Dataset Information

Secretion of IFN-? Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis.

ABSTRACT: In this study, we investigated the role of a matricellular protein galectin-9 (Gal-9) in pleural effusion related to tuberculosis (TB). Plasma and pleural fluid of a patient with extrapulmonary TB were analyzed for cytokine content by ELISA and Luminex. Peripheral blood mononuclear cells (PBMCs) and pleural fluid cells (PFCs) were examined for interferon-? (IFN-?) secretion by the enzyme-linked immunospot (ELISPOT) assay or IFN-? ELISA, for apoptosis and necrosis by Cell Death Detection ELISA, and also underwent cell sorting. The results indicate that compared to plasma, pleural fluid had increased levels of IFN-? (1.6 vs. 55.5 pg/mL), IL-10, IL-12p40, vascular endothelial growth factor (VEGF), and Gal-9 (3.0 vs. 936.0 pg/mL), respectively. PFCs culture supernatant exhibited higher concentration of Gal-9 compared to PBMCs in culture, consistent with enriched Gal-9 staining in the granuloma that is in closer vicinity to PFCs compared to PBMCs. PFCS displayed higher IFN-? secretion after stimulation with TB antigens ESAT-6/CFP-10. Furthermore, in PFCs, Gal-9 alone could stimulate IFN-? synthesis in culture or ELISPOT, which was inhibited by a Gal-9 antagonist lactose, and which may promote apoptosis and necrosis. These findings suggest that Gal-9 could modulate immune responses and participate in immunopathology of pleural effusion during TB.

SUBMITTER: Zhao J

PROVIDER: S-EPMC5535875 | biostudies-literature | 2017 Jun

REPOSITORIES: biostudies-literature

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Secretion of IFN-γ Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis.

Zhao Jingge J Shiratori Beata B Chagan-Yasutan Haorile H Matsumoto Makoto M Niki Toshiro T Tanaka Michinori M Takahashi Yayoi Y Usami Osumu O Ashino Yugo Y Hattori Toshio T

International journal of molecular sciences 20170628 7

In this study, we investigated the role of a matricellular protein galectin-9 (Gal-9) in pleural effusion related to tuberculosis (TB). Plasma and pleural fluid of a patient with extrapulmonary TB were analyzed for cytokine content by ELISA and Luminex. Peripheral blood mononuclear cells (PBMCs) and pleural fluid cells (PFCs) were examined for interferon-γ (IFN-γ) secretion by the enzyme-linked immunospot (ELISPOT) assay or IFN-γ ELISA, for apoptosis and necrosis by Cell Death Detection ELISA, a ...[more]

PMID: 28657598

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Project description:BackgroundPleural tuberculosis (PlTB) is the most common extrapulmonary manifestation of this infectious disease which still presents high mortality rates worldwide. Conventional diagnostic tests for PlTB register multiple limitations, including the lack of sensitivity of microbiological methods on pleural specimens and the need of invasive procedures such as pleural biopsy performance. In this scenario, the search for biological markers on pleural fluid (PF) has been the target of several studies as a strategy to overcome the limitations of PlTB diagnosis. This study aims to evaluate the use either isolated or in combination with adenosine deaminase (ADA), interferon-gamma (IFN-γ), interferon-gamma inducible protein of 10-kD (IP-10) levels on PF in order to guide an accurate anti-TB treatment in microbiologically non-confirmed cases.Methods and findingsEighty patients presenting pleural effusion under investigation were enrolled in a cross-sectional study conducted at Pedro Ernesto University Hospital, Rio de Janeiro, RJ, Brazil. Peripheral blood (PB) and PF samples collected from all patients were applied to the commercial IFN-γ release assay, QuantiFERON-TB Gold In-Tube, and samples were analyzed for IFN-γ and IP-10 by immunoassays. ADA activity was determined on PF by the colorimetric method. Based on microbiological and histological criteria, patients were categorized as follow: confirmed PlTB (n = 16), non-confirmed PlTB (n = 17) and non-PlTB (n = 47). The Mycobacterium tuberculosis antigen-specific production of IFN-γ and IP-10 on PB or PF did not show significant differences. However, the basal levels of these biomarkers, as well as the ADA activity on PF, were significantly increased in confirmed PlTB in comparison to non-PlTB group. Receiver operating characteristics curves were performed and the best cut-off points of these three biomarkers were estimated. Their either isolated or combined performances (sensitivity [Se], specificity [Sp], positive predictive value [PPV], negative predictive value [NPV] and accuracy [Acc]) were determined and applied to Venn's diagrams among the groups. Based on the confirmed PlTB cases, IFN-γ showed the best performance of them at a cut-off point of 2.33 IU/mL (Se = 93.8% and Sp = 97.9%) followed by ADA at a cut-off of 25.80 IU/L (Se = 100% and Sp = 84.8%) and IP-10 (Cut-point = 4,361.90 pg/mL, Se = 75% and Sp = 82.6%). IFN-γ plus ADA (cut-point: 25.80 IU/L) represent the most accurate biomarker combination (98.4%), showing Se = 93.7%, Sp = 100%, PPV = 100% and NPV = 97.9%. When this analysis was applied in non-confirmed PlTB, 15/17 (88.2%) presented at least two positive biomarkers in combination.ConclusionIFN-γ, IP-10, and ADA in PlTB effusions are significantly higher than in non-PlTB cases. IFN-γ is an excellent rule-in and rule-out test compared to IP-10 and ADA. The combination of IFN-γ and ADA, in a reviewed cut-off point, showed to be particularly useful to clinicians as their positive results combined prompts immediate treatment for TB while both negative results suggest further investigation.

Project description:IntroductionPleural tuberculosis (PlTB), the most common site of extrapulmonary TB, is characterized by a paucibacillary nature and a compartmentalized inflammatory response in the pleural cavity, both of which make diagnosis and management extremely challenging. Although transcriptional signatures for pulmonary TB have already been described, data obtained by using this approach for extrapulmonary tuberculosis and, specifically, for pleural tuberculosis are scarce and heterogeneous. In the present study, a set of candidate genes previously described in pulmonary TB was evaluated to identify and validate a transcriptional signature in clinical samples from a Brazilian cohort of PlTB patients and those with other exudative causes of pleural effusion.MethodsAs a first step, target genes were selected by a random forest algorithm with recursive feature elimination (RFE) from public microarray datasets. Then, peripheral blood (PB) and pleural fluid (PF) samples from recruited patients presenting exudative pleural effusion were collected during the thoracentesis procedure. Transcriptional analysis of the selected top 10 genes was performed by quantitative RT-PCR (RT-qPCR).ResultsReanalysis of the public datasets identified a set of candidate genes (CARD17, BHLHE40, FCGR1A, BATF2, STAT1, BTN3A1, ANKRD22, C1QB, GBP2, and SEPTIN4) that demonstrated a global accuracy of 89.5% in discriminating pulmonary TB cases from other respiratory diseases. Our validation cohort consisted of PlTB (n = 35) patients and non-TB (n = 34) ones. The gene expressions of CARD17, GBP2, and C1QB in PF at diagnosis were significantly different between the two (PlTB and non-TB) groups (p < 0.0001). It was observed that the gene expressions of CARD17 and GBP2 were higher in PlTB PF than in non-TB patients. C1QB showed the opposite behavior, being higher in the non-TB PF. After anti-TB therapy, however, GBP2 gene expression was significantly reduced in PlTB patients (p < 0.001). Finally, the accuracy of the three above-cited highlighted genes in the PF was analyzed, showing AUCs of 91%, 90%, and 85%, respectively. GBP2 was above 80% (sensitivity = 0.89/specificity = 0.81), and CARD17 showed significant specificity (Se = 0.69/Sp = 0.95) in its capacity to discriminate the groups.ConclusionCARD17, GBP2, and C1QB showed promise in discriminating PlTB from other causes of exudative pleural effusion by providing accurate diagnoses, thus accelerating the initiation of anti-TB therapy.

Project description:BackgroundIFN-γ is presently the only soluble immunological marker used to help diagnose latent Mycobacterium tuberculosis (M.tb) infection. However, IFN-γ is not available to distinguish latent from active TB infection. Moreover, extrapulmonary tuberculosis, such as tuberculous pleurisy, cannot be properly diagnosed by IFN-γ release assay. As a result, other disease- or infection-related immunological biomarkers that would be more effective need to be screened and identified.MethodologyA panel of 41 soluble immunological molecules (17 cytokines and 24 chemokines) was tested using Luminex liquid array-based multiplexed immunoassays. Samples, including plasma and pleural effusions, from healthy donors (HD, n = 12) or patients with latent tuberculosis infection (LTBI, n = 20), pulmonary tuberculosis (TB, n = 12), tuberculous pleurisy (TP, n = 15) or lung cancer (LC, n = 15) were collected and screened for soluble markers. Peripheral blood mononuclear cells (PBMCs) and pleural fluid mononuclear cells (PFMCs) were also isolated to investigate antigen-specific immune factors.Principal findingsFor the 41 examined factors, our results indicated that three patterns were closely associated with infection and disease. (1) Significantly elevated plasma levels of IL-2, IP-10, CXCL11 and CXCL12 were present in both patients with tuberculosis and in a sub-group participant with latent tuberculosis infection who showed a higher level of IFN-γ producing cells by ELISPOT assay compared with other latently infected individuals. (2) IL-6 and IL-9 were only significantly increased in plasma from active TB patients, and the two factors were consistently highly secreted after M.tb antigen stimulation. (3) When patients developed tuberculous pleurisy, CCL1, CCL21 and IL-6 were specifically increased in the pleural effusions. In particular, these three factors were consistently highly secreted by pleural fluid mononuclear cells following M.tb-specific antigen stimulation. In conclusion, our data imply that the specific secretion of soluble immunological factors, in addition to IFN-γ, may be used to evaluate M.tb infection and tuberculosis disease.

Dataset Information

Secretion of IFN-? Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis.

Publications

Secretion of IFN-γ Associated with Galectin-9 Production by Pleural Fluid Cells from a Patient with Extrapulmonary Tuberculosis.

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