↗