Assessment ofin vitroimmunity toM ycobacterium tuberculosisin a human peripheral blood infection model using a luciferase reporter construct ofM. tuberculosisH37Rv

Shaban

2010

Med Princ Pract

Objective: To identify T helper 1 (Th1)-cell stimulating and HLA-promiscuous peptides of MPT64 (Rv1980c), a major secreted antigen of Mycobacterium tuberculosis. Materials and Methods: Peripheral blood mononuclear cells (PBMCs) were obtained from 35 healthy subjects and typed for HLA-DR molecules using genomic methods. To identify subjects infected with M. tuberculosis, PBMCs were tested in antigen-induced proliferation assays with whole cells and culture filtrate antigens of M. tuberculosis, M. tuberculosis-specific antigens ESAT-6 and CFP10, and MPT64. Culture filtrate-induced T-cell lines were established in vitro from 12 M. tuberculosis-infected and HLA-heterogeneous healthy subjects, and tested with 20 overlapping synthetic peptides covering the sequence of MPT64 in Th1-cell assays, i.e. antigen-induced proliferation and/or IFN-γ secretion. In addition, T-cell lines from three HLA-heterogeneous subjects were tested for cytotoxic activity against peptide-pulsed antigen-presenting cells. Results: PBMCs from 12 of 35 subjects responded to M. tuberculosis-specific antigens ESAT-6 and CFP10 as well as to MPT64, which suggested that they were infected with M. tuberculosis. Ten of twelve T-cell lines established from these donors responded to MPT64, and nine T-cell lines responded to 1 or more of the peptides of MPT64 in anti- gen-induced proliferation assays. Furthermore, 18 of the 20 peptides of MPT64 were recognized by the T-cell lines in 1 or more assay systems, and at least 5 peptides were recognized by T-cell lines from HLA-DR-heterogeneous subjects. Conclusion: Th1-cell-reactive epitopes are scattered throughout the sequence of MPT64, and at least 5 of its peptides are presented to Th1-cells in a HLA-promiscuous manner.

Section: Introductionmentioning

confidence: 99%

Mapping of Th1-Cell Epitope Regions of Mycobacterium tuberculosis Protein MPT64 (Rv1980c) Using Synthetic Peptides and T-Cell Lines from M. tuberculosis-Infected Healthy Humans

Shaban

2010

Med Princ Pract

“…The protection against TB requires cellular immune responses mediated by T helper 1 (Th1)-type cells that secrete large quantities of gamma interferon (IFN-␥) (2,13,15), and therefore a primary criterion for selecting candidate antigens for vaccine design has been their ability to induce IFN-␥ responses (reviewed in reference 27). M. tuberculosis is rich in antigens that induce IFN-␥ secretion, and the presence of such antigens has been reported in purified cell walls, the cytosolic fraction, and short-term culture filtrates (ST-CF) (reviewed in reference 25).…”

mentioning

confidence: 99%

Efficient Testing of Large Pools of Mycobacterium tuberculosis RD1 Peptides and Identification of Major Antigens and Immunodominant Peptides Recognized by Human Th1 Cells

Al‐Attiyah

Hanif

et al. 2008

Clin Vaccine Immunol

Tuberculosis (TB) is among the most important diseases of worldwide distribution with respect to both morbidity and mortality. Annually, TB afflicts about 9 million people with 2 million deaths (18). The rapid spread of TB in developing countries of Africa and Asia is accelerated by the human immunodeficiency virus epidemic and the spread of multi-and extra-drug-resistant TB (46). To protect against TB, vaccination with Mycobacterium bovis BCG has been used for more than 70 years, but its efficacy varies tremendously in different parts of the world (49). In addition, the diagnostic value of the presently used skin test reagent, the purified protein derivative (PPD) of M. tuberculosis, is low due to cross-reactivity with environmental mycobacteria and the vaccine strains of M. bovis BCG (14). Furthermore, vaccination with M. bovis BCG is contraindicated in immunocompromised subjects, including AIDS patients who are usually at a very high risk of developing TB (19). Identification of M. tuberculosis antigens useful for diagnosis and development of new vaccines is therefore an important goal in TB research.The protection against TB requires cellular immune responses mediated by T helper 1 (Th1)-type cells that secrete large quantities of gamma interferon (IFN-␥) (2, 13, 15), and therefore a primary criterion for selecting candidate antigens for vaccine design has been their ability to induce IFN-␥ responses (reviewed in reference 27). M. tuberculosis is rich in antigens that induce IFN-␥ secretion, and the presence of such antigens has been reported in purified cell walls, the cytosolic fraction, and short-term culture filtrates (ST-CF) (reviewed in reference 25). However, several studies have suggested that antigens present in ST-CF are the primary inducers of IFN-␥ secretion and provide protection against TB in mice and guinea pigs (reviewed in reference 26). It has been previously shown that two M. tuberculosis-specific antigens present in ST-CF, i.e., ESAT6 and CFP10, are frequently recognized by human Th1 cells after natural infection and therefore are important in the specific diagnosis of active and latent TB (29,31,47). Interestingly, ESAT6 and CFP10, when tested together in IFN-␥ assays, have been shown to improve the diagnostic efficacy of TB compared to results with each antigen separately (20). In addition, vaccination with ESAT6 and CFP10 has been shown to provide protection against challenge with M. tuberculosis in animal models of TB (50). However, ESAT6 and CFP10 can be used either as vaccines or as diagnostic reagents but not as both. This necessitates the identification of additional M. tuberculosis antigens, some of which could be reserved for diagnosis and others for the development of new vaccines against TB.The analysis of the M. tuberculosis genome sequence has shown that genes encoding ESAT6 and CFP10 are located in region of difference 1 (RD1) that is deleted in all vaccine strains of M. bovis BCG but present in all of the tested strains and isolates of pathogenic M. tuberculosis and M. bov...

“…Protective CMI primarily involves interferon (IFN)-g release by antigen-activated CD4 1 T-helper (Th) type 1 cells, which activates macrophages to destroy intracellular mycobacteria (Flynn, 2004). The central role of IFN-g in the protection against TB has been suggested by many studies in both animals and humans (Flynn, 2004;Al-Attiyah et al, 2006a;Dietrich et al, 2006;Mustafa et al, 2006). On the other hand, the Th2 responses, characterized by the secretion of interleukin (IL)-4, IL-5 and anti-inflammatory cytokine IL-10, are associated with a lack of protection (Bai et al, 2004;Flynn, 2004).…”

Section: Introductionmentioning

confidence: 99%

Characterization of human cellular immune responses toMycobacterium tuberculosisproteins encoded by genes predicted in RD15 genomic region that is absent inMycobacterium bovisBCG

Al‐Attiyah

FEMS Immunol Med Microbiol

2010

RD15 is a genomic region of difference (RD) present in Mycobacterium tuberculosis H37Rv but absent in all strains of Mycobacterium bovis BCG. RD15 contains genes encoding proteins of mammalian cell entry (Mce3A-F), important for the invasion and survival of M. tuberculosis in host cells. In this study, we have evaluated cellular immune responses to RD15 proteins using peripheral blood mononuclear cells (PBMC) from pulmonary tuberculosis patients and M. bovis BCG-vaccinated healthy subjects. PBMC were tested for T-helper (Th) type 1 [antigen-induced proliferation and interferon (IFN)-gamma secretion] and anti-inflammatory [interleukin (IL)-10 secretion] responses to complex mycobacterial antigens and peptides corresponding to proteins of RD1 and RD15. In Th1 assays, complex mycobacterial antigens induced strong responses in both donor groups, and RD1 induced strong responses in tuberculosis patients and moderate responses in healthy subjects, whereas RD15 induced weak responses in tuberculosis patients and strong to moderate responses in healthy subjects. IL-10 secretion in both donor groups was strong to moderate in response to complex mycobacterial antigens, but weak in response to RD1 and RD15. Analysis of IFN-gamma : IL-10 ratios showed strong Th1 biases to complex mycobacterial antigens and RD1 in both donor groups, and to RD15 and RD1504 (Mce3A) in healthy subjects only. These results suggest that RD1504 is the best Th1-stimulating antigen present in RD15, and therefore may be a potential vaccine candidate against TB.