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1.
Elife ; 82019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31084714

RESUMO

Systems vaccinology approaches have been used successfully to define early signatures of the vaccine-induced immune response. However, the possibility that transcriptomics can also identify a correlate or surrogate for vaccine inflammation has not been fully explored. We have compared four licensed vaccines with known safety profiles, as well as three agonists of Toll-like receptors (TLRs) with known inflammatory potential, to elucidate the transcriptomic profile of an acceptable response to vaccination versus that of an inflammatory reaction. In mice, we looked at the transcriptomic changes in muscle at the injection site, the lymph node that drained the muscle, and the peripheral blood mononuclear cells (PBMCs)isolated from the circulating blood from 4 hr after injection and over the next week. A detailed examination and comparative analysis of these transcriptomes revealed a set of novel biomarkers that are reflective of inflammation after vaccination. These biomarkers are readily measurable in the peripheral blood, providing useful surrogates of inflammation, and provide a way to select candidates with acceptable safety profiles.

2.
EMBO Rep ; 20(4)2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30872316

RESUMO

Cyclic dinucleotides (CDNs) are important second messenger molecules in prokaryotes and eukaryotes. Within host cells, cytosolic CDNs are detected by STING and alert the host by activating innate immunity characterized by type I interferon (IFN) responses. Extracellular bacteria and dying cells can release CDNs, but sensing of extracellular CDNs (eCDNs) by mammalian cells remains elusive. Here, we report that endocytosis facilitates internalization of eCDNs. The DNA sensor cGAS facilitates sensing of endocytosed CDNs, their perinuclear accumulation, and subsequent STING-dependent release of type I IFN Internalized CDNs bind cGAS directly, leading to its dimerization, and the formation of a cGAS/STING complex, which may activate downstream signaling. Thus, eCDNs comprise microbe- and danger-associated molecular patterns that contribute to host-microbe crosstalk during health and disease.

3.
Artigo em Inglês | MEDLINE | ID: mdl-29624071

RESUMO

RATIONALE: Contacts of tuberculosis (TB) patients constitute an important target population for preventative measures as they are at high risk of infection with Mycobacterium tuberculosis and progression to disease. OBJECTIVES: We investigated biosignatures with predictive ability for incident tuberculosis. METHODS: In a case-control study nested within the Grand Challenges 6-74 longitudinal HIV-negative African cohort of exposed household contacts, we employed RNA sequencing, polymerase chain reaction (PCR) and the Pair Ratio algorithm in a training/test set approach. Overall, 79 progressors, who developed tuberculosis between 3 and 24 months following exposure, and 328 matched non-progressors, who remained healthy during 24 months of follow-up, were investigated. MEASUREMENTS AND MAIN RESULTS: A four-transcript signature (RISK4), derived from samples in a South African and Gambian training set, predicted progression up to two years before onset of disease in blinded test set samples from South Africa, The Gambia and Ethiopia with little population-associated variability and also validated on an external cohort of South African adolescents with latent Mycobacterium tuberculosis infection. By contrast, published diagnostic or prognostic tuberculosis signatures predicted on samples from some but not all 3 countries, indicating site-specific variability. Post-hoc meta-analysis identified a single gene pair, C1QC/TRAV27, that would consistently predict TB progression in household contacts from multiple African sites but not in infected adolescents without known recent exposure events. CONCLUSIONS: Collectively, we developed a simple whole blood-based PCR test to predict tuberculosis in household contacts from diverse African populations, with potential for implementation in national TB contact investigation programs.

4.
EMBO Mol Med ; 8(11): 1325-1339, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27729388

RESUMO

Accumulating evidence from experimental animal models suggests that antibodies play a protective role against tuberculosis (TB). However, little is known about the antibodies generated upon Mycobacterium tuberculosis (MTB) exposure in humans. Here, we performed a molecular and functional characterization of the human B-cell response to MTB by generating recombinant monoclonal antibodies from single isolated B cells of untreated adult patients with acute pulmonary TB and from MTB-exposed healthcare workers. The data suggest that the acute plasmablast response to MTB originates from reactivated memory B cells and indicates a mucosal origin. Through functional analyses, we identified MTB inhibitory antibodies against mycobacterial antigens including virulence factors that play important roles in host cell infection. The inhibitory activity of anti-MTB antibodies was directly linked to their isotype. Monoclonal as well as purified serum IgA antibodies showed MTB blocking activity independently of Fc alpha receptor expression, whereas IgG antibodies promoted the host cell infection. Together, the data provide molecular insights into the human antibody response to MTB and may thereby facilitate the design of protective vaccination strategies.


Assuntos
Anticorpos Antibacterianos/imunologia , Anticorpos Monoclonais/imunologia , Formação de Anticorpos , Linfócitos B/imunologia , Isotipos de Imunoglobulinas/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose Pulmonar/imunologia , Anticorpos Antibacterianos/isolamento & purificação , Anticorpos Monoclonais/isolamento & purificação , Humanos
5.
EMBO Mol Med ; 8(2): 86-95, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26682570

RESUMO

There is an urgent need for new tools to combat the ongoing tuberculosis (TB) pandemic. Gene expression profiles based on blood signatures have proved useful in identifying genes that enable classification of TB patients, but have thus far been complex. Using real-time PCR analysis, we evaluated the expression profiles from a large panel of genes in TB patients and healthy individuals in an Indian cohort. Classification models were built and validated for their capacity to discriminate samples from TB patients and controls within this cohort and on external independent gene expression datasets. A combination of only four genes distinguished TB patients from healthy individuals in both cross-validations and on separate validation datasets with very high accuracy. An external validation on two distinct cohorts using a real-time PCR setting confirmed the predictive power of this 4-gene tool reaching sensitivity scores of 88% with a specificity of around 75%. Moreover, this gene signature demonstrated good classification power in HIV(+) populations and also between TB and several other pulmonary diseases. Here we present proof of concept that our 4-gene signature and the top classifier genes from our models provide excellent candidates for the development of molecular point-of-care TB diagnosis in endemic areas.


Assuntos
Técnicas de Diagnóstico Molecular/métodos , Sistemas Automatizados de Assistência Junto ao Leito , Tuberculose/diagnóstico , Perfilação da Expressão Gênica/métodos , Humanos , Valor Preditivo dos Testes , Sensibilidade e Especificidade , Tuberculose/patologia
6.
BMC Genomics ; 16: 34, 2015 Feb 05.
Artigo em Inglês | MEDLINE | ID: mdl-25649146

RESUMO

BACKGROUND: The human pathogen Mycobacterium tuberculosis has the capacity to escape eradication by professional phagocytes. During infection, M. tuberculosis resists the harsh environment of phagosomes and actively manipulates macrophages and dendritic cells to ensure prolonged intracellular survival. In contrast to other intracellular pathogens, it has remained difficult to capture the transcriptome of mycobacteria during infection due to an unfavorable host-to-pathogen ratio. RESULTS: We infected the human macrophage-like cell line THP-1 with the attenuated M. tuberculosis surrogate M. bovis Bacillus Calmette-Guérin (M. bovis BCG). Mycobacterial RNA was up to 1000-fold underrepresented in total RNA preparations of infected host cells. We employed microbial enrichment combined with specific ribosomal RNA depletion to simultaneously analyze the transcriptional responses of host and pathogen during infection by dual RNA sequencing. Our results confirm that mycobacterial pathways for cholesterol degradation and iron acquisition are upregulated during infection. In addition, genes involved in the methylcitrate cycle, aspartate metabolism and recycling of mycolic acids were induced. In response to M. bovis BCG infection, host cells upregulated de novo cholesterol biosynthesis presumably to compensate for the loss of this metabolite by bacterial catabolism. CONCLUSIONS: Dual RNA sequencing allows simultaneous capture of the global transcriptome of host and pathogen, during infection. However, mycobacteria remained problematic due to their relatively low number per host cell resulting in an unfavorable bacterium-to-host RNA ratio. Here, we use a strategy that combines enrichment for bacterial transcripts and dual RNA sequencing to provide the most comprehensive transcriptome of intracellular mycobacteria to date. The knowledge acquired into the pathogen and host pathways regulated during infection may contribute to a solid basis for the deployment of novel intervention strategies to tackle infection.


Assuntos
Colesterol/biossíntese , Interações Hospedeiro-Patógeno/genética , Mycobacterium tuberculosis/genética , Tuberculose/genética , Animais , Bovinos , Colesterol/genética , Células Dendríticas/metabolismo , Células Dendríticas/microbiologia , Regulação Bacteriana da Expressão Gênica/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Macrófagos/microbiologia , Mycobacterium bovis/patogenicidade , Mycobacterium tuberculosis/patogenicidade , Fagócitos/metabolismo , Fagócitos/microbiologia , Transcriptoma/efeitos dos fármacos , Tuberculose/microbiologia
7.
Expert Rev Vaccines ; 13(5): 619-30, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24702486

RESUMO

Tuberculosis remains a major health threat and vaccines better than bacillus Calmette-Guérin (BCG) are urgently required. Here we describe our experience with a recombinant BCG expressing listeriolysin and deficient in urease. This potential replacement vaccine has demonstrated superior efficacy and safety over BCG in Mycobacterium tuberculosis aerosol-challenged mice and was safe in numerous animal models including immune-deficient mice, guinea pigs, rabbits and nonhuman primates. Phase I clinical trials in adults in Germany and South Africa have proven safety and a current Phase IIa trial is under way to assess immunogenicity and safety in its target population, newborns in a high tuberculosis incidence setting, with promising early results. Second-generation candidates are being developed to improve safety and efficacy.


Assuntos
Vacina BCG/administração & dosagem , Ensaios Clínicos como Assunto/tendências , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose/prevenção & controle , Animais , Humanos , Mycobacterium tuberculosis/fisiologia , Tuberculose/epidemiologia
8.
Expert Rev Mol Diagn ; 13(6): 625-37, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23895131

RESUMO

The success of our immune system depends on its ability to react efficiently, which in turn is supported by a large degree of plasticity as well as memory. Some aspects of this plasticity and memory are now known to be under epigenetic control - determined both by default, during differentiation, and by responses to environmental factors, including infectious agents. Thus, epigenetic marks in the immune system can occur as predetermined or as responsive marks and as such can potentially serve as diagnostic markers for disease susceptibility and disease progression or treatment response. Here, the authors review some examples of epigenetic control and epigenetic marks during the differentiation process of the immune system and memory formation, followed by some examples of epigenetic marks in the immune system subsequent to infection. These are used to illustrate the potential use of epigenetic marks as diagnostic markers in adverse immune system conditions and treatment thereof.


Assuntos
Biomarcadores , Metilação de DNA/genética , Epigênese Genética , Patologia Molecular , Diferenciação Celular/genética , Reprogramação Celular/genética , Histonas/metabolismo , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Imunidade/genética , Infecção/genética , Infecção/metabolismo
9.
Curr Opin Immunol ; 17(1): 79-87, 2005 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-15653315

RESUMO

Antigen processing and recognition is a key feature of antibacterial immune responses to intracellular bacteria. In contrast to viruses, which are primarily controlled by conventional MHC II- and MHC I-restricted CD4+ or CD8+ T cells, respectively, unconventional T cells participate additionally in antibacterial protection. These unconventional T cells include glycolipid-specific CD1-restricted T cells and phospholigand-specific gammadelta T cells. We are just beginning to understand the broad spectrum of antigen recognition and stimulation of distinct T-cell populations by bacterial pathogens. From the host perspective, a broad spectrum of different T-cell populations that recognize proteins, lipids and carbohydrates strengthens protective immunity. From the perspective of the pathogen, antigen presentation represents a bottleneck that should be exploited for evasion from, or devastation of, acquired immunity. Although several such mechanisms have been described in viral systems, few have thus far been elucidated in bacterial infections.


Assuntos
Apresentação do Antígeno/imunologia , Infecções Bacterianas/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Antígenos CD1/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Linfócitos T/imunologia
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