Chest Hospital of Guangzhou

Guangzhou, China

Chest Hospital of Guangzhou

Guangzhou, China

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Li L.,Sun Yat Sen University | Qiao D.,Sun Yat Sen University | Li Q.,Sun Yat Sen University | Zhang X.,Chest Hospital of Guangzhou | And 2 more authors.
Tuberculosis | Year: 2012

Tuberculous pleurisy (TBP) is a frequent extrapulmonary manifestation characterized by the accumulation of inflammatory cells that can sometimes be spontaneously self-cured. To achieve a greater insight into T cells at a local site, we systematically characterized and compared the numbers of antigen-specific T cells responding to BCG- or MTB-specific antigens. Our results showed that significantly higher levels of Th1 cytokines were produced by pleural fluid cells (PFCs) than PBMCs following stimulation with BCG or peptides from Mycobacterium tuberculosis (MTB)-specific antigens, ESAT-6 and CFP-10. The proportions of Th1 cells producing IL-2 alone or IL-2 and TNF-α were higher than those producing IFN-γ alone, following stimulation with ESAT-6 or CFP-10 peptides. The cells responding to BCG, ESAT-6 and CFP-10 displayed a CD45RA -CCR7 -CD62L -CD27 - effector/effector memory phenotype. The percentages and median fluorescence intensity (MFI) of polyfunctional CD4 + T cells were significantly higher following stimulation with peptides from ESAT-6 or CFP-10 than BCG. Our results demonstrated that significantly higher levels of polyfunctional CD4 + T cells for the epitopes of ESAT-6 or CFP-10 in PFCs may play an important role in the local control of tuberculosis (TB) infection. © 2011 Elsevier Ltd. All rights reserved.


Fu X.,Sun Yat Sen University | Liu Y.,Sun Yat Sen University | Li L.,Sun Yat Sen University | Li Q.,Sun Yat Sen University | And 5 more authors.
Immunology | Year: 2011

Natural killer (NK) cells are known as innate immune lymphocytes that respond rapidly when challenged by pathogens but little is known about adaptive immune features including memory related to NK cells from human beings. In the present study, we demonstrate for the first time that human NK cells expressing the memory-associated marker CD45RO were persistent in pleural fluid cells (PFCs) from tuberculous patients. CD45RO+ NK cells produced significantly more interferon-γ and were more cytotoxic compared with CD45RO- NK cells from PFCs when stimulated with interleukin-12 (IL-12). Consistently, IL-12 enhanced the expression of granzyme B, CD69, CD25, NKG2D, IL-12 receptors β1 and β2 on CD45RO+ NK cells from PFCs. Our experiments contribute to a better understanding of the NK cells from PFCs and indicate that human CD45RO+ NK cells from PFCs expressing a 'memory-like' phenotype may have an important role in defending against infection by Mycobacterium tuberculosis. © 2011 The Authors. Immunology © 2011 Blackwell Publishing Ltd.


Wu C.,Sun Yat Sen University | Ma J.,Sun Yat Sen University | Xu Y.,Sun Yat Sen University | Zhang X.,Chest Hospital of Guangzhou | And 2 more authors.
Tuberculosis | Year: 2014

Summary We recently reported that pleural fluid mononuclear cells (PFMCs) from tuberculous pleurisy stimulated with Bacillus Calmette-Guérin (BCG) or TB antigens produced high levels of cytokines. However, it was still unclear what mechanism of the PFMCs used to migrate into the pleural fluids in TB infection. In the present study, we found that CD3+CD4+ and CD3+CD8+ T cells from PFMCs expressed significantly high levels of CXCR3 compared to PBMCs. In addition, the levels of CXCL10 (the ligand for CXCR3) in pleural fluids were significantly higher than those in normal serum and cancerous fluids. After stimulation with BCG, PFMCs produced high levels of CXCL10. Importantly, the synthesis of CXCL10 was mainly dependent on the BCG-induced production of IFNs, because the neutralization of endogenous IFN-α or IFN-γ with mAbs significantly reduced the production of CXCL10 from BCG-stimulated PFMCs. In addition, the tubercular pleural fluid (TBPF) or exogenous CXCL10 induced the migration of PFMCs, indicating that IFN-α or IFN-γ modulated the immune response through the expression of CXCL10 to aid the recruitment and selective homing of activated/effector cells to the site of Mycobacterium tuberculosis (M.tb) infection. Taken together, the levels of CXCL10 in pleural fluids were high and BCG-stimulated PFMCs expressed high levels of CXCL10, and CXCL10 induced the migration of PFMCs into the pleural fluids in TB infection. © 2013 Elsevier Ltd. All rights reserved.


Li L.,Sun Yat Sen University | Qiao D.,Sun Yat Sen University | Fu X.,Sun Yat Sen University | Lao S.,Chest Hospital of Guangzhou | And 2 more authors.
PLoS ONE | Year: 2011

Important advances have been made in the immunodiagnosis of tuberculosis (TB) based on the detection of Mycobacterium tuberculosis (MTB)-specific T cells. However, the sensitivity and specificity of the immunological approach are relatively low because there are no specific markers for antigen-specific Th cells, and some of the Th cells that do not produce cytokines can be overlooked using this approach. In this study, we found that MTB-specific peptides of ESAT-6/CFP-10 can stimulate the expression of CD40L specifically in CD4+ T cells but not other cells from pleural fluid cells (PFCs) in patients with tuberculous pleurisy (TBP). CD4+CD40L+ but not CD4+CD40L- T cells express IFN-γ, IL-2, TNF-α, IL-17 or IL-22 after stimulation with MTB-specific peptides. In addition, CD4+CD40L+ T cells were found to be mostly polyfunctional T cells that simultaneously produce IFN-γ, IL-2 and TNF-α and display an effector or effector memory phenotype (CD45RA-CD45RO+CCR7-CD62L-ICOS-). To determine the specificity of CD4+CD40L+ T cells, we incubated PFCs with ESTA-6/CFP-10 peptides and sorted live CD4+CD40L+ and CD4+CD40L- T cells by flow cytometry. We further demonstrated that sorted CD4+CD40L+, but not CD4+CD40L- fractions, principally produced IFN-γ, IL-2, TNF-α, IL-17 and IL-22 following restimulation with ESTA-6/CFP-10 peptides. Taken together, our data indicate that the expression of CD40L on MTB-specific CD4+ T cells could be a good marker for the evaluation and isolation of MTB-specific Th cells and might also be useful in the diagnosis of TB. © 2011 Li et al.


Wu C.,Sun Yat Sen University | Li Z.,Sun Yat Sen University | Fu X.,Sun Yat Sen University | Yu S.,Sun Yat Sen University | And 2 more authors.
Oncotarget | Year: 2015

Natural killer T (NKT) cells from mouse and human play an important role in the immune responses against Mycobacterium tuberculosis. However, the function of CD3+TCRvβ11+ NKT cells at the local site of M. tuberculosis infection remains poorly defined. In the present study, we found that after stimulation with M. tuberculosis antigens, NKT cells isolated from tuberculosis (TB) pleural fluid mononuclear cells (PFMCs) produced IL-21 and other cytokines including IFN-γ, TNF-α, IL-2 and IL- 17. IL-21-expressing NKT cells in PFMCs displayed effector memory phenotype, expressing CD45ROhighCD62LlowCCR7low. Moreover, NKT cells expressed high levels of CXCR5 and all of IL-21-expressing NKT cells co-expressed CXCR5. The frequency of BCL-6-expression was higher in IL-21-expressing but not in non-IL-21-expressing CD3+TCRvβ11+ NKT cells. Sorted CD3+TCRvβ11+ NKT cells from PFMCs produced IFN-γ and IL-21 after stimulation, which expressed CD40L. Importantly, CD3+TCRvβ11+ NKT cells provided help to B cells for the production of IgG and IgA. Taken together, our data demonstrate that CD3+TCRvβ11+ NKT cells from a local site of M. tuberculosis infection produce IL-21, express CXCR5 and CD40L, help B cells to secrete IgG and IgA, and may participate in local immune responses against M. tuberculosis infection.


Li L.,Sun Yat Sen University | Qiao D.,Sun Yat Sen University | Zhang X.,Chest Hospital of Guangzhou | Liu Z.,Chest Hospital of Guangzhou | Wu C.,Sun Yat Sen University
Immunobiology | Year: 2011

Most of the studies evaluating the function of tuberculosis (TB)-specific T cells were only based on the ability to produce cytokines, which may not fully reflect the function of T cells. In the present study, we confirmed that Bacille Calmette Guerin (BCG) could significantly induce cytokine production by CD4+ T cells from BCG-vaccinated PPD+ donors. In addition, CD4+ T cells were activated, divided and proliferated in response to BCG stimulation. Phenotypic analysis showed that IFN-γ+CD4+ T cells displayed CD45RA-CCR7+/-CD62L-, indicating that these CD4+ T cells were central and effector memory cells. The analysis of cytokine profiles demonstrated that most of BCG-specific BrdU+CD4+ T cells produced Th1 cytokines in response to polyclonal stimulation. In addition, we found that regulatory T cells (Treg) suppressed BCG-induced proliferation and IFN-γ production by memory CD4+ T cells. The suppressive effects of Treg on BCG-specific responses of CD4+ T cells could be partially reversed by blocking the production of IL-10. Taken together, our results demonstrated that functional central and effector memory BCG-specific CD4+ T cells could be detected based on the activation, proliferation and division of these cells, and modulated by Treg in PBMCs from BCG-vaccinated PPD+ donors. © 2010 Elsevier GmbH.


Fu X.,Sun Yat Sen University | Yang B.,Sun Yat Sen University | Lao S.,Chest Hospital of Guangzhou | Fan Y.,Sun Yat Sen University | Wu C.,Sun Yat Sen University
Clinical Immunology | Year: 2013

We have previously shown that human memory-like NK cells were persistent in tuberculous pleurisy but it was unclear how NK cells migrated into the pleural fluids. At present, we found that NK cells from TB pleural fluid cells (PFCs) expressed significantly higher levels of CXCR3 and CXCR4 than NK cells from PBMCs. Migration assay demonstrated that IP-10 and SDF-1 induced more migration of NK cells from PFCs than PBMCs. CD45RO+ or CD45RO- NK cells from PFCs were co-cultured with autologous monocytes and stimulated with BCG. The results showed CD45RO+ but not CD45RO- NK cells produced significantly higher levels of IFN-γ, which was IL-12-dependent since anti-IL-12Rβ1 mAbs could significantly inhibit the IFN-γ by NK cells. Collectively, our data demonstrated that human Mycobacterium tuberculosis-specific NK cells were migrated into the local site of TB infection mainly via IP-10/CXCR3 and SDF-1/CXCR4 axis, memory-like NK cells might display an important role against M. tuberculosis infection. © 2013 Elsevier Inc.


Li L.,Sun Yat Sen University | Yang B.,Sun Yat Sen University | Zhang X.,Chest Hospital of Guangzhou | Lao S.,Chest Hospital of Guangzhou | Wu C.,Sun Yat Sen University
Tuberculosis | Year: 2014

Increasing evidences in animals and humans suggest that CD8+ T cells contribute significantly to immune defenses against Mycobacterium tuberculosis (Mtb). In the present study, we found that without any stimulation, CD8+ T cells in pleural fluid cells (PFCs) expressed significantly higher levels of CD69 than PBMCs from patients with tuberculous pleurisy (TBP). CD8+CD69+ T cells expressed significantly higher levels of CD45RO and HLA-DR and lower levels of CD45RA than CD8+CD69 - T cells, demonstrating that CD8+CD69+ T cells were activated memory cells. Furthermore, we found higher expression of CCR6 and lower expression of CCR7 and CD62L on CD8+CD69+ T cells compared with CD8+CD69- T cells, suggesting that the expression of CCR6 and reduced expression of CCR7 and CD62L might facilitate the migration of circulating CD8+CD69+ T cells into tuberculous pleural space. Importantly, following stimulation with culture filtrate protein of 10 kDa (CFP10) peptides, CD8+CD69+ T cells but not CD8+CD69- T cells expressed CD107a/b, IFN-γ and TNF-α, demonstrating that CD8+CD69+ T cells were MTB-specific cells. In addition, the majority of CD8 +CD69+ T cells were dominated by polyfunctional T cells. In summary, we demonstrated that CD69 as a useful marker for MTB-specific CD8+ T cells in PFCs from patients with TBP enabled a direct ex vivo estimation of the quantity, as well as the quality, of MTB-specific CD8 + responses. © 2014 Elsevier Ltd. All rights reserved.


Li L.,Sun Yat Sen University | Qiao D.,Sun Yat Sen University | Fu X.,Sun Yat Sen University | Lao S.,Chest Hospital of Guangzhou | And 2 more authors.
PLoS ONE | Year: 2011

Th1 cell-mediated immune responses at the site of active infection are important to restrict the growth of M.tuberculosis (MTB) and for the spontaneous resolution of patients with tuberculous pleurisy (TBP). In the present study, we found that without any stimulation, CD4 + T cells in pleural fluid cells (PFCs) from patients with TBP expressed significantly higher levels of CD69 than PBMCs from patients with tuberculosis (TB) or healthy donors. CD4 +CD69 + T cells expressed T-bet and IL-12Rβ2. After stimulation with MTB-specific antigens, CD4 +CD69 + T cells expressed significantly higher levels of IFN-γ, IL-2 and TNF-α than CD4 +CD69 - T cells, demonstrating that CD4 +CD69 + T cells were MTB-specific Th1 cells. In addition, CD4 +CD69 + T cells were mostly polyfunctional Th1 cells that simultaneously produced IFN-γ, IL-2, TNF-α and displayed an effector or effector memory phenotype (CD45RA -CCR7 -CD62L -CD27 -). Moreover, the percentages of CD4 +CD69 + T cells were significantly and positively correlated with polyfunctional T cells. Interestingly, sorted CD4 +CD69 + but not CD4 +CD69 - fractions by flow cytometry produced IFN-γ, IL-2 and TNF-α that were significantly regulated by CD4 +CD25 + Treg cells. Taken together, based on the expression of CD69, we found a direct quantitative and qualitative method to detect and evaluate the in vivo generated MTB-specific polyfunctional CD4 + T cells in PFCs from patients with TBP. This method can be used for the potential diagnosis and enrichment or isolation of MTB-specific Th1 cells in the investigations. © 2011 Li et al.


Ma J.,Sun Yat Sen University | Yang B.,Sun Yat Sen University | Yu S.,Sun Yat Sen University | Zhang Y.,Sun Yat Sen University | And 5 more authors.
FASEB Journal | Year: 2014

The mechanism by which IFN-α regulates the host response to Mycobacterium tuberculosis (M.tb) infection in humans is poorly understood. In the present study, we found that freshly isolated pleural fluid mononuclear cells (PFMCs) from tuberculous pleural effusion but not peripheral blood mononuclear cells (PBMCs) spontaneously expressed IFN-α and IL-1β in vivo. In addition, exogenous IFN-α significantly inhibited production of IL-1β in PFMCs after stimulation with Bacillus Calmette-Guérin (BCG). To further evaluate the effect of endogenous IFN-α on BCG-induced IL-1β production, a neutralizing antibody to IFN-α was added to the cultures of BCG-stimulated PFMCs. As expected, neutralization of IFN-α by antibody significantly enhanced the production of IL-1β. Notably, we showed that IFN-α inhibited production of IL-1β through 2 distinct mechanisms: IFN-α signaling, via the STAT1 transcription factor, suppressed caspase-1-dependent IL-1β maturation, and IFN-α induced the production of IL-10 in a STAT1-dependent manner in which IL-10 reduced the abundance of IL-1β. In contrast, we found that IFN-α enhanced the production of IFN-7, and IFN-7 also suppressed IL-1β production in the PFMCs during BCG stimulation. Our findings demonstrate that IFN-α employs distinct pathways for regulating IL-1β production and reveal that in the case of M.tb infection, the induction of IFN-α and IFN-7 might be associated with M.tb immune escape and disease progression in infected humans. © FASEB.

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