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Biomedical subjects

T H Ottenhoff

Publications and source records attributed to T H Ottenhoff.

At least 19 recordsLinked to original sources
PubMed2003

[Research into new methods for diagnosing, treating and preventing tuberculosis].

Tuberculosis control requires improved diagnostics, drugs, and vaccines. Their development is facilitated by progress in immunology, molecular biology, and genomics. In addition to sputum smear and culture, amplification techniques can already be used to diagnose tuberculosis and antigen-detection tests for this purpose are being developed. Molecular typing and DNA microarrays provide new insights in the natural history and transmission of tuberculosis. In addition to established drugs such as rifampicin, isoniazid, pyrazinamide, and ethambutol, a limited number of new drugs have been discovered such as rifampicin derivates and fluorochinolones. Improved screening techniques and insights from genomics may lead to new drugs being discovered. Factors hampering the development and evaluation of new vaccines include problems with extrapolation from animal models, incomplete natural immunity, and limited knowledge about protective immunity. However, new candidate vaccines are being developed and will be tested on humans in the near future.

Animals↗
PubMed2001

Mycobacterium leprae-specific, HLA class II-restricted killing of human Schwann cells by CD4+ Th1 cells: a novel immunopathogenic mechanism of nerve damage in leprosy.

Peripheral nerve damage is a major complication of reversal (or type-1) reactions in leprosy. The pathogenesis of nerve damage remains largely unresolved, but detailed in situ analyses suggest that type-1 T cells play an important role. Mycobacterium leprae is known to have a remarkable tropism for Schwann cells of the peripheral nerve. Reversal reactions in leprosy are often accompanied by severe and irreversible nerve destruction and are associated with increased cellular immune reactivity against M. leprae. Thus, a likely immunopathogenic mechanism of Schwann cell and nerve damage in leprosy is that infected Schwann cells process and present Ags of M. leprae to Ag-specific, inflammatory type-1 T cells and that these T cells subsequently damage and lyse infected Schwann cells. Thus far it has been difficult to study this directly because of the inability to grow large numbers of human Schwann cells. We now have established long-term human Schwann cell cultures from sural nerves and show that human Schwann cells express MHC class I and II, ICAM-1, and CD80 surface molecules involved in Ag presentation. Human Schwann cells process and present M. leprae, as well as recombinant proteins and peptides to MHC class II-restricted CD4(+) T cells, and are efficiently killed by these activated T cells. These findings elucidate a novel mechanism that is likely involved in the immunopathogenesis of nerve damage in leprosy.

Animals↗
PubMed2001

Natural T-helper immunity against human papillomavirus type 16 (HPV16) E7-derived peptide epitopes in patients with HPV16-positive cervical lesions: identification of 3 human leukocyte antigen class II-restricted epitopes.

Tumor-specific T-helper (Th) immunity was found to play a pivotal role in the natural and vaccine-induced immune defense against tumors. Since the majority of cervical cancers express human papillomavirus type 16 (HPV16) E7 oncoprotein, it is important to investigate the Th response against this target antigen in detail. By means of PBMC cultures from HLA-typed healthy donors, we identified the central part of HPV16 E7 (E7(41-72)) as the major immunogenic region within this antigen. Furthermore, we mapped 3 distinct Th epitopes within this region (DR15/E7(50-62), DR3/E7(43-77), DQ2/E7(35-50)). In a parallel approach, employing IFN-gamma ELISPOT analysis, we detected Th immunity against HPV16 E7 in subjects with HPV16+ lesions. Several of these responses matched with the 3 Th epitopes defined in our study. A number of other HPV16+ subjects did not display any E7-specific type 1 cytokine-producing T-cell immunity, indicating failure of the immune response. Our combined data argue for more extensive as well as longitudinal analysis of HPV16-specific T-cell immunity using the ELISPOT assay described, as well as for HPV-specific vaccination of individuals with HPV+ lesions.

Cancer Vaccines↗
PubMed2001

The identification of a common pathogen-specific HLA class I A*0201-restricted cytotoxic T cell epitope encoded within the heat shock protein 65.

Bacterial antigens recognized by CD8(+) T cells in the context of MHC class I are thought to play a crucial role in protection against pathogenic intracellular bacteria. Here, we demonstrate the induction of HLA-A*0201-restricted CD8(+) T cell responses against six new high-affinity HLA-A*0201-binding CTL epitopes, encoded within an immunodominant and highly conserved antigen of Mycobacteria, the heat shock protein 65 (hsp65). One of these epitopes, Mhsp65(9(369)), is identical in a large number of pathogenic bacteria, and is recognized in a CD8-independent fashion. Mhsp65(9(369)) could be presented by either mycobacterial hsp65-pulsed target cells or BCG-infected macrophages. Interestingly, T cells specific for this epitope did not recognize the corresponding human hsp65 homologue, probably due to structural differences as revealed by modeling studies. Furthermore, in vitro proteasome digestion analyses show that, whereas the mycobacterial hsp65 epitope is efficiently generated, the human hsp65 homologue is not, thus avoiding the induction of autoreactivity. Collectively, these findings describe high-affinity HLA class I-binding epitopes that are naturally processed and are recognized efficiently by MHC class I-restricted CD8(+) T cells, providing a rational basis for the development of subunit vaccine strategies against tuberculosis and other intracellular infectious diseases.

Amino Acid Sequence↗
PubMed2001

Repeatedly negative tuberculin skin tests followed by active tuberculosis in an immunocompetent individual.

We describe a woman who was repeatedly tuberculin (PPD) skin test negative after exposure to smear-positive tuberculosis (TB), but developed active TB with a positive skin test 7 years later. Molecular epidemiologic evidence is presented that the infection was contracted 7 years previously from the original source case. PPD skin testing is subject to many technical and biological variables and this report underscores that this tool can fail to detect latent TB infection in some cases. The causes of false-negative and false-positive PPD skin test results are reviewed.

Acute Disease↗
PubMed2001

Multifocal osteomyelitis caused by nontuberculous mycobacteria in patients with a genetic defect of the interferon-gamma receptor.

We describe three patients with multifocal osteomyelitis caused by Mycobacterium avium and a family history of one or more first degree family members diagnosed with various clinical presentations of infections with nontuberculous mycobacteria. There was a significant delay in the diagnosis and they had a protracted course of their illness, which responded only slowly to prolonged multi-drug treatment. In one patient, additional treatment with interferon-gamma (IFN-gamma) was necessary. Macrophages of these patients had decreased in vitro responsiveness to IFN-gamma. Genomic sequencing revealed that these patients and their affected family members were heterozygous for a previously described dominant negative mutation in the gene encoding the IFN-gamma binding receptor-1 chain. The clinical presentations of the infections with nontuberculous mycobacteria in these families, with spread limited to skin, bone and lymph nodes, is discussed in the light of the immune mechanisms that are responsible for the clearance of otherwise poorly pathogenic environmental mycobacteria.

Adult↗
PubMed2001

Presence of human T-cell responses to the Mycobacterium leprae 45-kilodalton antigen reflects infection with or exposure to M. leprae.

The ability of the 45-kDa serine-rich Mycobacterium leprae antigen to stimulate peripheral blood mononuclear cell (PBMC) proliferation and gamma interferon (IFN-gamma) production was measured in leprosy patients, household contacts, and healthy controls from areas of endemicity in Mexico. Almost all the tuberculoid leprosy patients gave strong PBMC proliferation responses to the M. leprae 45-kDa antigen (92.8%; n = 14). Responses were lower in lepromatous leprosy patients (60.6%; n = 34), but some responses to the 45-kDa antigen were detected in patients unresponsive to M. leprae sonicate. The proportion of positive responses to the M. leprae 45-kDa antigen was much higher in leprosy contacts (88%; n = 17) than in controls from areas of endemicity (10%; n = 20). None of 15 patients with pulmonary tuberculosis gave a positive proliferation response to the 45-kDa antigen. The 45-kDa antigen induced IFN-gamma secretion similar to that induced by the native Mycobacterium tuberculosis 30/31-kDa antigen in tuberculoid leprosy patients and higher responses than those induced by the other recombinant antigens (M. leprae 10- and 65-kDa antigens, thioredoxin, and thioredoxin reductase); in patients with pulmonary tuberculosis it induced lower IFN-gamma secretion than the other recombinant antigens. These results suggest that the M. leprae 45-kDa antigen is a potent T-cell antigen which is M. leprae specific in these Mexican donors. This antigen may therefore have diagnostic potential as a new skin test reagent or as an antigen in a simple whole-blood cytokine test.

Antigens, Bacterial↗
PubMed2001

Tuberculin skin testing compared with T-cell responses to Mycobacterium tuberculosis-specific and nonspecific antigens for detection of latent infection in persons with recent tuberculosis contact.

The tuberculin skin test (TST) is used for the identification of latent tuberculosis (TB) infection (LTBI) but lacks specificity in Mycobacterium bovis BCG-vaccinated individuals, who constitute an increasing proportion of TB patients and their contacts from regions where TB is endemic. In previous studies, T-cell responses to ESAT-6 and CFP-10, M. tuberculosis-specific antigens that are absent from BCG, were sensitive and specific for detection of active TB. We studied 44 close contacts of a patient with smear-positive pulmonary TB and compared the standard screening procedure for LTBI by TST or chest radiographs with T-cell responses to M. tuberculosis-specific and nonspecific antigens. Peripheral blood mononuclear cells were cocultured with ESAT-6, CFP-10, TB10.4 (each as recombinant antigen and as a mixture of overlapping synthetic peptides), M. tuberculosis sonicate, purified protein derivative (PPD), and short-term culture filtrate, using gamma interferon production as the response measure. LTBI screening was by TST in 36 participants and by chest radiographs in 8 persons. Nineteen contacts were categorized as TST negative, 12 were categorized as TST positive, and 5 had indeterminate TST results. Recombinant antigens and peptide mixtures gave similar results. Responses to TB10.4 were neither sensitive nor specific for LTBI. T-cell responses to ESAT-6 and CFP-10 were less sensitive for detection of LTBI than those to PPD (67 versus 100%) but considerably more specific (100 versus 72%). The specificity of the TST or in vitro responses to PPD will be even less when the proportion of BCG-vaccinated persons among TB contacts evaluated for LTBI increases.

Adolescent↗
PubMed2001

Improved immunogenicity and protective efficacy of a tuberculosis DNA vaccine encoding Ag85 by protein boosting.

C57BL/6 mice were vaccinated with plasmid DNA encoding Ag85 from Mycobacterium tuberculosis, with Ag85 protein in adjuvant, or with a combined DNA prime-protein boost regimen. While DNA immunization, as previously described, induced robust Th1-type cytokine responses, protein-in-adjuvant vaccination elicited very poor cytokine responses, which were 10-fold lower than those observed with DNA immunization alone. Injection of Ag85 DNA-primed mice with 30 to 100 microg of purified Ag85 protein in adjuvant increased the interleukin-2 and gamma interferon (IFN-gamma) response in spleen two- to fourfold. Further, intracellular cytokine analysis by flow cytometry also showed an increase in IFN-gamma-producing CD4(+) T cells in DNA-primed-protein-boosted animals, compared to those that received only the DNA vaccination. Moreover, these responses appeared to be better sustained over time. Antibodies were readily produced by all three methods of immunization but were exclusively of the immunoglobulin G1 (IgG1) isotype following protein immunization in adjuvant and preferentially of the IgG2a isotype following DNA and DNA prime-protein boost vaccination. Finally, protein boosting increased the protective efficacy of the DNA vaccine against an intravenous M. tuberculosis H37Rv challenge infection, as measured by CFU or relative light unit counts in lungs 1 and 2 months after infection. The capacity of exogenously given protein to boost the DNA-primed vaccination effect underlines the dominant role of Th1-type CD4(+) helper T cells in mediating protection.

Acyltransferases↗
PubMed2001

Uncommon presentations of tuberculosis: the potential value of a novel diagnostic assay based on the Mycobacterium tuberculosis-specific antigens ESAT-6 and CFP-10.

SETTING: Leiden University Medical Center, Leiden, the Netherlands. OBJECTIVE: To illustrate the potential value of a recently developed diagnostic assay for detection of tuberculosis (TB), based on T cell responses to the early secreted antigenic target 6 kDa protein (ESAT-6) and culture filtrate protein 10 (CFP-10). These antigens are Mycobacterium tuberculosis specific because they are expressed by M. tuberculosis but absent from M. bovis bacille Calmette-Guérin (BCG) and most environmental mycobacteria. In recent studies, the assay had a high sensitivity and specificity for detection of active TB. DESIGN: We describe five patients with uncommon presentations of tuberculosis, in whom the diagnosis was delayed by negative or conflicting results of diagnostic procedures aimed at detection of M. tuberculosis and an uninformative tuberculin skin test. IFN-gamma production in response to ESAT-6 and CFP-10 by peripheral blood mononuclear cells from these patients was evaluated before and during anti-tuberculosis treatment. RESULTS: In all five patients, IFN-gamma responses to ESAT-6 and/or CFP-10 were above the cut-off level defined in a previous study. During treatment, IFN-gamma responses generally increased. CONCLUSION: These results indicate that T cell responses to M. tuberculosis-specific antigens have potential diagnostic value when TB is suspected and the results of other diagnostic tests are inconclusive, especially in BCG-vaccinated individuals.

Adult↗
PubMed2000

Identification of major epitopes of Mycobacterium tuberculosis AG85B that are recognized by HLA-A*0201-restricted CD8+ T cells in HLA-transgenic mice and humans.

CD8(+) T cells are thought to play an important role in protective immunity to tuberculosis. Although several nonprotein ligands have been identified for CD1-restricted CD8(+) CTLs, epitopes for classical MHC class I-restricted CD8(+) T cells, which most likely represent a majority among CD8(+) T cells, have remained ill defined. HLA-A*0201 is one of the most prevalent class I alleles, with a frequency of over 30% in most populations. HLA-A2/K(b) transgenic mice were shown to provide a powerful model for studying induction of HLA-A*0201-restricted immune responses in vivo. The Ag85 complex, a major component of secreted Mycobacterium tuberculosis proteins, induces strong CD4(+) T cell responses in M. tuberculosis-infected individuals, and protection against tuberculosis in Ag85-DNA-immunized animals. In this study, we demonstrate the presence of HLA class I-restricted, CD8(+) T cells against Ag85B of M. tuberculosis in HLA-A2/K(b) transgenic mice and HLA-A*0201(+) humans. Moreover, two immunodominant Ag85 peptide epitopes for HLA-A*0201-restricted, M. tuberculosis-reactive CD8(+) CTLs were identified. These CD8(+) T cells produced IFN-gamma and TNF-alpha and recognized Ag-pulsed or bacillus Calmette-Guérin-infected, HLA-A*0201-positive, but not HLA-A*0201-negative or uninfected human macrophages. This CTL-mediated killing was blocked by anti-CD8 or anti-HLA class I mAb. Using fluorescent peptide/HLA-A*0201 tetramers, Ag85-specific CD8(+) T cells could be visualized in bacillus Calmette-Guérin-responsive, HLA-A*0201(+) individuals. Collectively, our results demonstrate the presence of HLA class I-restricted CD8(+) CTL against a major Ag of M. tuberculosis and identify Ag85B epitopes that are strongly recognized by HLA-A*0201-restricted CD8(+) T cells in humans and mice. These epitopes thus represent potential subunit components for the design of vaccines against tuberculosis.

Acyltransferases↗
PubMed2000

[From gene to disease; mutations in interleukin-12-receptor-beta 1- and interferon-gamma-receptor-1 lead to nontuberculous mycobacterial infections and salmonellosis].

In patients with severe and relapsing infections with non-tuberculous mycobacteria and Salmonella autosomal recessive as well as dominant-negative mutations are shown in genes for receptors of type-1 cytokines: the IL-12 receptor and the IFN-gamma receptor. In case of an impaired capacity to produce these cytokines or--given an abnormal receptor--to react to them, the cellular immune reaction does not run a normal course and the susceptibility to infection by intracellular pathogens is enhanced.

Immune Tolerance↗
PubMed2000

Residual type 1 immunity in patients genetically deficient for interleukin 12 receptor beta1 (IL-12Rbeta1): evidence for an IL-12Rbeta1-independent pathway of IL-12 responsiveness in human T cells.

Genetic lack of interleukin 12 receptor beta1 (IL-12Rbeta1) surface expression predisposes to severe infections by poorly pathogenic mycobacteria or Salmonella and causes strongly decreased, but not completely abrogated, interferon (IFN)-gamma production. To study IL-12Rbeta1-independent residual IFN-gamma production, we have generated mycobacterium-specific T cell clones (TCCs) from IL-12Rbeta1-deficient individuals. All TCCs displayed a T helper type 1 phenotype and the majority responded to IL-12 by increased IFN-gamma production and proliferative responses upon activation. This response to IL-12 could be further augmented by exogenous IL-18. IL-12Rbeta2 was found to be normally expressed in the absence of IL-12Rbeta1, and could be upregulated by IFN-alpha. Expression of IL-12Rbeta2 alone, however, was insufficient to induce signal transducer and activator of transcription (Stat)4 activation in response to IL-12, whereas IFN-alpha/IFN-alphaR ligation resulted in Stat4 activation in both control and IL-12Rbeta1-deficient cells. IL-12 failed to upregulate cell surface expression of IL-18R, integrin alpha6, and IL-12Rbeta2 on IL-12Rbeta1-deficient cells, whereas this was normal on control cells. IL-12-induced IFN-gamma production in IL-12Rbeta1-deficient T cells could be inhibited by the p38 mitogen-activated protein kinase (MAP) kinase inhibitor SB203580 and the MAP kinase kinase (MEK) 1/2 inhibitor U0126, suggesting involvement of MAP kinases in this alternative, Stat4-independent, IL-12 signaling pathway.Collectively, these results indicate that IL-12 acts as a partial agonist in the absence of IL-12Rbeta1. Moreover, the results reveal the presence of a novel IL-12Rbeta1/Stat4-independent pathway of IL-12 responsiveness in activated human T cells involving MAP kinases. This pathway is likely to play a role in the residual type 1 immunity in IL-12Rbeta1 deficiency.

Adolescent↗
PubMed2000

Detection of active tuberculosis infection by T cell responses to early-secreted antigenic target 6-kDa protein and culture filtrate protein 10.

The purified protein derivative (PPD) skin test has no predictive value for tuberculosis (TB) in Mycobacterium bovis bacillus Calmette-Guérin (BCG)-vaccinated individuals because of cross-reactive responses to nonspecific constituents of PPD. T cell responses to early-secreted antigenic target 6-kDa protein (ESAT-6) and the newly identified culture filtrate protein 10 (CFP-10), 2 proteins specifically expressed by M. tuberculosis (MTB) but not by BCG strains, were evaluated. Most TB patients responded to ESAT-6 (92%) or CFP-10 (89%). A minority of BCG-vaccinated individuals responded to both ESAT-6 and CFP-10, their history being consistent with latent infection with MTB in the presence of protective immunity. No responses were found in PPD-negative controls. The sensitivity and specificity of the assay were 84% and 100%, respectively, at a cutoff of 300 pg of interferon-gamma/mL. These data indicate that ESAT-6 and CFP-10 are promising antigens for highly specific immunodiagnosis of TB, even in BCG-vaccinated individuals.

Adolescent↗
PubMed2000

Glucocorticoids transform CD40-triggering of dendritic cells into an alternative activation pathway resulting in antigen-presenting cells that secrete IL-10.

Dendritic cell (DC) activation through CD40-CD40 ligand interactions is a key regulatory step for the development of protective T-cell immunity and also plays an important role in the initiation of T-cell responses involved in autoimmune diseases and allograft rejection. In contrast to previous reports, we show that the immunosuppressive drug dexamethasone (DEX) redirects rather than simply blocks this DC activation process. We found that DCs triggered through CD40 in the presence of DEX were unable to acquire high levels of costimulatory, adhesion, and major histocompatibility complex class I and II molecules and failed to express the maturation marker CD83, whereas antigen uptake was not affected. Moreover, DEX strikingly modified the CD40-activated DC cytokine secretion profile by suppressing the production of the proinflammatory cytokine interleukin (IL)-12 and potentiating the secretion of the anti-inflammatory cytokine IL-10. Accordingly, DEX-exposed CD40-triggered DCs displayed a decreased T-cell allostimulatory potential and a dramatically impaired ability to activate cloned CD4(+) T helper 1 (Th1) cells. Moreover, interaction between Th1 cells and these DCs rendered the T cells hyporesponsive to further antigen-specific restimulation. Collectively, our results demonstrate that DEX profoundly modulates CD40-dependent DC activation and suggest that the resulting alternatively activated DCs can be exploited for suppression of unwanted T-cell responses in vivo.

Antigen Presentation↗