RESUMEN
Immune activation is associated with increased risk of tuberculosis (TB) disease in infants. We performed a case-control analysis to identify drivers of immune activation and disease risk. Among 49 infants who developed TB disease over the first 2 years of life, and 129 healthy matched controls, we found the cytomegalovirus-stimulated (CMV-stimulated) IFN-γ response to be associated with CD8+ T cell activation (Spearman's rho, P = 6 × 10-8). A CMV-specific IFN-γ response was also associated with increased risk of developing TB disease (conditional logistic regression; P = 0.043; OR, 2.2; 95% CI, 1.02-4.83) and shorter time to TB diagnosis (Log Rank Mantel-Cox, P = 0.037). CMV+ infants who developed TB disease had lower expression of NK cell-associated gene signatures and a lower frequency of CD3-CD4-CD8- lymphocytes. We identified transcriptional signatures predictive of TB disease risk among CMV ELISpot-positive (area under the receiver operating characteristic [AUROC], 0.98, accuracy, 92.57%) and -negative (AUROC, 0.9; accuracy, 79.3%) infants; the CMV- signature was validated in an independent infant study (AUROC, 0.71; accuracy, 63.9%). A 16-gene signature that previously identified adolescents at risk of developing TB disease did not accurately classify case and control infants in this study. Understanding the microbial drivers of T cell activation, such as CMV, could guide new strategies for prevention of TB disease in infants.
Asunto(s)
Infecciones por Citomegalovirus/complicaciones , Infecciones por Citomegalovirus/inmunología , Tuberculosis/complicaciones , Tuberculosis/inmunología , Vacuna BCG , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Estudios de Casos y Controles , Citomegalovirus , Femenino , Humanos , Lactante , Inflamación , Interferón gamma/genética , Interferón gamma/metabolismo , Células Asesinas Naturales/inmunología , Activación de Linfocitos , Masculino , Mycobacterium tuberculosis , Factores de Riesgo , Sudáfrica , TranscriptomaRESUMEN
Non-lymphoid tissues (NLTs) harbor a pool of adaptive immune cells with largely unexplored phenotype and development. We used single-cell RNA-seq to characterize 35,000 CD4+ regulatory (Treg) and memory (Tmem) T cells in mouse skin and colon, their respective draining lymph nodes (LNs) and spleen. In these tissues, we identified Treg cell subpopulations with distinct degrees of NLT phenotype. Subpopulation pseudotime ordering and gene kinetics were consistent in recruitment to skin and colon, yet the initial NLT-priming in LNs and the final stages of NLT functional adaptation reflected tissue-specific differences. Predicted kinetics were recapitulated using an in vivo melanoma-induction model, validating key regulators and receptors. Finally, we profiled human blood and NLT Treg and Tmem cells, and identified cross-mammalian conserved tissue signatures. In summary, we describe the relationship between Treg cell heterogeneity and recruitment to NLTs through the combined use of computational prediction and in vivo validation.
Asunto(s)
Adaptación Fisiológica/inmunología , Análisis de la Célula Individual/métodos , Linfocitos T Reguladores/inmunología , Transcriptoma/inmunología , Adaptación Fisiológica/genética , Animales , Línea Celular Tumoral , Movimiento Celular/genética , Movimiento Celular/inmunología , Colon/inmunología , Colon/metabolismo , Humanos , Memoria Inmunológica/genética , Memoria Inmunológica/inmunología , Tejido Linfoide/inmunología , Tejido Linfoide/metabolismo , Ratones Transgénicos , Neoplasias Experimentales/genética , Neoplasias Experimentales/inmunología , Neoplasias Experimentales/patología , Piel/inmunología , Piel/metabolismo , Bazo/inmunología , Bazo/metabolismo , Linfocitos T Reguladores/metabolismoRESUMEN
Host microbial cross-talk is essential to maintain intestinal homeostasis. However, maladaptation of this response through microbial dysbiosis or defective host defense toward invasive intestinal bacteria can result in chronic inflammation. We have shown that macrophages differentiated in the presence of the bacterial metabolite butyrate display enhanced antimicrobial activity. Butyrate-induced antimicrobial activity was associated with a shift in macrophage metabolism, a reduction in mTOR kinase activity, increased LC3-associated host defense and anti-microbial peptide production in the absence of an increased inflammatory cytokine response. Butyrate drove this monocyte to macrophage differentiation program through histone deacetylase 3 (HDAC3) inhibition. Administration of butyrate induced antimicrobial activity in intestinal macrophages in vivo and increased resistance to enteropathogens. Our data suggest that (1) increased intestinal butyrate might represent a strategy to bolster host defense without tissue damaging inflammation and (2) that pharmacological HDAC3 inhibition might drive selective macrophage functions toward antimicrobial host defense.
Asunto(s)
Antiinfecciosos/farmacología , Butiratos/farmacología , Diferenciación Celular/efectos de los fármacos , Macrófagos/efectos de los fármacos , Monocitos/efectos de los fármacos , Animales , Diferenciación Celular/genética , Células Cultivadas , Colon/efectos de los fármacos , Colon/metabolismo , Colon/microbiología , Citocinas/genética , Citocinas/metabolismo , Disbiosis/microbiología , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Intestinos/efectos de los fármacos , Intestinos/microbiología , Macrófagos/metabolismo , Macrófagos/microbiología , Ratones Endogámicos C57BL , Microbiota/efectos de los fármacos , Microbiota/fisiología , Monocitos/metabolismo , Monocitos/microbiologíaRESUMEN
Tuberculosis (TB) remains a leading global cause of morbidity and mortality and an effective new vaccine is urgently needed. A major barrier to the rational development of novel TB vaccines is the lack of a validated immune correlate or biomarker of protection. Mycobacterial Growth Inhibition Assays (MGIAs) provide an unbiased measure of ability to control mycobacterial growth in vitro, and may represent a functional correlate of protection. However, the biological relevance of any potential correlate can only be assessed by determining the association with in vivo protection from either a controlled mycobacterial infection or natural development of TB disease. Our data demonstrate that the direct MGIA using peripheral blood mononuclear cells (PBMC) is measuring a biologically relevant response that correlates with protection from in vivo human BCG infection across two independent cohorts. This is the first report of an MGIA correlating with in vivo protection in the species-of-interest, humans, and furthermore on a per-individual as well as per-group basis. Control of mycobacterial growth in the MGIA is associated with a range of immune parameters measured post-BCG infection in vivo including the IFN-γ ELISpot response, frequency of PPD-specific IFN-γ or TNF-α producing CD4+ T cells and frequency of specific sub-populations of polyfunctional CD4+ T cells. Distinct transcriptomic profiles are associated with good vs. poor mycobacterial control in the MGIA, with good controllers showing enrichment for gene sets associated with antigen processing/presentation and the IL-23 pathway, and poor controllers showing enrichment for hypoxia-related pathways. This study represents an important step toward biologically validating the direct PBMC MGIA for use in TB vaccine development and furthermore demonstrates the utility of this assay in determining relevant immune mechanisms and pathways of protection.
Asunto(s)
Mycobacterium tuberculosis/crecimiento & desarrollo , Vacunas contra la Tuberculosis/administración & dosificación , Tuberculosis/prevención & control , Linfocitos T CD4-Positivos/inmunología , Técnicas de Cultivo de Célula , Humanos , Interleucina-23/genética , Interleucina-23/inmunología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/microbiología , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/inmunología , Tuberculosis/genética , Tuberculosis/inmunología , Tuberculosis/microbiología , Vacunas contra la Tuberculosis/inmunologíaRESUMEN
IL-23 is a key driver of pathogenic Th17 cell responses. It has been suggested that the transcription factor T-bet is required to facilitate IL-23-driven pathogenic effector functions; however, the precise role of T-bet in intestinal T cell responses remains elusive. Here, we show that T-bet expression by T cells is not required for the induction of colitis or the differentiation of pathogenic Th17 cells but modifies qualitative features of the IL-23-driven colitogenic response by negatively regulating IL-23R expression. Consequently, absence of T-bet leads to unrestrained Th17 cell differentiation and activation characterized by high amounts of IL-17A and IL-22. The combined increase in IL-17A/IL-22 results in enhanced epithelial cell activation and inhibition of either IL-17A or IL-22 leads to disease amelioration. Our study identifies T-bet as a key modulator of IL-23-driven colitogenic responses in the intestine and has important implications for understanding of heterogeneity among inflammatory bowel disease patients.
Asunto(s)
Linfocitos T CD4-Positivos/fisiología , Colitis/inmunología , Interleucina-23/metabolismo , Intestinos/inmunología , Proteínas de Dominio T Box/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Interleucina-17/metabolismo , Interleucinas/metabolismo , Masculino , Ratones Endogámicos C57BL , Miembro 3 del Grupo F de la Subfamilia 1 de Receptores Nucleares/metabolismo , Receptores de Interleucina/metabolismo , Interleucina-22RESUMEN
Vaccines to protect against tuberculosis (TB) are urgently needed. We performed a case-control analysis to identify immune correlates of TB disease risk in Bacille Calmette-Guerin (BCG) immunized infants from the MVA85A efficacy trial. Among 53 TB case infants and 205 matched controls, the frequency of activated HLA-DR(+) CD4(+) T cells associates with increased TB disease risk (OR=1.828, 95% CI=1.25-2.68, P=0.002, FDR=0.04, conditional logistic regression). In an independent study of Mycobacterium tuberculosis-infected adolescents, activated HLA-DR(+) CD4(+) T cells also associate with increased TB disease risk (OR=1.387, 95% CI=1.068-1.801, P=0.014, conditional logistic regression). In infants, BCG-specific T cells secreting IFN-γ associate with reduced risk of TB (OR=0.502, 95% CI=0.29-0.86, P=0.013, FDR=0.14). The causes and impact of T-cell activation on disease risk should be considered when designing and testing TB vaccine candidates for these populations.
Asunto(s)
Vacuna BCG/inmunología , Linfocitos T CD4-Positivos/inmunología , Activación de Linfocitos/inmunología , Vacunación , Adolescente , Formación de Anticuerpos/inmunología , Estudios de Casos y Controles , Estudios de Cohortes , Antígenos HLA-DR/inmunología , Humanos , Inmunidad , Inmunoglobulina G/inmunología , Lactante , Modelos Logísticos , Mycobacterium tuberculosis/inmunología , Placebos , Factores de Riesgo , Factores de Tiempo , Tuberculosis/sangre , Tuberculosis/inmunología , Vacunas contra la Tuberculosis/inmunología , Vacunas de ADNRESUMEN
BACKGROUND: Bacillus Calmette-Guerin (BCG) vaccine is the most widely administered vaccine in the world, yet its mechanism of action remains unclear. We hypothesize that certain immune pathways are associated with reduced mycobacterial growth following BCG challenge in human volunteers. METHODS: We used samples from a mycobacterial challenge in which previously BCG-vaccinated or BCG-naive adults in the United Kingdom were challenged intradermally with a standard dose of BCG. Any remaining BCG was quantified in a skin biopsy specimen obtained 2 weeks after challenge and used as a measure of BCG growth and functional antimycobacterial immunity. We measured the immune response over the 2-week challenge, using DNA microarrays and flow cytometry, and correlated this with mycobacterial growth. RESULTS: The magnitude of the immune response to BCG is greater in previously vaccinated volunteers, and this correlates with reduced mycobacterial growth but increased scarring at the vaccination site. In particular, the interferon γ and interleukin 17 pathways are strongly induced in previously vaccinated volunteers and correlate with reduced mycobacterial growth in this population. CONCLUSION: This study identifies pathways associated with control of mycobacterial growth in vivo in human volunteers and supports the use of BCG challenge as a tool for evaluating vaccine efficacy and identifying mechanisms of antimycobacterial immunity.
Asunto(s)
Vacuna BCG/inmunología , Citocinas/metabolismo , Regulación de la Expresión Génica/inmunología , Mycobacterium bovis/inmunología , Apoptosis , Citocinas/genética , Glucólisis , Humanos , Inmunidad Innata , Leucocitos Mononucleares/metabolismo , Enfermedades Cutáneas Bacterianas/inmunología , Enfermedades Cutáneas Bacterianas/microbiología , Linfocitos TRESUMEN
FOXP3(+) regulatory T cells (Treg cells) are abundant in the intestine, where they prevent dysregulated inflammatory responses to self and environmental stimuli. It is now appreciated that Treg cells acquire tissue-specific adaptations that facilitate their survival and function; however, key host factors controlling the Treg response in the intestine are poorly understood. The interleukin (IL)-1 family member IL-33 is constitutively expressed in epithelial cells at barrier sites, where it functions as an endogenous danger signal, or alarmin, in response to tissue damage. Recent studies in humans have described high levels of IL-33 in inflamed lesions of inflammatory bowel disease patients, suggesting a role for this cytokine in disease pathogenesis. In the intestine, both protective and pathological roles for IL-33 have been described in murine models of acute colitis, but its contribution to chronic inflammation remains ill defined. Here we show in mice that the IL-33 receptor ST2 is preferentially expressed on colonic Treg cells, where it promotes Treg function and adaptation to the inflammatory environment. IL-33 signalling in T cells stimulates Treg responses in several ways. First, it enhances transforming growth factor (TGF)-ß1-mediated differentiation of Treg cells and, second, it provides a necessary signal for Treg-cell accumulation and maintenance in inflamed tissues. Strikingly, IL-23, a key pro-inflammatory cytokine in the pathogenesis of inflammatory bowel disease, restrained Treg responses through inhibition of IL-33 responsiveness. These results demonstrate a hitherto unrecognized link between an endogenous mediator of tissue damage and a major anti-inflammatory pathway, and suggest that the balance between IL-33 and IL-23 may be a key controller of intestinal immune responses.
Asunto(s)
Interleucinas/inmunología , Intestinos/citología , Intestinos/inmunología , Linfocitos T Reguladores/inmunología , Animales , Colitis/inmunología , Colitis/patología , Colon/citología , Colon/inmunología , Colon/patología , Modelos Animales de Enfermedad , Femenino , Inmunidad Mucosa , Inflamación/inmunología , Inflamación/metabolismo , Inflamación/patología , Interleucina-23/inmunología , Interleucina-33 , Interleucinas/antagonistas & inhibidores , Interleucinas/metabolismo , Intestinos/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Receptores de Interleucina/metabolismo , Transducción de Señal/inmunología , Linfocitos T Reguladores/citología , Timo/citología , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
The first phase IIb safety and efficacy trial of a new tuberculosis vaccine since that for BCG was completed in October 2012. BCG-vaccinated South African infants were randomized to receive modified vaccinia virus Ankara, expressing the Mycobacterium tuberculosis antigen 85A (MVA85A), or placebo. MVA85A did not significantly boost the protective effect of BCG. Cryopreserved samples provide a unique opportunity for investigating the correlates of the risk of tuberculosis disease in this population. Due to the limited amount of sample available from each infant, preliminary work was necessary to determine which assays and conditions give the most useful information. Peripheral blood mononuclear cells (PBMC) were stimulated with antigen 85A (Ag85A) and purified protein derivative from M. tuberculosis in an ex vivo gamma interferon (IFN-γ) enzyme-linked immunosorbent spot assay (ELISpot) and a Ki67 proliferation assay. The effects of a 2-h or overnight rest of thawed PBMC on ELISpot responses and cell populations were determined. Both the ELISpot and Ki67 assays detected differences between the MVA85A and placebo groups, and the results correlated well. The cell numbers and ELISpot responses decreased significantly after an overnight rest, and surface flow cytometry showed a significant loss of CD4(+) and CD8(+) T cells. Of the infants tested, 50% had a positive ELISpot response to a single pool of flu, Epstein-Barr virus (EBV), and cytomegalovirus (CMV) (FEC) peptides. This pilot work has been essential in determining the assays and conditions to be used in the correlate study. Moving forward, PBMC will be rested for 2 h before assay setup. The ELISpot assay, performed in duplicate, will be selected over the Ki67 assay, and further work is needed to evaluate the effect of high FEC responses on vaccine-induced immunity and susceptibility to tuberculosis disease.