Use of ESAT-6 and CFP-10 antigens for diagnosis of extrapulmonary tuberculosis.
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Antigens, Bacterial↗
Biomedical subjects
M E Munk
Publications and source records attributed to M E Munk.
Performance of whole blood IFN-gamma test for tuberculosis diagnosis based on PPD or the specific antigens ESAT-6 and CFP-10.
OBJECTIVE: To evaluate the QuantiFERON-TB test in BCG-vaccinated, non-BCG-vaccinated and tuberculosis (TB) patient donor groups, and to compare its diagnostic performance with that of a blood test based on the Mycobacterium tuberculosis specific antigens ESAT-6 and CFP-10. DESIGN: Analysis of the IFN-gamma responses of whole blood cells from BCG-vaccinated or non-BCG-vaccinated donors or patients with tuberculosis, stimulated with PPD, ESAT-6 or CFP-10 antigens, and evaluation of the specificity and sensitivity of the test. RESULTS: None of the non-vaccinated donors showed positive responses to M. tuberculosis-PPD, ESAT-6 or CFP-10. In BCG-vaccinated donors, 9/19 (47%) donors responded to the QuantiFERON-TB test based on M. tuberculosis-PPD, whereas 2/19 (10.5%) responded to either ESAT-6 or CFP-10. Comparable levels of sensitivity were obtained with the QuantiFERON-TB test based on M. tuberculosis-PPD (79%) and ESAT-6 or CFP-10 antigens (72%). CONCLUSION: Our results demonstrate that the whole blood test based on M. tuberculosis-PPD did not efficiently distinguish BCG-vaccinated donors from individuals with disease due to M. tuberculosis. The introduction of new recombinant antigens specific for M. tuberculosis, such as ESAT-6 or CFP-10, should increase the specificity of the whole blood test and enable discrimination between TB infection, atypical mycobacterial reactivity and reactivity due to BCG vaccination. Such a test would provide a quantum improvement over the current practice of using the tuberculin skin test for TB control and elimination.
Adult↗
Specific immune-based diagnosis of tuberculosis.
Current diagnostic tests for tuberculosis based on tuberculin have poor specificity, and both BCG vaccination and exposure to non-tuberculosis mycobacteria produce a response similar to that induced by infection with Mycobacterium tuberculosis. The identification of regions of the M. tuberculosis genome that are not present in BCG and non-tuberculous mycobacteria provides a unique opportunity to develop new highly specific diagnostic reagents. We describe the current status of attempts to exploit this information and summarise recent research that has used defined antigens for an accurate and rapid test for tuberculosis infection based on the detection of T cells sensitised to M. tuberculosis either by blood tests in vitro or skin tests in vivo.
Antigens, Bacterial↗
Increased numbers of ESAT-6- and purified protein derivative-specific gamma interferon-producing cells in subclinical and active tuberculosis infection.
Numbers of gamma interferon (IFN-gamma)-producing cells reactive to ESAT-6 antigen were increased in recent converters to purified protein derivative positivity and in tuberculosis patients but not in unvaccinated or Mycobacterium bovis BCG-vaccinated healthy donors. ESAT-6-reactive IFN-gamma-producing cells in recent converters and tuberculosis patients recognized similar synthetic peptides. Thus, ESAT-6 is a potential candidate for use in detection of early, as well as active, tuberculosis and for control of the disease.
Amino Acid Sequence↗
Numbers of IFN-gamma-producing cells against ESAT-6 increase in tuberculosis patients during chemotherapy.
ESAT-6 is a specific Mycobacterium tuberculosis complex antigen and strong inducer of interferon-gamma (IFN-gamma) production by T cells from tuberculosis patient T-cells. We studied the frequency of IFN-gamma producing cells reacting to ESAT-6 during anti-tuberculosis chemotherapy. The numbers of IFN-gamma producing cells in the peripheral blood were higher in tuberculosis patients after discharge from specific anti-tuberculosis chemotherapy, compared with untreated patients. These results indicate that monitoring specific M. tuberculosis antigen reactivity during anti-tuberculosis chemotherapy may avoid premature termination of treatment and resistant strains.
Adult↗
Phenotypically activated gammadelta T lymphocytes in the peripheral blood of patients with tuberculosis.
Surface molecules with the potential relevance for resistance against Mycobacterium tuberculosis were investigated. The expression of lymphocyte function antigen-1, very late antigen (VLA)-4, l-selectin, intercellular adhesion molecule (ICAM)-1, major histocompatibility complex class II, Fas, and CD40 on alphabeta T cells, gammadelta T cells, NK cells, and monocytes of healthy donors and patients with tuberculosis were analyzed. A high activation status of gammadelta T cells and increased levels of soluble ICAM-1 in plasma of patients with tuberculosis versus healthy individuals was detected. Tuberculosis patients with and without an underlying systemic disease could be segregated by differential expression of VLA-4 and ICAM-1 on gammadelta T cells and on monocytes. The composition of peripheral blood mononuclear cells varied slightly, whereas the proportion of monocytes decreased significantly in patients with tuberculosis, compared with healthy controls. The activation phenotype of peripheral gammadelta T cells in patients with tuberculosis emphasizes the role of these T cells in controlling the inflammatory process during tuberculosis and perhaps other microbial infections.
Adolescent↗
Soluble tumor necrosis factor alpha receptors in sera from leprosy patients.
Serum levels of soluble tumor necrosis factor alpha receptor I (sTNF-RI) were elevated in patients with lepromatous (LL) reactional-state type II leprosy, and sTNF-RII levels were increased in patients with full tuberculoid (TT) or LL type II leprosy. The sTNF-R in sera from patients with type II leprosy, but not other forms of leprosy, inhibited recombinant TNF cytolytic activities in vitro. This suggests that sTNF-R regulatory activities are partially impaired in patients with leprosy.
Adult↗
Differential T cell responses to Mycobacterium tuberculosis ESAT6 in tuberculosis patients and healthy donors.
Vaccination against and diagnosis of tuberculosis are still insufficient. Proteins secreted by Mycobacterium tuberculosis induce strong immune responses in tuberculosis and constitute prime candidates for development of novel vaccines against tuberculosis as well as for immunodiagnostic assays. We investigated the role of the secreted proteins MPT63, MPT64 and ESAT6 from M. tuberculosis in healthy individuals and tuberculosis patients. None of the secreted proteins stimulated peripheral blood mononuclear cells from healthy donors. In contrast, CD4+ T cells from many tuberculosis patients were stimulated in an MHC class II-restricted fashion by ESAT6, but not by MPT63 or MPT64. T cell reactivities of tuberculosis patients were focused on the N-terminal region of ESAT6. The ESAT6 T cell epitopes were presented by different HLA-DR phenotypes. Cell cultures responding to either ESAT6 or synthetic peptides thereof showed mRNA transcripts for macrophage inflammatory protein (MIP)-1alpha, monocyte chemotactic protein (MCP)-1 or IL-8 and production of IFN-gamma and MIP-1alpha. Our results suggest that the secreted M. tuberculosis proteins MPT63, MPT64 or ESAT6 do not stimulate unprimed T cells, and that ESAT6 may be a potential candidate antigen for detection of clinical disease.
Adult↗
Human CD30+ cells are induced by Mycobacterium tuberculosis and present in tuberculosis lesions.
CD30 is a member of the tumor necrosis factor/nerve growth factor receptor family and evidence has been presented that activated CD4+ CD45Ro+ T cells of Th2 type selectively express CD30. Mycobacterium tuberculosis, a facultative intracellular bacterium capable of replicating in resting macrophages, is a potent inducer of IFN-gamma secretion by Th1 cells. We find increased CD30 expression by M. tuberculosis-stimulated alpha beta and gamma delta T cells, and elevated numbers of CD30+ alpha beta T cells in tuberculosis pleuritis and affected lung tissue. Furthermore, surface CD30 was associated with intracytoplasmic IFN-gamma expression and IFN-gamma production by M. tuberculosis-stimulated alpha beta and gamma delta T cells. Thus, our results indicate that M. tuberculosis is a potent inducer of CD30 expression in Th1 cells and argue against exclusive correlation of CD30 expression with Th2 cell responses.
Animals↗
The superantigen exfoliative toxin induces cutaneous lymphocyte-associated antigen expression in peripheral human T lymphocytes.
Several immune-mediated dermatoses including psoriasis and atopic dermatitis can be exacerbated by bacterial infections. Superantigen producing bacteria can be isolated from skin lesions of these dermatoses. Consistent with superantigen effects, skewed T cell receptor variable gene usage has been demonstrated within these lesions. Therefore, the question arises whether superantigen induce a skin-seeking phenotype within peripheral T cells. In this study, we investigated the in vitro influence of the V beta 2-selective superantigen exfoliative toxin from Staphylococcus aureus on the expression of the cutaneous lymphocyte-associated antigen on peripheral T lymphocytes of healthy donors. We demonstrate that exfoliative toxin dramatically upregulates cutaneous lymphocyte-associated antigen expression on T cell receptor V beta 2+ lymphocytes. Up to 69% of V beta 2+ lymphocytes expressed cutaneous lymphocyte-associated antigen after 5 days of in vitro culture. Additionally, exfoliative toxin also increased cutaneous lymphocyte-associated antigen expression in CD3+ T cell receptor V beta 2- lymphocytes indicating a different effect as caused by the superantigen-T cell receptor V beta 2 interaction. Our findings suggest influence of bacterial superantigens on T lymphocyte skin homing in vivo.
Antigens, Differentiation, T-Lymphocyte↗
Human gamma/delta T-cell response to Listeria monocytogenes protein components in vitro.
Listeria monocytogenes is a facultative intracellular pathogen that replicates inside mononuclear phagocytes and induces specific cellular immunity. Listeriosis encompasses many clinical syndromes and meningitis is the most frequent clinical manifestation. Human alpha/beta and gamma/delta T cells have been shown to respond to L. monocytogenes antigens and to play an important role in resistance against listerial infection. We investigated the nature of listerial ligands and the influence of the major virulence factor, listeriolysin (hly), on the stimulation of human gamma/delta T cells from healthy individuals. We found that a listerial somatic protein ligand, which is sensitive to proteinase treatment, stimulated gamma/delta T cells in vitro; the majority of Listeria-responsive gamma/delta T cells expressed V gamma 9V delta 2 T-cell receptor chains and human leucocyte antigen-DR molecules; gamma/delta T-cell responses to hly+ and hly- Listeria strains were comparable; L. monocytogenes strains of different virulence stimulated gamma/delta T cells equally. Thus, protein components of L. monocytogenes unrelated to virulence activate human gamma/delta T cells in vitro.
Bacterial Proteins↗
Low-molecular-weight protein ligands from Onchocerca volvulus preferentially stimulate the human gammadelta T cell Vdelta1+ subset.
Onchocerciasis is a chronic infectious disease caused by the filarial nematode Onchocerca volvulus. A minor population of human gammadelta T cells expressing Vdelta1 chains is preferentially stimulated by O. volvulus ligands in vitro. Therefore, the nature of the parasite ligand and the effector functions of Vdelta1+ T cells stimulated by O. volvulus was investigated. A 5- to 30-kDa ligand from the adult parasite lysate that is sensitive to proteinase treatment was identified. Presentation for preferential stimulation of Vdelta1+ T cells required processing. After in vitro stimulation with O. volvulus in the presence of interleukin-2, Vdelta1+ T cells produced interferon-gamma but not interleukin-4 and exhibited NK cytolytic activities. It is concluded that somatic 5- to 30-kDa protein ligands from O. volvulus stimulate Vdelta1+ T cells and that Vdelta1+ T cells play a role in immunity to O. volvulus.
Animals↗
Increased numbers of interleukin-12-producing cells in human tuberculosis.
Numbers of interleukin-12 (IL-12) producing cells were quantitated in the peripheral blood of healthy donors and tuberculosis patients by the ELISPOT assay. We observed that (i) stimulation with mycobacteria increases numbers of IL-12 producers from healthy donors and (ii) tuberculosis patients have larger numbers of IL-12 producers than healthy donors. Our data emphasize the importance of Il-12 in immunity to tuberculosis.
Adult↗
Frequencies of IFN gamma- and IL-4-producing cells during Mycobacterium bovis BCG infection in two genetically susceptible mouse strains: role of alpha/beta T cells and NK1.1 cells.
Frequencies of IFN gamma- and IL-4-producing spleen cells in response to Mycobacterium bovis BCG infection were determined in C57BL/6 and BALB/c mice. Both mouse strains express equal innate susceptibility to M. bovis BCG (Bcgs), but differ in their NK1.1 and T-cell activities. M. bovis BCG infection induced higher frequencies (f approximately 1/500) of antigen-induced IFN gamma-secreting spleen cells in C57BL/6 mice as compared to BALB/c mice (f approximately 1/8000). Concanavalin A stimulated almost equal numbers of IFN gamma-secreting cells in both mouse strains (f approximately 1/50). Treatment with anti-NK1.1 mAb of M. bovis BCG-infected C57BL/6 mice did not alter frequencies of IFN gamma-secreting cells. Equally low numbers of antigen-induced IL-4-producing cells (f approximately 1/3000) were determined in both C57BL/6 and BALB/c mice during M. bovis BCG infection and treatment of C57BL/6 mice with anti-NK1.1 mAb had no measurable effect on IL-4 producers. Finally, frequencies of IFN gamma-producing cells were markedly reduced (10-fold) in M. bovis BCG-infected TCR-beta-/- gene deletion mutants as compared to their heterozygous controls. Our findings verify that M. bovis BCG infection primarily induces IFN gamma-secreting alpha/beta T cells of TH1 type and show that the frequencies of these IFN gamma producers differ in the two Bcgs mouse strains C57BL/6 and BALB/c.
Animals↗
Paracoccidioides brasiliensis-stimulated human gamma/delta T cells support antibody production by B cells.
Paracoccidioidomycosis patients show hyperactive humoral immune responses. Consequently, we investigated whether cytokines in supernatants from Paracoccidioides brasiliensis-stimulated gamma/delta T cells support B-cell activation. We detected proliferation of B cells and increased immunoglobulin M (IgM) and IgG production. Thus, gamma/delta T cells may participate in polyclonal B-cell activation during paracoccidioidomycosis.
Antibodies, Fungal↗
Human cord blood T-cell receptor alpha beta cell responses to protein antigens of Paracoccidioides brasiliensis yeast forms.
Paracoccidioides brasiliensis causes a chronic granulomatous mycosis, prevalent in South America, and cell-mediated immunity represents the principal mode of protection against this fungal infection. We investigated the response of naive cord blood T cells to P. brasiliensis lysates. Our results show: (1) P. brasiliensis stimulates T-cell expansion, interleukin-2 (IL-2) production and differentiation into cytotoxic T cells; (2) T-cell stimulation depends on P. brasiliensis processing and major histocompatibility complex (MHC) class II expression; (3) the responsive T-cell population expresses alpha beta T-cell receptors (TCR) with different V beta gene products, CD4 and CD45RO; (4) the P. brasiliensis components involved in T-cell expansion primarily reside in a high molecular weight (100,000 MW) and a low molecular weight (< 1000 MW) protein fraction. These results indicate that protein antigens of P. brasiliensis stimulate cord blood CD4 alpha beta T cells, independent from in vivo presensitization, and thus question direct correlation of positive in vitro responses with protective immunity in vivo.
Antigens, Fungal↗
Functions of T-cell subsets and cytokines in mycobacterial infections.
Phagocytosed M. tuberculosis either multiply inside the endocytic compartment of mononuclear phagocytes or they are destroyed by the host cell. Due to this macrophage-shelter (ab)used by mycobacteria, tuberculosis is controlled by the cellular immune response. Protection against mycobacteria depends on alpha/beta T-cells expressing the CD4 or CD8 phenotype. T-cell-mediated immunity amplifies macrophage capacities to kill and digest the bacilli. Specific alpha/beta T-cells produce several cytokines that attract and activate macrophages and additional lymphocytes, such as: interferon-gamma (IFN-gamma) which has the capacity to activate several antimicrobial properties of macrophages; tumour necrosis factor-alpha (TNF-alpha) a key cytokine involved in granuloma formation; interleukins 2, 6 and 8 (IL-2; IL-6 and IL-8); and interleukin 12 (IL-12), a candidate cytokine for the induction of Th1 cells. Furthermore, CD4+ and CD8+ T-cells display cytotoxic activity, which permits them to control mycobacterial growth through destruction of the infected cells. Escaping bacteria are subsequently ingested and destroyed by surrounding macrophages activated by T-cells. There is evidence to associate gamma/delta T-cells with antimycobacterial immunity, such as their preferential accumulation in inflammatory lesions, in necrotic areas of tuberculous lymphadenitis, and potent in vitro stimulation by M. tuberculosis components. In addition, M. tuberculosis activated gamma/delta T-cells are cytolytic and secrete several cytokines. Hence, clinical tuberculosis is associated with T-cell reactivity which controls the local concentrations of tubercle bacilli. Taken together, the cellular response, cytokine regulation, and the definition of target molecules are important aspects for the understanding of pathological immune mechanisms in tuberculosis.
CD4-Positive T-Lymphocytes↗
Prevention of autoimmune lysis by T cells with specificity for a heat shock protein by antisense oligonucleotide treatment.
T lymphocytes with specificity for the bacterial heat shock protein (hsp) 60 recognize stressed host cells, thus possibly promoting pathogenesis of certain infectious and autoimmune diseases. Here, we show that autoimmune destruction of stressed Schwann cells and macrophages by cytotoxic T lymphocytes raised against mycobacterial hsp60 can be inhibited by the use of hsp60-specific antisense oligodeoxynucleotides (A-ODNs). The inhibitory effect of hsp60 A-ODNs was specific because lysis of murine cytomegalovirus-infected host cells by virus-specific cytotoxic lymphocytes was not affected. Immunoblot analysis and immunoprecipitation studies suggest that different forms of stress increase hsp60 synthesis in Schwann cells and that this neosynthesis is reduced by hsp60 A-ODNs. These findings (i) provide evidence for participation of endogenous hsp60 in the recognition of stressed host cells by mycobacterial hsp60-crossreactive T cells and (ii) suggest the feasibility of inhibiting autoimmune reactions by target-cell treatment with specific A-ODNs.
Animals↗