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INTRODUCTION: Kaposi sarcoma (KS) is an angioproliferative disease caused by human herpesvirus 8 (HHV-8) and is mediated by cytokines in an immunodeficient environment. This study aimed to compare IL-6, IL-10, and TNFα levels among AIDS patients with disseminated KS (DKS), treatment naïve patients living with HIV (PLWHIV) without DKS, and healthy controls. Secondary outcomes were to compare cytokines levels in patients with DKS and unfavorable outcomes, as well as an analysis of the behavior of cytokines over time. METHODS: This cohort study was performed at two centers in Mexico City. Three groups were included. Group 1: HIV+ treatment naïve with DKS, Group 2: HIV+ treatment naïve without KS, and Group 3: HIV negative, healthy controls. Plasmatic IL-6, IL-10, and TNFα levels were measured at baseline and over time in Groups 1 and 2. RESULTS: Seventy-six patients were included: 39 (52%) in Group 1, 17 (22%) in Group 2, and 20 (26%) in Group 3. The median baseline IL-6, IL-10, and TNFα levels were significantly higher in group 1. In group 1, baseline IL-6 was higher in patients who died than in survivors (14.4 vs 5.8 pg/mL p=0.048). Patients with severe immune reconstitution inflammatory syndrome due to KS (S-IRIS-KS) had higher IL-6 values than those without it (14.4 vs 5.8 pg/mL p=0.004). In the repeated-measures model in group 1, IL-10 levels were higher in patients who died (p<0.001) and developed IRIS-KS (p=0.01). CONCLUSIONS: IL-6, IL-10, and TNF α levels were markedly higher in patients with DKS. IL-6 and IL-10 levels were higher in patients with unfavorable outcomes.
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BACKGROUND: Persistent respiratory symptoms and lung abnormalities post-COVID-19 are public health problems. This study evaluated biomarkers to stratify high-risk patients to the development or persistence of post-COVID-19 interstitial lung disease. METHODS: One hundred eighteen patients discharged with residual lung abnormalities compatible with interstitial lung disease (COVID-ILD patients) after a severe COVID-19 were followed for 1 year (post-COVID-ILD patients). Physical examination, pulmonary function tests, and chest high-resolution computed tomography (HRCT) were performed. Soluble forms (s) of PD-L1, PD-L2, TIM-3, and GAL-9 were evaluated in serum and cell culture supernatant, as well as T-cells subsets and the transmembrane expression of PD-L1 and PD-L2 on the cell surface. RESULTS: Eighty percent of the post-COVID-ILD patients normalized their lung function at 1-year follow-up, 8% presented COVID-independent ILD, and 12% still showed functional and HRCT alterations. PD-L2 levels were heterogeneous during acute COVID-19 (aCOVID); patients who increased (at least 30%) their sPD-L2 levels at 1 year post-COVID-19 and exhibited altered CD4/CD8 ratio showed persistence of chest tomographic and functional alterations. By contrast, patients who decreased sPD-L2 displayed a complete lung recovery. sPD-L1, sTIM-3, and sGAL-9 increased significantly during aCOVID and decreased in all patients after 1-year follow-up. CONCLUSION: Increased sPD-L2 and an altered CD4/CD8 ratio after 12 months of aCOVID are associated with the persistence of lung lesions, suggesting that they may contribute to lung damage post-COVID-19.
Subject(s)
CD4-CD8 Ratio , COVID-19 , Lung , SARS-CoV-2 , Humans , COVID-19/immunology , COVID-19/blood , COVID-19/complications , Male , Female , Middle Aged , Aged , Lung/immunology , Lung/pathology , Lung/diagnostic imaging , SARS-CoV-2/immunology , Lung Diseases, Interstitial/immunology , Lung Diseases, Interstitial/blood , Biomarkers/blood , B7-H1 Antigen/blood , Respiratory Function Tests , Tomography, X-Ray Computed , Follow-Up Studies , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , AdultABSTRACT
Introduction: The proteolytic activity of A Disintegrin and Metalloproteinase 17 (ADAM17) regulates the release of tumor necrosis factor (TNF) and TNF receptors (TNFRs) from cell surfaces. These molecules play important roles in tuberculosis (TB) shaping innate immune reactions and granuloma formation. Methods: Here, we investigated whether single nucleotide polymorphisms (SNPs) of ADAM17 influence TNF and TNFRs levels in 224 patients with active TB (ATB) and 118 healthy close contacts. Also, we looked for significant associations between SNPs of ADAM17 and ATB status. TNF, TNFR1, and TNFR2 levels were measured in plasma samples by ELISA. Four SNPs of ADAM17 (rs12692386, rs1524668, rs11684747, and rs55790676) were analyzed in DNA isolated from peripheral blood leucocytes. The association between ATB status, genotype, and cytokines was analyzed by multiple regression models. Results: Our results showed a higher frequency of rs11684747 and rs55790676 in close contacts than ATB patients. Coincidentally, heterozygous to these SNPs of ADAM17 showed higher plasma levels of TNF compared to homozygous to their respective ancestral alleles. Strikingly, the levels of TNF and TNFRs distinguished participant groups, with ATB patients displaying lower TNF and higher TNFR1/TNFR2 levels compared to their close contacts. Conclusion: These findings suggest a role for SNPs of ADAM17 in genetic susceptibility to ATB.
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Background: The 5th edition of the World Health Organization Classification of Hematolymphoid Tumors recently defined immune deficiency/dysregulation (IDD)-associated-lymphoid-proliferations in HIV settings, where information is scarce, often gone under or misdiagnosed. Objectives: To describe the clinical picture, histopathology, and outcomes of IDD-associated-lymphoidproliferations Epstein-Barr virus+ (EBV) in people living with HIV without organ transplantation, antiretroviral therapy (ART) treated. Materials and Methods: HIV+ patients diagnosed with IDD-associated-lymphoid-proliferations seen at an academic medical center in Mexico from 2016 to 2019 were included. Immunohistochemical studies, in situ hybridization, and polymerase chain reaction analysis for EBV and LMP1 gene deletions were performed and correlated with clinical data. Results: We included 27 patients, all men who have sex with men, median age 36 years (interquartile range [IQR] 22-54). The median baseline CD4+ T cells were 113/mL (IQR 89-243), the CD4+/CD8+ ratio was 0.15 (IQR: 0.09-0.22), and the HIV viral load was 184,280 copies/mL (IQR: 76,000-515,707). Twenty patients (74.07%) had IDD-associated-lymphoid-proliferations hyperplasia plasma cell type EBV+, 3 (11.1%) had hyperplasia mononucleosis-like type (IM-type), 1 patient (3.70%) had florid follicular hyperplasia, 3 (11.1%) IDD-associated-lymphoid-proliferations polymorphic type, and there were 22 cases (81.4%) of synchronic Kaposi Sarcoma. Two patients were diagnosed with Hodgkin lymphoma following a second positron emission tomography-computed tomography scan-guided biopsy. The median follow-up was 228 weeks (IQR 50-269); 6 patients died (22.2%) of causes unrelated to IDD-associated-lymphoid-proliferations related. Conclusion: IDD-associated-lymphoid-proliferations EBV+ occured in severely immunosuppressed HIV+ patients, a high percentage of whom had concomitant Kaposi sarcoma. The prognosis was good in patients treated only with ART.
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Chronic infections induce CD4+ T-cells with cytotoxic functions (CD4 CTLs); at present, it is still unknown whether latent tuberculosis (LTB) and active tuberculosis (ATB) induce CD4 CTLs. Plasma and cells from four patient groups-uninfected contact (UC), LTB, and ATB (divided as sensitive [DS-TB]- or resistant [DR-TB]-drug)-were evaluated by flow cytometry, q-PCR, and proteomics. The data showed that ATB patients had an increased frequency of CD4+ T-cells and a decreased frequency of CD8+ T-cells. The latter displays an exhausted-like profile characterized by CD39, CD279, and TIM-3 expression. ATB had a high frequency of CD4 + perforin+ cells, suggesting a CD4 CTL profile. The expression (at the transcriptional level) of granzyme A, granzyme B, granulysin, and perforin, as well as the genes T-bet (Tbx21) and NKG2D (Klrk1), in enriched CD4+ T-cells, confirmed the cytotoxic signature of CD4+ T-cells during ATB (which was stronger in DS-TB than in DR-TB). Moreover, proteomic analysis revealed the presence of HSP70 (in DS-TB) and annexin A5 (in DR-TB), which are molecules that have been associated with favoring the CD4 CTL profile. Finally, we found that lipids from Mycobacterium tuberculosis increased the presence of CD4 CTLs in DR-TB patients. Our data suggest that ATB is characterized by exhausted-like CD8+ T-cells, which, together with a specific microenvironment, favor the presence of CD4 CTLs.
Subject(s)
CD4-Positive T-Lymphocytes , CD8-Positive T-Lymphocytes , Granzymes , Hepatitis A Virus Cellular Receptor 2 , Perforin , Tuberculosis , Humans , CD8-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/immunology , CD4-Positive T-Lymphocytes/metabolism , Male , Granzymes/metabolism , Granzymes/genetics , Granzymes/immunology , Perforin/metabolism , Perforin/genetics , Perforin/immunology , Adult , Female , Hepatitis A Virus Cellular Receptor 2/metabolism , Hepatitis A Virus Cellular Receptor 2/immunology , Tuberculosis/immunology , Tuberculosis/microbiology , Latent Tuberculosis/immunology , Latent Tuberculosis/microbiology , Middle Aged , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Cytotoxic/metabolism , Mycobacterium tuberculosis/immunology , T-Box Domain Proteins/metabolism , T-Box Domain Proteins/genetics , T-Box Domain Proteins/immunology , Antigens, CD/metabolism , Antigens, CD/immunology , Antigens, CD/genetics , NK Cell Lectin-Like Receptor Subfamily K/metabolism , NK Cell Lectin-Like Receptor Subfamily K/immunology , NK Cell Lectin-Like Receptor Subfamily K/genetics , Proteomics/methods , Antigens, Differentiation, T-Lymphocyte , ApyraseABSTRACT
Valganciclovir (VGC) was used in a randomized clinical trial in patients with disseminated Kaposi Sarcoma/human immunodeficiency virus (DKS/HIV) as add-on therapy to evaluate the proinflammatory axis tumor necrosis factor (TNF) and its receptors (TNFRs) in T cells. Two treatment schedules were used: an experimental regime (ER) and a conventional treatment (CT). Mononuclear cells from patients with DKS/HIV were obtained at baseline (W0), 4 (W4), and 12 weeks (W12). Ten DKS/HIV patients received CT (antiretroviral therapy [cART]) and 10 ER (valganciclovir [VGC] initially, plus cART at the fourth week). HIV+ without KS and HIV- patient groups were included as controls. Correlation between T-cell subsets and HHV-8 viral load (VL) and a multivariate linear regression was performed. Data showed that DKS/HIV patients have an increased frequency of CD8+ T cells, which display a high density of CD8 expression. The ER scheme increases naïve and central memory CD4+ T cells at W4 and W12 of follow-up and induces a balanced distribution of activated CD4+ T-cell subsets. Moreover, ER decreases solTNFR2 since W4 and CT decreased the transmembrane forms of TNF axis molecules. Although CT induces a positive correlation between HHV-8 VL and TNFRs, the use of ER positively correlates with TNF and TNFRs levels through follow-up and a moderate correlation with HHV-8 VL and TNF soluble levels. In conclusion, VGC, as an add-on therapy in DKS/HIV patients, gradually modulates the activation of CD4+ T-cell subsets and the TNF/TNFRs axis, suggesting a better regulation of the inflammatory status.
Subject(s)
HIV Infections , Sarcoma, Kaposi , Sulfonamides , Humans , Sarcoma, Kaposi/drug therapy , Sarcoma, Kaposi/metabolism , HIV Infections/metabolism , Valganciclovir/metabolism , Valganciclovir/therapeutic use , CD4-Positive T-Lymphocytes/metabolism , T-Lymphocyte Subsets , CD8-Positive T-Lymphocytes/metabolism , Tumor Necrosis Factor-alpha/metabolism , Viral LoadABSTRACT
T-cell Immunoglobulin and Mucin Domain 3 (TIM-3) is an immune checkpoint receptor known to regulate T-cell activation and has been targeted for immunotherapy in cancer and other diseases. However, its expression and function in other cell types, such as macrophages, are poorly understood. This study investigated TIM-3 expression in human macrophages polarized to M1 (stimulated with IFN-γ and LPS) and M2 (stimulated with IL-4 and IL-13) phenotypes using an in vitro model. Our results show that M1 macrophages have a lower frequency of TIM-3+ cells compared to M2 macrophages at 48 and 72 hr poststimulation. Additionally, we observed differential levels of soluble ADAM 10, an enzyme responsible for TIM-3 release, in the supernatants of M1 and M2 macrophages at 72 hr. We also found that the TIM-3 intracellular tail might associate with lymphocyte-specific protein 1 (LSP-1), a protein implicated in cell motility and podosome formation. These findings enhance our understanding of TIM-3 function in myeloid cells such as macrophages and may inform the development of immunotherapies with reduced immune-related adverse effects.
Subject(s)
Hepatitis A Virus Cellular Receptor 2 , Macrophages , Microfilament Proteins , Humans , Hepatitis A Virus Cellular Receptor 2/metabolism , Microfilament Proteins/metabolismABSTRACT
Introduction: Tuberculosis (TB) is a bacterial infection caused by Mycobacterium tuberculosis (M.tb). B cells are the central mediator of the humoral response; they are responsible for producing antibodies in addition to mediating other functions. The role of the cellular response during the TB spectrum by B cells is still controversial. Methods: In this study, we evaluated the distribution of the circulating B cell subsets in patients with active and latent TB (ATB and LTB, respectively) and how they respond to stimuli of protein or lipid from M.tb. Results: Here, we show that ATB patients show an immune fingerprinting. However, patients with drug-sensitive- (DS-TB) or drug-resistant- (DR-TB) TB have altered frequencies of circulating B cells. DS-TB and DR-TB display a unique profile characterized by high systemic levels of IFN-γ, IL-10, IgG, IgG/IgM ratio, and total B cells. Moreover, B cells from DR-TB are less efficient in producing IL-10, and both DS-TB and DR-TB produce less IFN-γ in response to M.tb antigens. Conclusion: These results provide new insights into the population dynamics of the cellular immune response by B cells against M.tb and suggest a fingerprinting to characterize the B-cell response on DR-TB.
Subject(s)
Latent Tuberculosis , Mycobacterium tuberculosis , Tuberculosis , Humans , Interleukin-10 , Interferon-gamma/metabolism , Immunoglobulin GABSTRACT
TNF and IFN-γ trigger cell damage during SARS CoV-2 infection; these cytokines can induce senescence and a cell death process called PANoptosis. This study included 138 vaccine-naïve COVID-19 patients, who were divided into four groups (Gp) according to the plasma level of TNF and IFN-γ (High [Hi] or Normal-Low [No-Low]), Gp 1: TNFHi/IFNγHi; Gp 2: TNFHi/IFNγNo-Low; Gp 3: TNFNo-Low/IFNγHi; and Gp 4: TNFNo-Low/IFNγNo-Low. Thirty-five apoptosis-related proteins and molecules related to cell death and senescence were evaluated. Our results showed that groups did not display differences in age and comorbidities. However, 81% of the Gp 1 patients had severe COVID-19, and 44% died. Notably, the p21/CDKN1A was increased in Gp 2 and Gp 3. Moreover, Gp 1 showed higher TNFR1, MLKL, RIPK1, NLRP3, Caspase 1, and HMGB-1 levels, suggesting elevated TNF and IFN-γ levels simultaneously activate diverse cell death pathways because it is not observed when only one of these cytokines is increased. Thus, high TNF/IFN-γ levels are predominant in severe COVID-19 status, and patients display cell alterations associated with the activation of diverse cell death pathways, including a possible senescent phenotype.
Subject(s)
COVID-19 , Interferon-gamma , Humans , Cell Death , Cytokines , Interferon-gamma/metabolism , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
Tuberculosis (TB) remains a public health problem worldwide and is one of the deadliest infectious diseases, only after the current COVID-19 pandemic. Despite significant advances in the TB field, there needs to be more immune response comprehension; for instance, the role played by humoral immunity is still controversial. This study aimed to identify the frequency and function of B1 and immature/transitional B cells in patients with active and latent TB (ATB and LTB, respectively). Here we show that LTB patients have an increased frequency of CD5+ B cells and decreased CD10+ B cells. Furthermore, LTB patients stimulated with mycobacteria's antigens increase the frequency of IFN-γ-producing B cells, whereas cells from ATB do not respond. Moreover, under the mycobacterial protein stimulus, LTB promotes a pro-inflammatory environment characterized by a high level of IFN-γ but also can produce IL-10. Regarding the ATB group, they cannot produce IFN-γ, and mycobacterial lipids and proteins stimulate only the IL-10 production. Finally, our data showed that in ATB, but not in LTB, B cell subsets correlate with clinical and laboratory parameters, suggesting that these CD5+ and CD10+ B cell subpopulations have the potential to be biomarkers to differentiate between LTB and ATB. In conclusion, LTB has increased CD5+ B cells, and these cells can maintain a rich microenvironment of IFN-γ, IL-10, and IL-4. In contrast, ATB only maintains an anti-inflammatory environment when stimulated with mycobacterial proteins or lipids.
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Mycobacterium tuberculosis (Mtb) modulates diverse cell death pathways to escape the host immune responses and favor its dissemination, a complex process of interest in pathogenesis-related studies. The main virulence factors of Mtb that alter cell death pathways are classified according to their origin as either non-protein (for instance, lipomannan) or protein (such as the PE family and ESX secretion system). The 38 kDa lipoprotein, ESAT-6 (early antigen-secreted protein 6 kDa), and another secreted protein, tuberculosis necrotizing toxin (TNT), induces necroptosis, thereby allowing mycobacteria to survive inside the cell. The inhibition of pyroptosis by blocking inflammasome activation by Zmp1 and PknF is another pathway that aids the intracellular replication of Mtb. Autophagy inhibition is another mechanism that allows Mtb to escape the immune response. The enhanced intracellular survival (Eis) protein, other proteins, such as ESX-1, SecA2, SapM, PE6, and certain microRNAs, also facilitate Mtb host immune escape process. In summary, Mtb affects the microenvironment of cell death to avoid an effective immune response and facilitate its spread. A thorough study of these pathways would help identify therapeutic targets to prevent the survival of mycobacteria in the host.
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INTRODUCTION: High HHV-8 viral load (VL) in Kaposi Sarcoma (KS) has been associated with Severe Immune Reconstitution Inflammatory Syndrome (Severe-IRIS-KS), which can occur after initiating cART, and leads to high mortality, particularly in patients with pulmonary involvement. We investigate if valganciclovir (as an anti-HHV-8 agent) initiated before cART reduces the mortality associated with Severe-IRIS-KS and the incidence of Severe-IRIS-KS. METHODS: Open-label parallel-group randomized clinical trial in AIDS cART naïve patients with disseminated KS (DKS) as defined by at least two of the following: pulmonary, lymph-node, or gastrointestinal involvement, lymphedema, or ≥30 skin lesions. In the experimental group (EG), patients received valganciclovir 900 mg BID four weeks before cART and continued until week 48; in the control group (CG), cART was initiated on week 0. Non-severe-IRIS-KS was defined as: an increase in the number of lesions plus a decrease of ≥one log10 HIV-VL, or an increase of ≥50cells/mm3 or ≥2-fold in baseline CD4+cells. Severe-IRIS-KS was defined as abrupt clinical worsening of KS lesions and/or fever after ruling out another infection following cART initiation, and at least three of the following: thrombocytopenia, anemia, hyponatremia, or hypoalbuminemia. RESULTS: 40 patients were randomized and 37 completed the study. In the ITT analysis, at 48 weeks, total mortality was the same in both groups (3/20), severe-IRIS-KS attributable mortality was 0/20 in the EG, compared with 3/20 in the CG (p = 0.09), similar to the per-protocol analysis: 0/18 in the EG, and 3/19 in the control group (p = 0.09). The crude incidence rate of severe-IRIS-KS was four patients developed a total of 12 episodes of Severe-IRIS-KS in the CG and two patients developed one episode each in the EG. Mortality in patients with pulmonary KS was nil in the EG (0/5) compared with 3/4 in the CG (P = 0.048). No difference was found between groups in the number of non-S-IRIS-KS events. Among survivors at week 48, 82% achieved >80% remission. CONCLUSIONS: Although mortality attributable to KS was lower in the EG the difference was not statistically significant.
Subject(s)
Anemia , HIV Infections , Herpesvirus 8, Human , Sarcoma, Kaposi , Humans , HIV Infections/complications , HIV Infections/drug therapy , Valganciclovir/therapeutic use , Antiretroviral Therapy, Highly Active , Anemia/complicationsABSTRACT
Staphylococcal enterotoxin B (SEB) is a protein produced by Staphylococcus aureus, which is toxic to humans. It is well known for its ability to stimulate the exacerbated activation of proinflammatory CD4+ T cells (Th1 profile), and in vitro studies have been conducted to understand its mechanism of action and its potential use as an immune therapy. However, the efficiency of the SEB1741 aptamer in blocking SEB has not been experimentally demonstrated. METHODS: Enrichment CD4+ T cells were stimulated with SEB, and as a blocker, we used the SEB1741 aptamer, which was previously synthesised by an "in silico" analysis, showing high affinity and specificity to SEB. The efficiency of the SEB1741 aptamer in blocking CD4+ T cell activation was compared with that of an anti-SEB monoclonal antibody. Flow cytometry and Bio-Plex were used to evaluate the T-cell function. RESULTS: In vitro, SEB induced the activation of CD4+ T cells and favoured a Th1 profile; however, the SEB1741 aptamer was highly efficient in decreasing the frequency of CD4+ T cells positive to ki-67 and CD69 cells, this means that proliferation and activation of CD4+ T cells was decreased. Moreover, the production of interleukin 2 (IL-2) and interferon-gamma (IFN-γ) was affected, suggesting that the Th1 profile is not present when the SEB1441 aptamer is used. Thus, the SEB1741 function was similar to that of anti-SEB. CONCLUSIONS: The SEB1741 aptamer is a valuable tool for blocking CD4+ T cell activation and the subsequent release of proinflammatory cytokines by SEB stimulation.
Subject(s)
CD4-Positive T-Lymphocytes , Enterotoxins , Humans , Enterotoxins/metabolism , Cytokines/metabolism , Staphylococcus aureus , Lymphocyte ActivationABSTRACT
Platelets play a major role in coagulation and hemostasis; evidence supports the hypothesis that they also contribute to immunological processes. Increased platelet counts have been associated with poor prognosis in tuberculosis (TB). Platelet-monocyte aggregates have been reported in patients with TB, but it is still unclear if only one monocyte subpopulation is correlated to the platelet count; moreover, the platelet-monocyte axis has not been studied during latent tuberculosis (LTB). In this study, mononuclear cells and plasma were obtained from patients diagnosed with active drug-sensitive TB (DS-TB, n = 10) and LTB (n = 10); cytokines and growth factors levels associated to platelets were evaluated, and correlations with monocyte subpopulations were performed to identify a relationship between them, as well as an association with the degree of lung damage. Our data showed that, compared to LTB, DS-TB patients had an increased frequency of platelets, monocytes, and neutrophils. Although DS-TB patients showed no significant difference in the frequency of classical and non-classical monocytes, the classical monocytes had increased CD14 intensity of expression and frequency of TLR-2+. Furthermore, the plasma levels of angiogenic factors such as vascular endothelial growth factor (VEGF-A), platelet-derived growth factor (PDGF-BB), and platelet factor-4 (PF4), and pro-inflammatory cytokines like interleukin 6 (IL-6), interleukin 1 beta (IL-1ß), and interferon-γ-inducible protein 10 (IP-10) were increased in DS-TB patients. In addition, PF-4 and VEGF-A correlated positively with the frequency of classical monocytes and the platelet count. Using a principal component analysis, we identified four groups of DS-TB patients according to their levels of pro-inflammatory cytokines, angiogenic factors, and degree of lung damage. This study establishes that there is a correlation between VEGF-A and PF4 with platelets and classical monocytes during active TB, suggesting that those cell subpopulations are the major contributors of these molecules, and together, they control the severity of lung damage by amplification of the inflammatory environment.
Subject(s)
Monocytes , Tuberculosis , Humans , Platelet Factor 4/metabolism , Vascular Endothelial Growth Factor A/metabolism , Tuberculosis/metabolism , Cytokines/metabolism , Inflammation/metabolism , Immunologic Factors/metabolismABSTRACT
The PDZ (PSD95, Dlg and ZO-1) genes encode proteins that primarily function as scaffolds of diverse signaling pathways. To date, 153 PDZ genes have been identified in the human genome, most of which have multiple protein isoforms widely studied in epithelial and neural cells. However, their expression and function in immune cells have been poorly studied. Herein, we aimed to assess the transcriptional profiles of 83 PDZ genes in human macrophages (Mɸ) and dendritic cells (DCs) and changes in their relative expression during cell PRR stimulation. Significantly distinct PDZ gene transcriptional profiles were identified under different stimulation conditions. Furthermore, a distinct PDZ gene transcriptional signature was found in Mɸ and DCs under the same phagocytic stimuli. Notably, more than 40 PDZ genes had significant changes in expression, with potentially relevant functions in antigen-presenting cells (APCs). Given that several PDZ proteins are targeted by viral products, our results support that many of these proteins might be viral targets in APCs as part of evasion mechanisms. Our results suggest a distinct requirement for PDZ scaffolds in Mɸ and DCs signaling pathways activation. More assessments on the functions of PDZ proteins in APCs and their role in immune evasion mechanisms are needed.
Subject(s)
Immune Evasion , Macrophages , Dendritic Cells , Humans , Macrophages/metabolism , Signal TransductionABSTRACT
Mycobacterium tuberculosis has developed diverse mechanisms to survive inside phagocytic cells, such as macrophages. Phagocytosis is a key process in eliminating invading pathogens; thus, M. tuberculosis efficiently disrupts phagosome maturation to ensure infection. However, inflammatory cytokines produced by macrophages in response to early M. tuberculosis infection are key to promoting bacterial clarification. IFN-γ enhances M. tuberculosis engulfment and destruction by reprogramming macrophages from phagocytosis to macropinocytosis. Here, we show that the transcription factor Krüppel-like factor 10 (Klf10) plays a positive role in M. tuberculosis survival and infection by negatively modulating IFN-γ levels. Naïve Klf10-deficient macrophages produce more IFN-γ upon stimulation than wild-type macrophages, thus enhancing bacterial uptake and bactericidal activity achieved by macropinocytosis. Moreover, Klf10â»/ â» macrophages showed cytoplasmic distribution of coronin 1 correlated with increased pseudopod count and length. In agreement with these observations, Klf10â»/ â» mice showed improved bacterial clearance from the lungs and increased viability. Altogether, our data indicate that Klf10 plays a critical role in M. tuberculosis survival by preventing macrophage reprogramming from phagocytosis to macropinocytosis by negatively regulating IFN-γ production upon macrophage infection.
Subject(s)
Kruppel-Like Transcription Factors , Macrophages , Mycobacterium tuberculosis , Tuberculosis , Animals , Early Growth Response Transcription Factors , Interferon-gamma , Kruppel-Like Transcription Factors/genetics , Macrophages/microbiology , Mice , Phagocytosis , PinocytosisABSTRACT
In recent years, a growing body of evidence has shown the presence of a subpopulation of macrophages that express CD3, especially in the context of mycobacterial infections. Despite these findings, the function of these cells has been poorly understood. Furthermore, the low frequency of CD3+ macrophages in humans limits the study of this subpopulation. This work aimed to evaluate the expression of CD3 in a murine macrophage cell line and its potential for the study of CD3 signaling. The murine macrophage cell line RAW was used to evaluate CD3 expression at the transcriptional and protein levels and the effect of in vitro infection with the Mycobacterium bovis Bacillus Calmette-Guérin (BCG) on these. Our data showed that RAW macrophages express CD3, both the ε and ζ chains, and it is further increased at the transcriptional level after BCG infection. Furthermore, our data suggest that CD3 can be found on the cell surface and intracellularly. However, this molecule is internalized constantly, mainly after activation with anti-CD3 stimulus, but interestingly, it is stably maintained at the transcriptional level. Finally, signaling proteins such as NFAT1, c-Jun, and IKK-α are highly expressed in RAW macrophages. They may play a role in the CD3-controlled signaling pathway to deliver inflammatory cytokines such as TNF and IL-6. Our study provides evidence to support that RAW cells are a suitable model to study the function and signaling of the CD3 complex in myeloid cells.
Subject(s)
BCG Vaccine , Mycobacterium bovis , Animals , BCG Vaccine/pharmacology , Humans , Macrophages/metabolism , Mice , Mycobacterium bovis/physiology , Signal Transduction , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Macrophages are necessary to eliminate pathogens. However, some pathogens have developed mechanisms to avoid the immune response. One of them is modulating the cell death mechanism to favor pathogen survival. In this study, we evaluated if virulent Mycobacterium tuberculosis (M. tb) can simultaneously activate more than one cell death mechanism. We infected human monocyte-derived macrophages (MDM) in vitro with avirulent (H37Ra) and virulent (H37Rv) strains, and then we measured molecules involved in apoptosis, necroptosis, and pyroptosis. Our data showed that H37Rv infection increased the BCL-2 transcript and protein, decreased the BAX transcript, and increased phosphorylated BCL-2 at the protein level. Moreover, H37Rv infection increased the expression of the molecules involved in the necroptotic pathway, such as ASK1, p-38, RIPK1, RIPK3, and caspase-8, while H37Ra increased caspase-8 and decreased RIPK3 at the transcriptional level. In addition, NLRP3 and CASP1 expression was increased at low MOI in both strains, while IL-1ß was independent of virulence but dependent on infection MOI, suggesting the activation of pyroptosis. These findings suggest that virulent M. tb inhibits the apoptosis mediated by BCL-2 family molecules but, at the same time, increases the expression of molecules involved in apoptosis, necroptosis, and pyroptosis at the transcriptional and protein levels, probably as a mechanism to avoid the immune response and guarantee its survival.
ABSTRACT
Idiopathic pulmonary fibrosis (IPF) is a progressive and irreversible lung disease of unknown etiology. Myofibroblasts are organized in peculiar subepithelial fibroblasts foci (FF), where they abnormally persist and exclude lymphocytes by unclear mechanisms. FF are the source of an excessive extracellular matrix, which results in progressive stiffening and destruction of the lung architecture. We hypothesized that the absence of T cells inside the FF could be related, at least partially, to an inefficient function of lymphocytes induced by IPF fibroblasts. Here, we evaluated the effect of a supernatant from IPF fibroblasts on T-cell apoptosis and migration capacity. Data showed that IPF fibroblasts secrete pro-apoptotic molecules (both from extrinsic and intrinsic pathways), generating a microenvironment that induces apoptosis of T cells at 3 h of culture, despite a weak anti-apoptotic profile exhibited by these T cells. At 24 h of culture, the supernatants from both IPF and control fibroblasts provoked T-cell death. However, at this time of culture, IPF fibroblasts caused a marked decrease in T-cell migration; in contrast, control lung fibroblasts induced an increase of T-cell migration. The reduction of T-cell migratory capacity provoked by IPF fibroblasts was associated with a negative regulation of RHOA and ROCK, two essential GTPases for migration, and was independent of the expression of chemokine receptors. In conclusion, our findings demonstrate that IPF fibroblasts/myofibroblasts induce apoptosis and affect T-cell migration, revealing a mechanism involved in the virtual absence of T lymphocytes inside the FF.
Subject(s)
Idiopathic Pulmonary Fibrosis , Apoptosis , Fibroblasts/metabolism , Humans , Idiopathic Pulmonary Fibrosis/metabolism , Myofibroblasts/metabolism , T-Lymphocytes/metabolismABSTRACT
Human herpesvirus-8 infection (HHV-8) is the causative agent of Kaposi sarcoma (KS) and is highly prevalent among people living with HIV (KS/HIV). It has been reported that valganciclovir (VGC) reduces HHV-8 replication in KS/HIV patients. However, currently it is unclear if VGC modifies the frequency and induces changes in markers of immune regulation of immune cells necessary to eliminate HHV8-infected cells, such as Natural Killer (NK) and NK T cells (NKT). This study evaluated the effect of VGC used as antiviral HHV8 therapy in KS patients on the frequency of NK and NKT subpopulations based on the CD27 and CD57 expression, and the immunosenescence markers, PD-1 and KLRG1. Twenty KS/HIV patients were followed-up at baseline (W0), 4 (W4), and 12 weeks (W12) of the study protocol. Among them, 10 patients received a conventional treatment scheme (CT), solely antiretroviral therapy (ART), and 10 patients received a modified treatment regime (MT), including VGC plus ART. In both groups, bleomycin/vincristine was administrated according to the treating physician's decision. The soluble levels of IL-15, PD-L1, PD-L2, and E-cadherin were quantified across the follow-up. Our results showed that the higher IL-15 levels and lower NK frequencies cells in KS/HIV patients reach almost normal values with both treatments regimes at W12. CD27+ NK and NKT cell frequencies increased since W4 on KS/HIV patients with MT. Furthermore, PD-1 expression decreased while KLRG1 increased on NK and NKT subpopulations at W12, and it is accompanied by increased PD-L1 plasma level since W4. Our study highlights the disruption of NK and NKT subpopulations in patients with KS/HIV and explores VGC treatment's contribution to immune reconstitution during the first weeks of treatment.