Tuberculosis Antigen Stimulation Research Articles

Tuberculous Pleurisy in a Patient with a History of Breast Cancer: Diagnostic Challenges and Management Options

Background: Tuberculosis (TB) and cancer are increasingly prevalent diseases that can be challenging to diagnose due to similarities in clinical and radiological findings. This case report describes a 46-year-old woman with a history of breast cancer who developed tuberculous pleurisy (TP). Case presentation: A 46-year-old woman who underwent mastectomy and chemotherapy for BC in 2023 had hypertension, but no history of TB. The patient presented with dry cough, fever, and stomach discomfort, with an oxygen saturation level of 60%, but no respiratory distress before their appointment. Microscopy and culture tests were negative for Mycobacterium TB, A positive result was observed with an IFN-γ level of 0.35 IU/ml and 26% of the negative control after TB antigen stimulation. Histological analysis using hematoxylin and eosin staining showed Langhans giant cells, epithelioid cell granulomas, caseous necrosis, and necrotic foci. These findings indicate granulomatous inflammation with no signs of malignancy. Following the diagnosis, the patient received a daily dose of 300 mg isoniazid, 600 mg rifampin, 1500 mg pyrazinamide, and 10 mg pyridoxine for six months without any adverse effects. Conclusion: Physicians must employ a combination of diagnostic techniques, including morphological and microbiological confirmation, to accurately diagnose pleural effusion in this patient population.

Distinct TB-antigen stimulated cytokine profiles as predictive biomarkers for unfavorable treatment outcomes in pulmonary tuberculosis.

The assessment of tuberculosis (TB) treatment outcomes predominantly relies on sputum culture conversion status. To enhance treatment management, it is crucial to identify non-sputum-based biomarkers that can predict unfavorable outcomes. Cytokines are widely studied as diagnostic biomarkers for active TB. However, their potential as indicators for unfavorable treatment outcomes remains uncertain. This study was conducted within a well-characterized cohort comprising newly diagnosed patients with drug-sensitive pulmonary TB, confirmed through sputum smear and culture positivity. Our objective was to elucidate the TB antigen-stimulated cytokine profile at pre-treatment and at 2 months into anti-TB treatment (ATT) in patients with unfavorable treatment outcomes (cases, n = 27) in comparison to recurrence-free, microbiologically cured controls (n = 31). Whole blood was stimulated with TB antigens using the QuantiFERON In-tube gold method, and plasma supernatants were subjected to a panel of 14 cytokine measurements. In our study, pre-treatment analysis revealed that eight cytokines (IL-2, IFN-γ, TNF-α, IL-6, IL-10, IL-17A, IL-18, and GM-CSF) were significantly elevated at baseline in cases compared to cured controls, both in unstimulated conditions and following TB antigen (CFP10, ESAT6, and TB7.7) stimulation. A similar pattern was observed at the 2-month mark of ATT, with eight cytokines (IL-2, IL-10, IL-13, IFN-γ, IL-6, IL-12p70, IL-17A, and TNF-α) showing significant differences between the groups. Importantly, no variations were detected following mitogen stimulation, underscoring that these distinctive immune responses are primarily driven by TB-specific antigens. Our findings indicate that individuals with unfavorable TB treatment outcomes display a characteristic cytokine profile distinct from TB-cured patients, even before commencing ATT. Therefore, the levels of specific cytokine pre-treatment and at the 2-month point in the course of treatment may serve as predictive immune markers for identifying individuals at risk of unfavorable TB treatment outcomes, with these responses being predominantly influenced by TB-specific antigens.

Combination of HLA-DR on Mycobacterium tuberculosis-Specific Cells and Tuberculosis Antigen/Phytohemagglutinin Ratio for Discriminating Active Tuberculosis From Latent Tuberculosis Infection.

BackgroundNovel approaches for tuberculosis (TB) diagnosis, especially for distinguishing active TB (ATB) from latent TB infection (LTBI), are urgently warranted. The present study aims to determine whether the combination of HLA-DR on Mycobacterium tuberculosis (MTB)-specific cells and TB antigen/phytohemagglutinin (TBAg/PHA) ratio could facilitate MTB infection status discrimination.MethodsBetween June 2020 and June 2021, participants with ATB and LTBI were recruited from Tongji Hospital (Qiaokou cohort) and Sino-French New City Hospital (Caidian cohort), respectively. The detection of HLA-DR on MTB-specific cells upon TB antigen stimulation and T-SPOT assay were simultaneously performed on all subjects.ResultsA total of 116 (54 ATB and 62 LTBI) and another 84 (43 ATB and 41 LTBI) cases were respectively enrolled from Qiaokou cohort and Caidian cohort. Both HLA-DR on IFN-γ+TNF-α+ cells and TBAg/PHA ratio showed discriminatory value in distinguishing between ATB and LTBI. Receiver operator characteristic (ROC) curve analysis showed that HLA-DR on IFN-γ+TNF-α+ cells produced an area under the ROC curve (AUC) of 0.886. Besides, TBAg/PHA ratio yield an AUC of 0.736. Furthermore, the combination of these two indicators resulted in the accurate discrimination with an AUC of 0.937. When the threshold was set as 0.36, the diagnostic model could differentiate ATB from LTBI with a sensitivity of 92.00% and a specificity of 81.82%. The performance obtained in Qiaokou cohort was further validated in Caidian cohort.ConclusionsThe combination of HLA-DR on MTB-specific cells and TBAg/PHA ratio could serve as a robust tool to determine TB disease states.

IL-2 Restores T-Cell Dysfunction Induced by Persistent Mycobacterium tuberculosis Antigen Stimulation

Tuberculosis (TB) is a chronic disease mainly caused by Mycobacterium tuberculosis. The function of T cells usually decreased and even exhausted in severe TB such as multiple drug resistant TB (MDR-TB), which might lead to the failure of treatment in return. The mechanism of T cell dysfunction in TB is still not clear. In this study we set up a mouse model of T cell dysfunction by persistent M. tuberculosis antigen stimulation and investigated the therapeutic role of interleukin 2 (IL-2) in it. C57BL/6 mice were primed with Mycobacterium bovis Bacillus Calmette-Guérin (BCG) and boosted repeatedly with a combination of M. tuberculosis fusion proteins Mtb10.4-HspX (MH) plus ESAT6-Ag85B-MPT64 <190−198>-Mtb8.4-Rv2626c (LT70) or MH plus ESAT6 and CFP10 with adjuvant of N, N′-dimethyl-N, N′-dioctadecylammonium bromide (DDA) plus polyinosinic-polycytidylic acid (Poly I:C). Following persistent antigen stimulation, the mice were treated with IL-2 and the therapeutic effects were analyzed. The results showed that compared with the mice that received transient antigen stimulation (boost twice), persistent antigen stimulation (boost more than 10 times) resulted in decrease of antigen specific IFN-γ and IL-2 production, reduction of memory CD8+ T cells, over-expression of immune checkpoint programmed cell death protein 1 (PD-1), and impaired the protective immunity against bacterial challenge. Treating the T cell functionally exhausted mice with IL-2 restored antigen-specific T cell responses and protective efficacy. In conclusion, persistent stimulation with M. tuberculosis antigens induced T cell dysfunction, which could be restored by complement of IL-2.

M. tuberculosis infection and antigen specific cytokine response in healthcare workers frequently exposed to tuberculosis

Tuberculosis (TB) is the leading cause of death due to an infectious agent, but only a small fraction of those infected develop the disease. Cytokines are involved in the mediation and regulation of immunity, and their secretion patterns may reflect the infection status. To increase our understanding of immune response to M. tuberculosis infection, we conducted a cross-sectional study investigating M. tuberculosis infection status and comparing the release profiles of cytokines GM-CSF, IFN-γ, IL-1β, IL-10, IL-12 (p70), IL-2, IL-4, IL-5, IL-6, IL-8, TNF-α, in community controls (CCs) and healthy healthcare workers (HCWs) highly exposed to TB. Among HCWs and CCs, the probability of latent M. tuberculosis (LTB+) infection was respectively 5.4 (p = 0.002) and 3.4 (p = 0.006) times higher in men than women. The odds ratio of LTB infection was 4 times higher among HCWs in direct contact with active TB patients than other HCW (p = 0.01). Whole blood supernatant cytokine responses to M. tuberculosis antigens showed differential pro-inflammatory responses between HCWs and CCs. CCsLTB− had higher IL-1β responses than HCWsLTB− (p = 0.002). HCWsLTB+ had significantly higher IL-8 responses to M. tuberculosis antigens than HCWsLTB− (p = 0.003) and CCsLTB− (p = 0.015). HCWsLTB+/− showed weak but positive TNF-α responses to M. tuberculosis antigen stimulation compared to CCsLTB+/− (p ≤ 0.015). Looking at T-helper (1 and 2) responses, HCWsLTB+ and CCsLTB+ had significantly higher IFN-γ and IL-2 responses compared to HCWsLTB− and CCsLTB− (p < [0.0001–0.003]). Also, TB antigen induced IL-5 secretion was significantly higher in HCWsLTB+ and CCsLTB+ than in non-infected CCsLTB− (p < [0.005–0.04]). M. tuberculosis antigen specific responses in HCWsLTB+ varied based on active TB exposure gradient. HCWsLTB+ who were highly exposed to active TB (≥3 hours per day) had significantly higher IFN-γ and IL-8 responses (p ≤ 0.02) than HCWs LTB+ not in direct contact with active TB patients. HCWsLTB+ working with active TB patients for 5 to 31 years had a significantly enhanced secretion of proinflammatory cytokines (GM-CSF, IFN-γ, IL-1β, IL-2, IL-6, IL-8, IL-12p70, TNF-α) compared to HCWsLTB− (p < [0.0001–0.01]). Secretion of anti-inflammatory/Th2 cytokines IL-5 and IL-10 was also higher in HCWsLTB+ than HCWsLTB−. In conclusion, LTBI individuals controlling the M. tuberculosis infection have an enhanced TB specific Th1-cytokines/proinflammatory response combined with selected Th2 type/anti-inflammatory cytokines induction.

The exhausted CD4+CXCR5+ T cells involve the pathogenesis of human tuberculosis disease

ObjectivesCD4+CXCR5+ T cells have previously been established. However, their decreased frequency during tuberculosis (TB) disease is only partially understood. The aim of this study was to explore the depletion of CD4+CXCR5+ T cells in human TB. MethodsThe frequency and function of CD4+CXCR5+ T cells were evaluated in active TB (ATB) patients and healthy control subjects. The function of CD4+CXCR5+ T cells was determined after blockade of inhibitory receptors. ResultsThe frequency of CD4+CXCR5+ T cells was decreased in ATB patients. The expression of activation markers (HLA-DR and ICOS) and inhibitory receptors (Tim-3 and PD-1) on CD4+CXCR5+ T cells was increased in the ATB group. TB-specific antigen stimulation induced higher expression of inhibitory receptors than phytohemagglutinin stimulation in the ATB group. In contrast, TB antigen stimulation did not induce a significantly increased expression of IL-21 and Ki-67 on CD4+CXCR5+ T cells. However, blockade of inhibitory receptors Tim-3 and PD-1 not only increased the frequency of CD4+CXCR5+ T cells, but also restored their proliferation and cytokine secretion potential. ConclusionsThe increased expression of inhibitory receptors involves the depletion of CD4+CXCR5+ T cells, and blockade of inhibitory receptors can restore the function of CD4+CXCR5+ T cells in ATB patients.

Coincident Type 2 diabetes mellitus modulates the systemic and antigen – specific induction of Type 1, Type 17, and other cytokines in both active and latent tuberculosis

Abstract Type 2 diabetes mellitus (DM) is known to be a major risk factor for the development of active tuberculosis, although its influence on active and latent Mycobacterium tuberculosis infection and disease remains poorly characterized. To identify the influence of coincident DM on cytokine levels in active pulmonary tuberculosis and latent tuberculosis, we examined circulating levels of a panel of cytokines in the plasma of individuals with PTB-DM or PTB alone and LTB-DM or LTB alone. PTB-DM is characterized by elevated circulating levels of type 1 (IFNγ, TNFα, and IL-2), type 2 (IL-5), type 17 (IL-17A) but decreased circulating levels of IL-10 family of cytokines (IL-19, IL-20, IL-22 and IL-24). This was associated with increased systemic levels of other pro-inflammatory cytokines (IL-1β, IL-6, and IL-18) and an anti-inflammatory cytokine (IL-10) but not type 1 IFNs. In contrast, LTB-DM is characterized by diminished circulating levels of type 1 (IFNγ, IL-2, and TNFα), type 17 (IL-17F), IL-10 family (IL-10, IL-19, IL-20 and IL-24) and other pro-inflammatory (IL-1β and IL-18) cytokines. Moreover, PTB-DM showed increased levels of pro-inflammatory cytokines upon tuberculosis antigen stimulation whereas LTB-DM showed diminished pro-inflammatory cytokines upon tuberculosis antigen stimulation. Our data reveal that PTB-DM is characterized by heightened cytokine responsiveness, indicating that chronic inflammation underlying DM potentially contributes to increased immune pathology. On the other hand, LTB-DM is characterized by diminished production of cytokines, implicated in the control of M. tuberculosis, allowing for a potential immunological mechanism that could account for the increased risk of active tuberculosis.

Genetic polymorphisms of CCL2 associated with susceptibility to latent tuberculous infection in Thailand.

The extent to which individuals exhibit genetic susceptibility to tuberculosis (TB) is still unclear. Genetic variations in chemokine genes might influence the early clearance of Mycobacterium tuberculosis, affecting TB susceptibility. To study single nucleotide polymorphisms (SNPs) of chemokine genes CCL2, CXCL9, CXCL10 and CXCL11, and their association with TB susceptibility. Of 248 participants enrolled, 49 had active TB, 43 had latent tuberculous infection (LTBI) and 156 were non-infected, including 24 healthy controls with no known TB exposure. These populations were divided into two groups based on TB exposure: susceptible (n = 92) and resistant (early clearance) (n = 132). Only CCL2 SNPs (-2518A/G) were significantly associated with increased TB susceptibility. Based on adjusted multivariate analysis, persons with the GG genotype at this SNP were twice as susceptible to TB as those with the AA genotype (P = 0.018, OR 2.880, 95%CI 1.201-6.903). Risk of LTBI was three times higher among those with GG (P = 0.003, OR 3.358, 95%CI 1.525-7.396 for AA+AG vs. GG and P = 0.012, OR 3.706, 95%CI 1.340-10.254 for AA vs. GG). Persons with the GG genotype produced significantly lower CCL2 levels in response to M. tuberculosis antigen stimulation (AA+AG vs. GG, P = 0.038). The CCL2 polymorphism (-2518A/G) was associated with susceptibility to LTBI in a North-East Thai populations.

Aplicación y utilidad actual de las técnicas de interferón-γ en el diagnóstico de la tuberculosis

Utility of the in-vitro immunodiagnostic methods, based on the detection of interferon-γ released by T-cells after specific Mycobacterium tuberculosis antigen stimulation (IGRA), has been an improvement in the accuracy of the latent tuberculosis infection diagnosis. IGRA have a well-known higher specificity than the tuberculin skin testing (TST). Moreover, they can obtain a larger number of positive results than the TST in immunocompromised patients. IGRA have shown a high correlation with M. tuberculosis exposure, but their positive and negative predictive value are similar than those obtained by TST. Nevertheless, given their high specificity, they allow reducing number of unnecessary preventive treatments. In addition, these in-vitro techniques are less affected than TST by the different immunosuppressing status. In this review is discussed up-to-date applicability of IGRA in different patient groups: contact studies, pediatric population, immunosuppressed patients, health care workers and active tuberculosis patients. Furthermore, it has been included possible future directions for latent tuberculosis infection and active tuberculosis diagnosis.

Regulation network of serum cytokines induced by tuberculosis-specific antigens reveals biomarkers for tuberculosis diagnosis.

In this study, we identified potential serum biomarkers for the diagnosis of active tuberculosis (TB) and screening for latent TB infections (LTBIs). Peripheral blood samples from 40 healthy individuals, 40 patients with TB, and 40 LTBI individuals were stimulated with the TB-specific antigens ESAT-6 and CFP-10. Human inflammatory cytokine arrays were used to detect the expression of inflammatory cytokines. Cytokines with significant changes were screened to construct a cytokine regulation network. The levels of the cytokines CCL1 (I-309), CXCL9 (MIG), IL-10, IL-6, CSF2, CSF3, IL-8, IL-1α, IL-7, TGF-β1, CCL2, IL-2, IL-13, and TNFα were significantly upregulated in the active TB group. The levels of CCL3, IL-1β, CCL8, IFNγ, and CXCL10 were significantly increased in the TB groups compared to those in the healthy control group. sTNF RII was upregulated in the LTBI group. CCL4 and MIP1d were significantly increased in all groups.The upregulated cytokines were mainly found in the IFNγ and IL-1α regulatory networks. Importantly, we found that CXCL10 (IP-10), CCL3, CCL8, and IL-1β may be more suitable than IFNγ for active or latent TB infection screening. Furthermore, we found that levels of CCL1 (I-309), CXCL9 (MIG), IL-10, IL-6, CSF2, CSF3, IL-8, IL-1α, IL-7, TGF-β1, CCL2, IL-2, and IL-13 after TB antigen stimulation may help distinguish between active and latent TB.

Expansion of Pathogen-Specific T-Helper 1 and T-Helper 17 Cells in Pulmonary Tuberculosis With Coincident Type 2 Diabetes Mellitus

Type 2 diabetes mellitus (DM) is a major risk factor for the development of active pulmonary tuberculosis, although the immunological mechanisms underlying this interaction remain unexplored. The influence of poorly controlled diabetes on pathogen-specific T-helper 1 (Th1) and T-helper 17 (Th17) responses have not been examined. To identify the role of Th1 and Th17 cells in tuberculosis with coincident DM, we examined mycobacteria-specific immune responses in the whole blood of individuals who had tuberculosis with DM and compared them to those in individuals who had tuberculosis without DM. Tuberculosis coincident with DM is characterized by elevated frequencies of monofunctional and dual-functional CD4(+) Th1 cells following Mycobacterium tuberculosis antigen stimulation and elevated frequencies of Th17 subsets at both baseline and following antigen stimulation. This was associated with increased systemic (plasma) levels of both Th1 and Th17 cytokines and decreased baseline frequencies of natural regulatory T cells but not interleukin 10 or transforming growth factor β. Therefore, our data reveal that tuberculosis in persons with DM is characterized by elevated frequencies of Th1 and Th17 cells, indicating that DM is associated with an alteration in the immune response to tuberculosis, leading to a biased induction of Th1- and Th17-mediated cellular responses and likely contributing to increased immune pathology in M. tuberculosis infection.

CD137 differentially regulates innate and adaptive immunity against Mycobacterium tuberculosis

Protective immunity against Mycobacterium tuberculosis is primarily mediated by the interaction of antigen-specific T cells and antigen presenting cells, which often depends on the interplay of cytokines produced by these cells. Costimulatory signals represent a complex network of receptor–ligand interactions that qualitatively and quantitatively influence immune responses. Thus, here we investigated the function of CD137 and CD137L, molecules known to have a central role in immune regulation, during human tuberculosis (TB). We demonstrated that M. tuberculosis antigen stimulation increased both CD137 and CD137L expression on monocytes and NK cells from TB patients and healthy donors, but only up-regulated CD137 on T lymphocytes. Blockage of the CD137 pathway enhanced the levels of interferon (IFN)-γ and tumor necrosis factor (TNF)-α produced by monocytes and NK against M. tuberculosis. In contrast, CD137 blockage significantly decreased the specific degranulation of CD8+ T cells and the percentage of specific IFN-γ and TNF-α producing lymphocytes against the pathogen. Furthermore, inhibition of the CD137 pathway markedly increased T-cell apoptosis. Taken together, our results demonstrate that CD137:CD137L interactions regulate the innate and adaptive immune response of the host against M. tuberculosis.

Elevated expression of Tim-3 on CD8 T cells correlates with disease severity of pulmonary tuberculosis

T cell immunoglobulin and mucin domain-containing molecule 3 (Tim-3) plays an important role in regulating T cell responses and induction of peripheral tolerance. In this study, the association of Tim-3 with active pulmonary tuberculosis (TB) was investigated. Tim-3 expression on CD8 T cells and its influence on production of effector molecules and proliferation of CD8 T cells were examined. Pulmonary TB patients had significantly elevated Tim-3 expression on total CD8 T cells (P<0.0001) and on antigen-specific CD8 T cells (P=0.0028) compared with tuberculin-positive healthy controls. The expression of Tim-3 on CD8 T cells was significantly higher in smear/culture-positive TB patients than smear/culture-negative TB patients (P<0.0001). Tim-3 expressing CD8 T cells had reduced production of IFN-γ, decreased degranulation and lower proliferation in response to Mycobacterium tuberculosis antigen stimulation. Blocking of Tim-3 signaling led to significantly increased production of IFN-γ. The elevated expression of Tim-3 on CD8 T cells correlates with functional defects of CD8 T cells and disease severity of pulmonary TB.

Indeterminate Interferon-γ Release Assay Results in Children

Indeterminate Interferon-γ Release Assay Results in Children

Quantitative evaluation of T-cell response after specific antigen stimulation in active and latent tuberculosis infection in adults and children

We have evaluated the quantitative T-cell response after specific Mycobacterium tuberculosis antigen stimulation in active tuberculosis (TB) and latent TB infection (LTBI) patients. In adults, the median number of T cells after RD1 antigen stimulation was significantly higher in active TB patients than in LTBI patients. In children, the number of responder T cells against the specific antigens was higher in active TB than in LTBI patients, although the differences were not significant. In summary, in patients with suspected clinical TB, although there is overlapping in the number of responder T cells between both groups, a T-cell count above the described threshold could suggest active TB, especially in patients with a high probability of having active TB and low probability of having LTBI. In addition, the results are consistent with the current evidence that T-cell response may indicate mycobacterial burden and disease activity.