RESUMO
Tuberculosis, caused by Mycobacterium tuberculosis, remains an enduring global health challenge due to the limited efficacy of existing treatments. Although much research has focused on immune failure, the role of host macrophage biology in controlling the disease remains underappreciated. Here we show, through multi-modal single-cell RNA sequencing in a murine model, that different alveolar macrophage subsets play distinct roles in either advancing or controlling the disease. Initially, alveolar macrophages that are negative for the CD38 marker are the main infected population. As the infection progresses, CD38+ monocyte-derived and tissue-resident alveolar macrophages emerge as significant controllers of bacterial growth. These macrophages display a unique chromatin organization pre-infection, indicative of epigenetic priming for pro-inflammatory responses. Moreover, intranasal BCG immunization increases the numbers of CD38+ macrophages, enhancing their capability to restrict Mycobacterium tuberculosis growth. Our findings highlight the dynamic roles of alveolar macrophages in tuberculosis and open pathways for improved vaccines and therapies.
Assuntos
ADP-Ribosil Ciclase 1 , Pulmão , Macrófagos Alveolares , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis , Animais , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/microbiologia , Macrófagos Alveolares/metabolismo , Mycobacterium tuberculosis/imunologia , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Camundongos , ADP-Ribosil Ciclase 1/metabolismo , ADP-Ribosil Ciclase 1/genética , Modelos Animais de Doenças , Feminino , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia , Glicoproteínas de Membrana/metabolismo , Glicoproteínas de Membrana/imunologia , Vacina BCG/imunologia , Análise de Célula Única , Tuberculose/imunologia , Tuberculose/microbiologiaRESUMO
Background: To effectively control tuberculosis (TB), it is crucial to distinguish between active TB disease and latent TB infection (LTBI) to provide appropriate treatment. However, no such tests are currently available. Immune responses associated with active TB and LTBI are dynamic and exhibit distinct patterns. Comparing these differences is crucial for developing new diagnostic methods and understanding the etiology of TB. This study aimed to investigate the relationship between pro- and anti-inflammatory CD4+ cytokine production following stimulation with two types of latency-associated Mycobacterium tuberculosis (M.tb) antigens to allow differentiation between active TB and LTBI. Methods: Cryopreserved PBMCs from patients with active TB disease or LTBI were stimulated overnight with replication-related antigen [ESAT-6/CFP-10 (E/C)] or two latency-associated antigens [heparin-binding hemagglutinin (HBHA) and alpha-crystallin-like protein (Acr)]. Responses were analyzed using multiparameter flow cytometry: active TB disease (n=15), LTBI (n=15) and ELISA: active TB disease (n=26) or LTBI (n=27). Results: CD4+ central memory T cells (Tcm) specific to E/C and CD4+ effector memory T cells specific to Acr and HBHA were higher in LTBI than in TB patients. IFN-γ+Tcm and IL-17+ Tem cells was higher in the LTBI group (p= 0.012 and p=0.029 respectively), but IL-10+ Tcm was higher in the active TB group (p= 0.029) following HBHA stimulation. Additionally, following stimulation with HBHA, IL-10 production from CD4+ T cells was significantly elevated in patients with active TB compared to those with LTBI (p= 0.0038), while CD4+ T cell production of IL-17 and IFN-γ was significantly elevated in LTBI compared to active TB (p= 0.0076, p< 0.0001, respectively). HBHA also induced more CCR6+IL-17+CD4Tcells and IL-17+FoxP3+CD25+CD4Tcells in LTBI than in TB patients (P=0.026 and P=0.04, respectively). HBHA also induced higher levels of IFN-γ+IL-10+CD4+ T cells in patients with active TB (Pp=0.03) and higher levels of IFN-γ+IL-17+ CD4+ T cells in those with LTBI (p=0.04). HBHA-specific cytokine production measured using ELISA showed higher levels of IFN-γ in participants with LTBI (P=0.004) and higher levels of IL-10 in those with active TB (P=0.04). Conclusion: Stimulation with HBHA and measurement of CD4+ T cell production of IFN-γ, IL-10, and IL-17 could potentially differentiate active TB from LTBI. The characteristics of cytokine-expressing cells induced by HBHA also differed between participants with active TB and LTBI.
Assuntos
Antígenos de Bactérias , Linfócitos T CD4-Positivos , Interferon gama , Interleucina-10 , Interleucina-17 , Tuberculose Latente , Mycobacterium tuberculosis , Tuberculose , Humanos , Masculino , Feminino , Linfócitos T CD4-Positivos/imunologia , Adulto , Interleucina-17/imunologia , Interleucina-17/metabolismo , Mycobacterium tuberculosis/imunologia , Interleucina-10/imunologia , Interferon gama/metabolismo , Interferon gama/imunologia , Pessoa de Meia-Idade , Tuberculose Latente/imunologia , Tuberculose/imunologia , Antígenos de Bactérias/imunologia , Idoso , Adulto Jovem , LectinasRESUMO
The control of bacterial growth is key to the prevention and treatment of tuberculosis (TB). Granulomas represent independent foci of the host immune response that present heterogeneous capacity for control of bacterial growth. At the whole tissue level, B cells and CD4 or CD8 T cells have an established role in immune protection against TB. Immune cells interact within each granuloma response, but the impact of granuloma immune composition on bacterial replication remains unknown. Here we investigate the associations between immune cell composition, including B cell, CD4, and CD8 T cells, and the state of replicating Mycobacterium tuberculosis (Mtb) within the granuloma. A measure of ribosomal RNA synthesis, the RS ratio®, represents a proxy measure of Mtb replication at the whole tissue level. We adapted the RS ratio through use of in situ hybridization, to identify replicating and non-replicating Mtb within each designated granuloma. We applied a regression model to characterize the associations between immune cell populations and the state of Mtb replication within each respective granuloma. In the evaluation of nearly 200 granulomas, we identified heterogeneity in both immune cell composition and proportion of replicating bacteria. We found clear evidence of directional associations between immune cell composition and replicating Mtb. Controlling for vaccination status and endpoint post-infection, granulomas with lower CD4 or higher CD8 cell counts are associated with a higher percent of replicating Mtb. Conversely, changes in B cell proportions were associated with little change in Mtb replication. This study establishes heterogeneity across granulomas, demonstrating that certain immune cell types are differentially associated with control of Mtb replication. These data suggest that evaluation at the granuloma level may be imperative to identifying correlates of immune protection.
Assuntos
Linfócitos T CD8-Positivos , Granuloma , Mycobacterium tuberculosis , Mycobacterium tuberculosis/imunologia , Humanos , Granuloma/imunologia , Granuloma/microbiologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Linfócitos T CD4-Positivos/imunologia , Linfócitos B/imunologia , Masculino , Tuberculose/imunologia , Tuberculose/microbiologiaRESUMO
Tuberculosis(TB) of the Central nervous system (CNS) is a rare and highly destructive disease. The emergence of drug resistance has increased treatment difficulty, leaving the Bacillus Calmette-Guérin (BCG) vaccine as the only licensed preventative immunization available. This study focused on identifying the epitopes of PknD (Rv0931c) and Rv0986 from Mycobacterium tuberculosis(Mtb) strain H37Rv using an in silico method. The goal was to develop a therapeutic mRNA vaccine for preventing CNS TB. The vaccine was designed to be non-allergenic, non-toxic, and highly antigenic. Codon optimization was performed to ensure effective translation in the human host. Additionally, the secondary and tertiary structures of the vaccine were predicted, and molecular docking with TLR-4 was carried out. A molecular dynamics simulation confirmed the stability of the complex. The results indicate that the vaccine structure shows effectiveness. Overall, the constructed vaccine exhibits ideal physicochemical properties, immune response, and stability, laying a theoretical foundation for future laboratory experiments.
Assuntos
Simulação por Computador , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Mycobacterium tuberculosis , Tuberculose do Sistema Nervoso Central , Humanos , Mycobacterium tuberculosis/imunologia , Tuberculose do Sistema Nervoso Central/prevenção & controle , Tuberculose do Sistema Nervoso Central/imunologia , Vacinas contra a Tuberculose/imunologia , Epitopos/imunologia , Epitopos/química , Vacinas de mRNA , Vacinas Sintéticas/imunologiaRESUMO
BACKGROUNDS: Mycobacterium tuberculosis (Mtb), the pathogen responsible for tuberculosis, secretes a multitude of proteins that modulate the host's immune response to ensure its own persistence. The region of difference (RD) genes encoding proteins play key roles in TB immunity and pathogenesis. Nevertheless, the roles of the majority of RD-encoded proteins remain to be elucidated. OBJECTS: To elucidate the role of Rv2652c located in RD13 in Mtb on bacterial growth, bacterial survival, and host immune response. METHODS: We constructed the strain MS_Rv2652c which over-expresses Mtb RD-encoding protein Rv2652c in M. smegmatis (MS), and compared it with the wild strain in the bacterial growth, bacterial survival, virulence of Rv2652c, and determined the effect of MS_Rv2652c on host immune response in macrophages. RESULTS: Rv2652c protein is located at cell wall of MS_Rv2652c strain and also an integral component of the Mtb H37Rv cell wall. Rv2652c can enhance the resistance of recombinant MS to various stressors. Moreover, Rv2652c inhibits host proinflammatory responses via modulation of the NF-κB pathway, thereby promoting Mtb survival in vitro and in vivo. CONCLUSION: Our data suggest that cell wall protein Rv2652c plays an important role in creating a favorable environment for bacterial survival by modulating host signals and could be established as a potential TB drug target.
Assuntos
Proteínas de Bactérias , Macrófagos , Mycobacterium tuberculosis , Mycobacterium tuberculosis/imunologia , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/imunologia , Animais , Camundongos , Macrófagos/imunologia , Macrófagos/microbiologia , Macrófagos/metabolismo , Tuberculose/imunologia , Tuberculose/microbiologia , Humanos , Interações Hospedeiro-Patógeno/imunologia , Virulência , Mycobacterium smegmatis/imunologia , Viabilidade Microbiana/imunologia , NF-kappa B/metabolismo , Camundongos Endogâmicos C57BL , Parede Celular/imunologia , Parede Celular/metabolismoRESUMO
BACKGROUND AND OBJECTIVE: To diagnose tuberculosis infection (TBI), whole blood is incubated with M.tuberculosis (Mtb)-specific peptides and the release of interferon-γ (IFN-γ) is measured in IFN-γ-release assays (IGRAs). Hyperglycaemia and fluctuations in blood glucose may modulate IFN-γ-release. Here, we investigated if glucose intake affects IFN-γ-release or IGRA results in IGRAs taken during an oral glucose tolerance test (OGTT). METHODS: Persons with TB disease (TB) or TBI underwent a standard 75-g OGTT at the start and end of treatment for TB or TBI. Blood for the IGRA QuantiFERON-TB Gold Plus (QFT) containing Mtb-specific tubes (TB1 and TB2), a non-specific mitogen tube (MIT) and an empty control tube (NIL) was drawn at sample-timepoints -15 (baseline), 60, 90, 120 and 240 min during the OGTT. Blood glucose was measured in parallel at all timepoints. IFN-γ-release (after subtraction of NIL) at each timepoint was compared with baseline using linear-mixed-model analysis. RESULTS: Twenty-four OGTTs from 14 participants were included in the final analysis. Compared to baseline, IFN-γ-release was increased at sample-timepoint 240 min for TB1; geometric mean (95% confidence interval) 3.0 (1.5-6.2) vs 2.5 (1.4-4.4) IU/mL (p = 0.047), and MIT; 182.6 (103.3-322.9) vs 146.0 (84.0-254.1) IU/mL (p = 0.002). Plasma glucose levels were not associated with IFN-γ-release and the QFT test results were unaffected by the OGTT. CONCLUSION: Ingestion of glucose after a 10-h fast was associated with increased IFN-γ-release after 240 min in the MIT tube. However, there was no association between plasma glucose levels at the QFT sampling timepoint and IFN-γ-release. Furthermore, the QFT test results were not affected by glucose intake. The overall effect of an OGTT and prevailing plasma glucose levels on IFN-γ-release in IGRAs seem limited. TRIAL REGISTRATION: Trial registration ID: NCT04830462 ( https://clinicaltrials.gov/study/NCT04830462 ). Registration date: 05-Apr-2021.
Assuntos
Glicemia , Teste de Tolerância a Glucose , Testes de Liberação de Interferon-gama , Interferon gama , Mycobacterium tuberculosis , Tuberculose , Humanos , Masculino , Feminino , Interferon gama/sangue , Pessoa de Meia-Idade , Adulto , Testes de Liberação de Interferon-gama/métodos , Tuberculose/sangue , Tuberculose/imunologia , Mycobacterium tuberculosis/imunologia , Glicemia/análise , Glucose/administração & dosagem , IdosoRESUMO
Bacille Calmette-Guérin (BCG) remains the only licensed vaccine against tuberculosis (TB). While BCG protects against TB in children, its protection against pulmonary TB in adults is suboptimal, and the development of a better TB vaccine is a global health priority. Previously, we reported two recombinant BCG strains effective against murine TB with low virulence and lung pathology in immunocompromised mice and guinea pigs. We have recently combined these two recombinant BCG strains into one novel vaccine candidate (BCGΔBCG1419c::ESAT6-PE25SS) and evaluated its immunogenicity, efficacy and safety profile in mice. This new vaccine candidate is non-inferior to BCG in protection against TB, presents reduced pro-inflammatory immune responses and displays an enhanced safety profile.
Assuntos
Vacina BCG , Hospedeiro Imunocomprometido , Vacinas Sintéticas , Animais , Vacina BCG/imunologia , Vacina BCG/efeitos adversos , Vacina BCG/genética , Camundongos , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/efeitos adversos , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Feminino , Tuberculose/prevenção & controle , Tuberculose/imunologia , Mycobacterium bovis/imunologia , Mycobacterium bovis/genética , Mycobacterium bovis/patogenicidade , Modelos Animais de Doenças , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidade , Camundongos Endogâmicos C57BL , Pulmão/microbiologia , Pulmão/patologia , Pulmão/imunologia , Tuberculose Pulmonar/prevenção & controle , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/microbiologia , Eficácia de VacinasRESUMO
Severe and chronic infections, including pneumonia, sepsis, and tuberculosis (TB), induce long-lasting epigenetic changes that are associated with an increase in all-cause postinfectious morbidity and mortality. Oncology studies identified metabolic drivers of the epigenetic landscape, with the tricarboxylic acid (TCA) cycle acting as a central hub. It is unknown if the TCA cycle also regulates epigenetics, specifically DNA methylation, after infection-induced immune tolerance. The following studies demonstrate that lipopolysaccharide and Mycobacterium tuberculosis induce changes in DNA methylation that are mediated by the TCA cycle. Infection-induced DNA hypermethylation is mitigated by inhibitors of cellular metabolism (rapamycin, everolimus, metformin) and the TCA cycle (isocitrate dehydrogenase inhibitors). Conversely, exogenous supplementation with TCA metabolites (succinate and itaconate) induces DNA hypermethylation and immune tolerance. Finally, TB patients who received everolimus have less DNA hypermethylation demonstrating proof of concept that metabolic manipulation can mitigate epigenetic scars.
Assuntos
Ciclo do Ácido Cítrico , Metilação de DNA , Tolerância Imunológica , Lipopolissacarídeos , Mycobacterium tuberculosis , Tuberculose , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/imunologia , Humanos , Animais , Tuberculose/imunologia , Tuberculose/genética , Tuberculose/microbiologia , Camundongos , Epigênese Genética , Succinatos/metabolismo , Everolimo/farmacologia , Ácido Succínico/metabolismoRESUMO
Mycobacterium tuberculosis (Mtb) infection represents a global health problem and is characterized by formation of granuloma with a necrotic center and a systemic inflammatory response. Inflammasomes have a crucial role in the host immune response towards Mtb. These intracellular multi-protein complexes are assembled in response to pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs). Inflammasome platforms activate caspases, leading to the maturation of the proinflammatory cytokines interleukin (IL)-1 and 18 and the cleavage of gasdermin D (GSDMD), a pore-forming protein responsible for cytokine release and pyroptotic cell death. Recent in vitro and in vivo findings have highlighted the importance of inflammasome signaling and subsequent necrotic cell death in Mtb-infected innate immune cells. However, we are just beginning to understand how inflammasomes contribute to disease or to a protective immune response in tuberculosis (TB). A detailed molecular understanding of inflammasome-associated pathomechanisms may foster the development of novel host-directed therapeutics or vaccines with improved activity. In this mini-review, we discuss the regulatory and molecular aspects of inflammasome activation and the associated immunological consequences for Mtb pathogenesis.
Assuntos
Inflamassomos , Mycobacterium tuberculosis , Tuberculose , Humanos , Inflamassomos/imunologia , Inflamassomos/metabolismo , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Tuberculose/microbiologia , Animais , Inflamação/imunologia , Transdução de Sinais/imunologia , Imunidade Inata , Interações Hospedeiro-Patógeno/imunologiaRESUMO
Tuberculosis (TB) remains one of the leading causes of death among infectious diseases, with 10.6 million new cases and 1.3 million deaths reported in 2022, according to the most recent WHO report. Early studies have shown an expansion of γδ T cells following TB infection in both experimental models and humans, indicating their abundance among lung lymphocytes and suggesting a role in protective immune responses against Mycobacterium tuberculosis (M. tuberculosis) infection. In this study, we hypothesized that distinct subsets of γδ T cells are associated with either protection against or disease progression in TB. To explore this, we applied large-scale scRNA-seq and bulk RNA-seq data integration to define the phenotypic and molecular characteristics of peripheral blood γδ T cells. Our analysis identified five unique γδ T subclusters, each with distinct functional profiles. Notably, we identified a unique cluster significantly enriched in the TCR signaling pathway, with high CD81 expression as a conserved marker. This distinct molecular signature suggests a specialized role for this cluster in immune signaling and regulation of immune response against M. tuberculosis. Flow cytometry confirmed our in silico results, showing that the mean fluorescence intensity (MFI) values of CD81 expression on γδ T cells were significantly increased in individuals with latent TB infection (TBI) compared to those with active TB (ATB). This finding underscores the importance of CD81 and its associated signaling mechanisms in modulating the activity and function of γδ T cells under TBI conditions, providing insights into potential therapeutic targets for TB management.
Assuntos
Mycobacterium tuberculosis , Receptores de Antígenos de Linfócitos T gama-delta , Humanos , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Mycobacterium tuberculosis/imunologia , Tetraspanina 28/metabolismo , Análise de Célula Única , Tuberculose Latente/imunologia , Tuberculose/imunologia , Tuberculose/microbiologia , Feminino , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Adulto , MasculinoRESUMO
The cytokine IFNγ is a principal effector of macrophage activation and immune resistance to mycobacterial infection; however, pathogenic mycobacteria are capable of surviving in IFNγ-activated macrophages by largely unknown mechanisms. In this study, we find that pathogenic mycobacteria, including M. bovis BCG and M. tuberculosis can sense IFNγ to promote their proliferative activity and virulence phenotype. Moreover, interaction with the host intracellular environment increases the susceptibility of mycobacteria to IFNγ through upregulating expression of mmpL10, a mycobacterial IFNγ receptor, thereby facilitating IFNγ-dependent survival and growth of mycobacteria in macrophages. Transmission electron microscopy analysis reveals that IFNγ triggers the secretion of extracellular vesicles, an essential virulence strategy of intracellular mycobacteria, while proteomics identifies numerous pivotal IFNγ-induced effectors required for mycobacterial infection in macrophages. Our study suggests that sensing host IFNγ is a crucial virulence mechanism used by pathogenic mycobacteria to survive and proliferate inside macrophages.
Assuntos
Interferon gama , Macrófagos , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis , Interferon gama/metabolismo , Interferon gama/imunologia , Macrófagos/microbiologia , Macrófagos/metabolismo , Macrófagos/imunologia , Animais , Camundongos , Mycobacterium tuberculosis/patogenicidade , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/metabolismo , Mycobacterium bovis/imunologia , Mycobacterium bovis/metabolismo , Humanos , Interações Hospedeiro-Patógeno/imunologia , Virulência , Receptores de Interferon/metabolismo , Receptores de Interferon/genética , Receptor de Interferon gama , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/imunologia , Ativação de Macrófagos , Infecções por Mycobacterium/microbiologia , Infecções por Mycobacterium/imunologia , Infecções por Mycobacterium/metabolismo , Infecções por Mycobacterium/patologiaRESUMO
BACKGROUND: PPE 59, which is absent from bacillus Calmette Guérin (BCG) strains, seems to induce a humoral immune response in patients with tuberculosis (TB). Additional studies are needed to better evaluate this protein in immune response to tuberculosis. OBJECTIVES: To evaluate the response of antibodies to PPE59 in TB individuals, its combination with IgG response to other, previously tested mycobacterial antigens (Ag) and with sputum smear microbiology (SM) results. METHODS: We have cloned and expressed the rv3429 gene that encodes PPE59, then IgG, IgM, and IgA against PPE59 antigens measured by enzyme-linked immunosorbent assay (ELISA) in 212 sera samples obtained from the following subject cohorts: TB residents from Italy (79) and in Brazil (52); and an all-Brazilian cohort of 55 patients with other respiratory disorders; 10 patients infected with non-tuberculous mycobacteria, and 16 asymptomatic subjects. Drawing on results from a previous study(17) of serum samples from Brazilian subjects tested for IgG by ELISA against mycobacterial antigens ESAT-6, 16kDa, MT10.3, MPT-64 and 38kDa, the results were analysed in combination with those of the PPE59 and SM tests. FINDINGS: Keeping the specificity rate at 97%, the overall PPE59 IgA sensitivity was 42.7%, while IgG and IgM showed lower performance (p < 0.0001). Combining PPE59 IgA/16kDa IgG results increased sensitivity to 71%, and even higher rates when the results were combined with SM results (86.5%, p = 0.001), at 88.9% specificity. Positive IgA was associated with pulmonary image alterations of high TB probability (p < 0.05). MAIN CONCLUSIONS: Tests with TB patients found a moderate frequency of positivity for PPE59 IgA. However, the higher level of sensitivity attained in combination with PPE59 IgA/16kDa IgG/SM results unheard of before, although imperfect, suggests that this may be a potential additional tool for rapid detection of TB in low-resource areas.
Assuntos
Anticorpos Antibacterianos , Antígenos de Bactérias , Biomarcadores , Ensaio de Imunoadsorção Enzimática , Imunoglobulina A , Imunoglobulina G , Imunoglobulina M , Humanos , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/imunologia , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Feminino , Biomarcadores/sangue , Adulto , Imunoglobulina A/sangue , Sensibilidade e Especificidade , Pessoa de Meia-Idade , Mycobacterium tuberculosis/imunologia , Tuberculose/diagnóstico , Tuberculose/imunologia , Tuberculose/sangue , Adulto Jovem , Tuberculose Pulmonar/diagnóstico , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/sangue , Escarro/microbiologia , Brasil , Proteínas de Bactérias/imunologia , Idoso , Adolescente , Estudos de CoortesRESUMO
Bacille-Calmette-Guérin (BCG) is the only approved vaccine against Mycobacterium tuberculosis (MTB), offering protection not only against tuberculosis (TB) but also non-related infections. 'Trained immunity' of innate immune cells is considered one of the mechanisms of this broad protection derived through BCG. Here, we investigated the effect of BCG on Natural Killer (NK) cells, a key innate immune cell type, and their subsequent responses to mycobacterial and HIV antigens. We found that BCG-induced KLRG1+ NK cells exhibit significantly higher production of IFNγ, compared to KLRG1- cells, indicating their memory-like responses upon exposure to these antigens (p < 0.05). These findings may be important in regions of high burden of HIV and TB where BCG is routinely administered.
Assuntos
Vacina BCG , Infecções por HIV , Memória Imunológica , Interferon gama , Células Matadoras Naturais , Lectinas Tipo C , Mycobacterium tuberculosis , Receptores Imunológicos , Tuberculose , Células Matadoras Naturais/imunologia , Receptores Imunológicos/imunologia , Memória Imunológica/imunologia , Lectinas Tipo C/imunologia , Lectinas Tipo C/metabolismo , Humanos , Interferon gama/imunologia , Interferon gama/metabolismo , Vacina BCG/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Infecções por HIV/imunologia , Antígenos de Bactérias/imunologia , Antígenos Virais/imunologia , Imunização/métodosRESUMO
Introduction: Tuberculosis (TB) remains a significant health concern in India, and its complexity is exacerbated by the rising occurrence of non-communicable diseases such as diabetes mellitus (DM). Recognizing that DM is a risk factor for active TB, the emerging comorbidity of TB and PDM (TB-PDM) presents a particular challenge. Our study focused on the impact of PDM on cytokine and chemokine profiles in patients with pulmonary tuberculosis TB) who also have PDM. Materials and methods: We measured and compared the cytokine (GM-CSF, IFN-γ, IL-1α/IL-1F1, IL-1ß/IL-1F2, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p70, IL-13, IL-17/IL-17A, IL-18/IL-1F4, TNF-α) and chemokine (CCL1, CCL2, CCL3, CCL4, CCL11, CXCL1, CXCL2, CXCL9, CXCL10, and CXCL11) levels in plasma samples of TB-PDM, only TB or only PDM using multiplex assay. Results: We observed that PDM was linked to higher mycobacterial loads in TB. Patients with coexisting TB and PDM showed elevated levels of various cytokines (including IFNγ, TNFα, IL-2, IL-17, IL-1α, IL-1ß, IL-6, IL-12, IL-18, and GM-CSF) and chemokines (such as CCL1, CCL2, CCL3, CCL4, CCL11, CXCL1, CXCL9, CXCL10, and CXCL11). Additionally, cytokines such as IL-18 and GM-CSF, along with the chemokine CCL11, were closely linked to levels of glycated hemoglobin (HbA1c), hinting at an interaction between glycemic control and immune response in TB patients with PDM. Conclusion: Our results highlight the complex interplay between metabolic disturbances, immune responses, and TB pathology in the context of PDM, particularly highlighting the impact of changes in HbA1c levels. This emphasizes the need for specialized approaches to manage and treat TB-PDM comorbidity.
Assuntos
Citocinas , Estado Pré-Diabético , Tuberculose Pulmonar , Humanos , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/sangue , Citocinas/sangue , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Estado Pré-Diabético/imunologia , Estado Pré-Diabético/sangue , Quimiocinas/sangue , Biomarcadores/sangue , Mycobacterium tuberculosis/imunologia , Índia/epidemiologiaRESUMO
Tuberculosis (TB) is caused by infection with the bacterial pathogen Mycobacterium tuberculosis (M.tb) in the respiratory tract. There was an estimated 10.6 million people newly diagnosed with TB, and there were approximately 1.3 million deaths caused by TB in 2022. Although the global prevalence of TB has remained high for decades and is an annual leading cause of death attributed to infectious diseases, only one vaccine, Bacillus Calmette-Guérin (BCG), has been approved so far to prevent/attenuate TB disease. Correlates of protection or immunological mechanisms that are needed to control M.tb remain unknown. The protective role of antibodies after BCG vaccination has also remained largely unclear; however, recent studies have provided evidence for their involvement in protection against disease, as biomarkers for the state of infection, and as potential predictors of outcomes. Interestingly, the antibodies generated post-vaccination with BCG are linked to the activation of innate immune cascades, providing further evidence that antibody effector functions are critical for protection against respiratory pathogens such as M.tb. In this review, we aim to provide current knowledge of antibody application in TB diagnosis, prevention, and treatment. Particularly, this review will focus on 1) The role of antibodies in preventing M.tb infections through preventing Mtb adherence to epithelium, antibody-mediated phagocytosis, and antibody-mediated cellular cytotoxicity; 2) The M.tb-directed antibody response generated after vaccination and how humoral profiles with different glycosylation patterns of these antibodies are linked with protection against the disease state; and 3) How antibody-mediated immunity against M.tb can be further explored as early diagnosis biomarkers and different detection methods to combat the global M.tb burden. Broadening the paradigm of differentiated antibody profiling and antibody-based detection during TB disease progression offers new directions for diagnosis, treatment, and preventative strategies. This approach involves linking the aforementioned humoral responses with the disease state, progression, and clearance.
Assuntos
Anticorpos Antibacterianos , Vacina BCG , Mycobacterium tuberculosis , Tuberculose , Humanos , Mycobacterium tuberculosis/imunologia , Anticorpos Antibacterianos/imunologia , Tuberculose/imunologia , Tuberculose/prevenção & controle , Vacina BCG/imunologia , Animais , Imunidade Inata , Vacinação , BiomarcadoresRESUMO
Tuberculosis (TB), a deadly disease caused by Mycobacterium tuberculosis (Mtb) infection, remains one of the top killers among infectious diseases worldwide. How to increase targeting effects of current anti-TB chemotherapeutics and enhance anti-TB immunological responses remains a big challenge in TB and drug-resistant TB treatment. Here, mannose functionalized and polyetherimide protected graphene oxide system (GO-PEI-MAN) was designed for macrophage-targeted antibiotic (rifampicin) and autophagy inducer (carbamazepine) delivery to achieve more effective Mtb killings by combining targeted drug killing and host immunological clearance. GO-PEI-MAN system demonstrated selective uptake by in vitro macrophages and ex vivo macrophages from macaques. The endocytosed GO-PEI-MAN system would be transported into lysosomes, where the drug loaded Rif@Car@GO-PEI-MAN system would undergo accelerated drug release in acidic lysosomal conditions. Rif@Car@GO-PEI-MAN could significantly promote autophagy and apoptosis in Mtb infected macrophages, as well as induce anti-bacterial M1 polarization of Mtb infected macrophages to increase anti-bacterial IFN-γ and nitric oxide production. Collectively, Rif@Car@GO-PEI-MAN demonstrated effectively enhanced intracellular Mtb killing effects than rifampicin, carbamazepine or GO-PEI-MAN alone in Mtb infected macrophages, and could significantly reduce mycobacterial burdens in the lung of infected mice with alleviated pathology and inflammation without systemic toxicity. This macrophage targeted nanosystem synergizing increased drug killing efficiency and enhanced host immunological defense may be served as more effective therapeutics against TB and drug-resistant TB.
Assuntos
Antituberculosos , Grafite , Macrófagos , Mycobacterium tuberculosis , Rifampina , Tuberculose , Grafite/química , Animais , Mycobacterium tuberculosis/efeitos dos fármacos , Mycobacterium tuberculosis/imunologia , Tuberculose/tratamento farmacológico , Tuberculose/imunologia , Tuberculose/microbiologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Rifampina/farmacologia , Rifampina/administração & dosagem , Rifampina/uso terapêutico , Camundongos , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Antituberculosos/administração & dosagem , Autofagia/efeitos dos fármacos , Macaca , Nanopartículas , Células RAW 264.7RESUMO
BACKGROUND: The polarization of macrophages plays a critical role in the immune response to infectious diseases, with M2 polarization shown to be particularly important in various pathological processes. However, the specific mechanisms of M2 macrophage polarization in Mycobacterium tuberculosis (Mtb) infection remain unclear. In particular, the roles of Granulin (GRN) and tumor necrosis factor receptor 2 (TNFR2) in the M2 polarization process have not been thoroughly studied. OBJECTIVE: To investigate the effect of macrophage M2 polarization on Mtb infection and the mechanism of GRN and TNFR2 in M2 polarization. METHODS: Forty patients with pulmonary tuberculosis (PTB) and 40 healthy volunteers were enrolled in this study, and peripheral blood samples were taken to detect the levels of TNFR2 and GRN mRNA by Quantitative Reverse Transcription Polymerase Chain Reaction (RT-qPCR); monocytes were isolated and then assessed by Flow Cytometry (FC) for M1 and M2 macrophage levels. To further validate the function of TNFR2 in macrophage polarization, we used interleukin 4 (IL-4) to induce mouse monocyte macrophages RAW264.7 to M2 polarized state. The expression of TNFR2 was detected by Western Blot and RT-qPCR. Next, we constructed a GRN knockdown plasmid and transfected it into IL-4-induced mouse monocyte macrophage RAW264.7, and detected the expression of TNFR2, M1 macrophage-associated factors tumor necrosis factor-α (TNF-α), inducible nitric oxide synthase (iNOS), and interleukin 6 (IL-6), and the M2 macrophage-associated factors CD206, IL-10, and Arginase 1 (Arg1); Immunofluorescence staining was used to monitor the expression of CD86+ and CD206+, and FC was used to analyze the macrophage phenotype. Subsequently, immunoprecipitation was used to detect the binding role of GRN and TNFR2. Finally, the effects of GRN and TNFR2 in macrophage polarization were further explored by knocking down GRN and simultaneously overexpressing TNFR2 and observing the macrophage polarization status. RESULTS: The results of the study showed elevated expression of TNFR2 and GRN and predominance of M2 type in macrophages in PTB patients compared to healthy volunteers (p < 0.05). Moreover, TNFR2 was highly expressed in M2 macrophages (p < 0.05). Additionally, GRN knockdown was followed by elevated expression of M1 polarization markers TNF-α, iNOS and IL-6 (p < 0.05), decreased levels of M2 polarization-associated factors CD206, IL-10 and Arg1 (p < 0.05), and macrophage polarization towards M1. Subsequently, we found that GRN binds to TNFR2 and that GRN upregulates TNFR2 expression (p < 0.05). In addition, knockdown of GRN elevated M1 polarization marker expression, decreased M2 polarization marker expression, and increased M1 macrophages and decreased M2 macrophages, whereas concurrent overexpression of TNFR2 decreased M1 polarization marker expression, elevated M2 polarization marker expression, and decreased M1 macrophages and increased M2 macrophages. CONCLUSION: TNFR2 and GRN are highly expressed in PTB patients and GRN promotes macrophage M2 polarization by upregulating TNFR2 expression.
Assuntos
Macrófagos , Mycobacterium tuberculosis , Progranulinas , Receptores Tipo II do Fator de Necrose Tumoral , Humanos , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Receptores Tipo II do Fator de Necrose Tumoral/genética , Macrófagos/metabolismo , Macrófagos/imunologia , Animais , Camundongos , Mycobacterium tuberculosis/imunologia , Feminino , Adulto , Masculino , Progranulinas/metabolismo , Progranulinas/genética , Pessoa de Meia-Idade , Tuberculose Pulmonar/imunologia , Tuberculose Pulmonar/metabolismo , Tuberculose Pulmonar/microbiologia , Células RAW 264.7 , Estudos de Casos e Controles , Ativação de Macrófagos , Óxido Nítrico Sintase Tipo II/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Lectinas Tipo C/metabolismo , Lectinas Tipo C/genéticaRESUMO
The limitations of TB treatment are the long duration and immune-dampening effects of anti-tuberculosis therapy. The Cell wall plays a crucial role in survival and virulence; hence, enzymes involved in its biosynthesis are good therapeutic targets. Here, we identify Mycobacterium tuberculosis (Mtb) GlmM, (GlmMMtb) engaged in the UDP-GlcNAc synthesis pathway as an essential enzyme. We generated a conditional knockdown strain, Rv-glmMkD using the CRISPR interference-mediated gene silencing approach. Depletion of GlmMMtb affects the morphology and thickness of the cell wall. The Rv-glmMkD strain attenuated Mtb survival in vitro, in the host macrophages (ex vivo), and in a murine mice infection model (in vivo). Results suggest that the depletion of GlmMMtb induces M1 macrophage polarization, prompting a pro-inflammatory cytokine response, apparent from the upregulation of activation markers, including IFNÉ£ and IL-17 that resists the growth of Mtb. These observations provide a rationale for exploring GlmMMtb as a potential therapeutic target.
Assuntos
Proteínas de Bactérias , Macrófagos , Mycobacterium tuberculosis , Tuberculose , Mycobacterium tuberculosis/imunologia , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidade , Animais , Camundongos , Tuberculose/imunologia , Tuberculose/microbiologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Macrófagos/imunologia , Macrófagos/microbiologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Feminino , Interações Hospedeiro-Patógeno/imunologia , Modelos Animais de Doenças , HumanosRESUMO
Tuberculosis (TB) has been and still is a global emergency for centuries. Prevention of disease through vaccination would have a major impact on disease prevalence, but the only available current vaccine, BCG, has insufficient impact. In this article, a novel subunit vaccine against TB was developed, using the Ag85B-ESAT6-Rv2034 fusion antigen, two adjuvants - CpG and MPLA, and a cationic pH-sensitive liposome as a delivery system, representing a new TB vaccine delivery strategy not previously reported for TB. In vitro in human dendritic cells (DCs), the adjuvanted formulation induced a significant increase in the production of (innate) cytokines and chemokines compared to the liposome without additional adjuvants. In vivo, the new vaccine administrated subcutaneously significantly reduced Mycobacterium tuberculosis (Mtb) bacterial load in the lungs and spleens of mice, significantly outperforming results from mice vaccinated with the antigen mixed with adjuvants without liposomes. In-depth analysis underpinned the vaccine's effectiveness in terms of its capacity to induce polyfunctional CD4+ and CD8+ T-cell responses, both considered essential for controlling Mtb infection. Also noteworthy was the differential abundance of various CD69+ B-cell subpopulations, which included IL17-A-producing B-cells. The vaccine stimulated robust antigen-specific antibody titers, further extending its potential as a novel protective agent against TB.
Assuntos
Adjuvantes Imunológicos , Lipossomos , Mycobacterium tuberculosis , Vacinas contra a Tuberculose , Tuberculose , Vacinas de Subunidades Antigênicas , Animais , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/imunologia , Mycobacterium tuberculosis/imunologia , Camundongos , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Humanos , Feminino , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/farmacologia , Concentração de Íons de Hidrogênio , Tuberculose/prevenção & controle , Tuberculose/imunologia , Camundongos Endogâmicos C57BL , Células Dendríticas/imunologia , Cátions , Antígenos de Bactérias/imunologia , Antígenos de Bactérias/administração & dosagem , Citocinas/metabolismo , Linfócitos T CD8-Positivos/imunologia , Pulmão/imunologia , Pulmão/microbiologia , Linfócitos T CD4-Positivos/imunologia , Carga BacterianaRESUMO
Severe defects in human IFNγ immunity predispose individuals to both Bacillus Calmette-Guérin disease and tuberculosis, whereas milder defects predispose only to tuberculosis1. Here we report two adults with recurrent pulmonary tuberculosis who are homozygous for a private loss-of-function TNF variant. Neither has any other clinical phenotype and both mount normal clinical and biological inflammatory responses. Their leukocytes, including monocytes and monocyte-derived macrophages (MDMs) do not produce TNF, even after stimulation with IFNγ. Blood leukocyte subset development is normal in these patients. However, an impairment in the respiratory burst was observed in granulocyte-macrophage colony-stimulating factor (GM-CSF)-matured MDMs and alveolar macrophage-like (AML) cells2 from both patients with TNF deficiency, TNF- or TNFR1-deficient induced pluripotent stem (iPS)-cell-derived GM-CSF-matured macrophages, and healthy control MDMs and AML cells differentiated with TNF blockers in vitro, and in lung macrophages treated with TNF blockers ex vivo. The stimulation of TNF-deficient iPS-cell-derived macrophages with TNF rescued the respiratory burst. These findings contrast with those for patients with inherited complete deficiency of the respiratory burst across all phagocytes, who are prone to multiple infections, including both Bacillus Calmette-Guérin disease and tuberculosis3. Human TNF is required for respiratory-burst-dependent immunity to Mycobacterium tuberculosis in macrophages but is surprisingly redundant otherwise, including for inflammation and immunity to weakly virulent mycobacteria and many other infectious agents.