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Duan Z.-L.,Wenzhou University | Liu H.-F.,Wenzhou University | Huang X.,Institute of Tuberculosis Control | Huang X.,Sun Yat Sen University | And 7 more authors.
Virus Research | Year: 2015

In this study, we set out to identify dengue virus serotype 2 (DENV-2)-specific HLA-A*2402-restricted epitopes and determine the characteristics of T cells generated to these epitopes. We screened the full-length amino-acid sequence of DENV-2 to find potential epitopes using the SYFPEITHI algorithm. Twelve putative HLA-A*2402-binding peptides conserved in hundreds of DENV-2 strains were synthesized, and the HLA restriction of peptides was tested in HLA-A*2402 transgenic mice. Nine peptides (NS4b228-237, NS2a73-81, E298-306, M141-149, NS4a96-105, NS4b159-168, NS5475-484, NS1162-171, and NS5611-620) induced high levels of peptide-specific IFN-γ-secreting cells in HLA-A*2402 transgenic mice. Apart from IFN-γ, NS4b228-237-, NS2a73-81- and E298-306-specific CD8+ cells produced TNF-α and IL-6 simultaneously, whereas M141-149- and NS5475-484- CD8+ cells produced only IL-6. Moreover, splenic mononuclear cells (SMCs) efficiently recognized and killed peptide-pulsed splenocytes. Furthermore, each of nine peptides could be recognized by splenocytes from DENV-2-infected HLA-A*2402 transgenic mice. The SMCs from HLA-A*2402 transgenic mice immunized with nine immunogenic peptides efficiently killed DENV-2-infected splenic monocytes. The present identified epitopes have the potential to be new diagnostic tools for characterization of T-cell immunity in DENV infection and may serve as part of a universal epitope-based vaccine. © 2014 Elsevier B.V. Source


Wang J.,Institute of Tuberculosis Control | Wang J.,Sun Yat Sen University | Wang J.,Guangzhou University | Huang C.,Institute of Tuberculosis Control | And 16 more authors.
Journal of Infection | Year: 2015

Objective: To explore the role of myeloid-related protein 8/14 in mycobacterial infection. Methods: The mRNA and protein expression levels of MRP8 or MRP14 were measured by real-time PCR and flow cytometry, respectively. Role of MRP8/14 was tested by overexpression or RNA interference assays. Flow cytometry and colony forming unit were used to test the phagocytosis and the survival of intracellular Mycobacterium bovis BCG (BCG), respectively. Autophagy mediated by MRP8/14 was detected by Western blot and immunofluorescence. The colocalization of BCG phagosomes with autophagosomes or lysosomes was by detected by confocal microscopy. ROS production was detected by flow cytometry. Results: MRP8/14 expressions were up-regulated in human monocytic THP1 cells and primary macrophages after mycobacterial challenge. Silencing of MRP8/14 suppressed bacterial killing, but had no influence on the phagocytosis of BCG. Importantly, silencing MRP8/14 decreased autophagy and BCG phagosome maturation in THP1-derived macrophages, thereby increasing the BCG survival. Additionally, we demonstrated that MRP8/14 promoted autophagy in a ROS-dependent manner. Conclusions: The present study revealed a novel role of MRP8/14 in the autophagy-mediated elimination of intracellular BCG by promoting ROS generation, which may provide a promising therapeutic target for tuberculosis and other intracellular bacterial infectious diseases. © 2014 The British Infection Association. Source


Zhan X.,Institute of Tuberculosis Control | Zhan X.,Sun Yat Sen University | Fang Y.,Guangzhou Chest Hospital | Hu S.,Institute of Tuberculosis Control | And 12 more authors.
Molecular Immunology | Year: 2015

During chronic inflammation, prolonged over-reactive immune response may lead to tissue destruction, while immune suppression hinders tissue repair and pathogen elimination. Therefore, precise regulation of the immune response is needed to avoid immuno-pathology. Interferon-gamma (IFN-γ) is widely used in clinical treatment of inflammatory diseases. However, the underlying mechanism remains unclear. Here, we evaluated the role of IFN-γ on CD11b+Gr-1+ myeloid cell differentiation and function, using a heat-killed Mycobacterium bovis BCG-induced chronic inflammation model. After challenge with heat-killed BCG, two subpopulations of CD11b+Gr-1+ myeloid cells were generated in the mouse spleen. Phenotypical, morphological and functional analysis indicated that the CD11b+Gr-1high Ly6Ghigh Ly6Clow subset was neutrophil-like myeloid-derived inducer cells (N-MDICs), which promoted T cell activation, while the other subset was CD11b+Gr-1low Ly6Gneg Ly6Chigh monocyte-like myeloid-derived suppressor cells (M-MDSCs) that displayed extensive suppressor function. IFN-γ treatment dampened N-MDICs-mediated T cell activation through up-regulating T cell suppressive mediators, reactive oxygen species (ROS) and arginase I. While for M-MDSCs, IFN-γ reduced their suppressing activity by decreasing the arginase activity. Our study provides evidence that IFN-γ balances the over-reactive vs compromised immune response through different regulation of distinct myeloid subsets, and therefore displays significant therapeutic potential for effective immuno-therapy of chronic inflammatory diseases. © 2015 Elsevier Ltd. Source


Wang J.,Institute of Tuberculosis Control | Wang J.,Sun Yat Sen University | Wu M.,Institute of Tuberculosis Control | Wu M.,Sun Yat Sen University | And 11 more authors.
Molecular Immunology | Year: 2014

mycobacterial infection. Macrophages play a critical role in the host immune response against mycobacterial infection. Our previous study has demonstrated that microRNA-155 (miR-155), one of the most important small non-coding RNAs in the immune system, promotes oxygen-independent mycobacterial killing in macrophages. However, little is known regarding the role of miR-155 in modulating oxygen-dependent mycobactericidal response in macrophages, including the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS). In the present study, we demonstrated that miR-155 was increased in macrophages after Mycobacterium bovis bacille Calmette-Guérin (BCG) infection. Moreover, the BCG-induced upregulation of miR-155 in macrophages was dependent on TLR2, NF-κB and JNK signaling pathways. More importantly, our study explored that miR-155 significantly elevated ROS production in macrophages, although miR-155 had no influence on the inducible nitric oxide synthase (iNOS) expression or nitric oxide (NO) production. In addition, our study demonstrated that miR-155 repressed the expression of src homology 2 (SH2) containing inositol 5-phosphatase1 (SHIP1), and knockdown of SHIP1 greatly increased ROS production in BCG-infected macrophages. Collectively, these data indicate that miR-155 modulates ROS but not RNS production by targeting SHIP1, which may provide a better understanding of the host anti-mycobacterial response. © 2014 Elsevier Ltd. Source

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