In vitro stimulation with HBV therapeutic vaccine candidate Nasvac activates B and T cells from chronic hepatitis B patients and healthy donors

Hepatitis B virus (HBV) chronic infections remain a considerable health problem worldwide. The standard therapies have demonstrated limited efficacy, side effects or need life-long treatments. Nowadays therapeutic vaccination is a promising option. Recently, we developed a new vaccine formulation called Nasvac, based on the combination of surface and core antigens from HBV. Clinical trials already performed showed good efficacy in virus control. However, the exact mode of action of Nasvac formulation remains unclear. So far the functional impairment of DCs during persistent HBV infection is a controversial issue. On the other hand, it is known that B cells may function as antigen presenting cells (APC) activating T cells. The hepatitis B core antigen contained in Nasvac vaccine is able to bind and activate a high frequency of naive human B cells. In the present study the surface expression of activation and exhaustion markers on B cells and the subsequent activation of T cells after in vitro stimulation with Nasvac antigens were evaluated in chronic HBV patients and healthy donors. B- and T-cell phenotype and proliferation were assessed by flow cytometry. Our results indicate that in contrast to exhaustions markers B cell activation markers were increased on both study groups after Nasvac stimulation. A shift toward an activation phenotype was observed for both B and T cells. The present work suggests that B cells could act as efficient APCs for Nasvac antigens in humans, which might suggest the use of activated B cells as immunotherapeutic strategy for chronic hepatitis B.     

Serum amyloid A chemoattracts immature dendritic cells and indirectly provokes monocyte chemotaxis by induction of cooperating CC and CXC chemokines

Serum amyloid A (SAA) is an acute phase protein that is upregulated in inflammatory diseases and chemoattracts monocytes, lymphocytes, and granulocytes via its G protein‐coupled receptor formyl peptide receptor like 1/formyl peptide receptor 2 (FPRL1/FPR2). Here, we demonstrated that the SAA1α isoform also chemoattracts monocyte‐derived immature dendritic cells (DCs) in the Boyden and μ‐slide chemotaxis assay and that its chemotactic activity for monocytes and DCs was indirectly mediated via rapid chemokine induction. Indeed, SAA1 induced significant amounts (≥5 ng/mL) of macrophage inflammatory protein‐1α/CC chemokine ligand 3 (MIP‐1α/CCL3) and interleukin‐8/CXC chemokine ligand 8 (IL‐8/CXCL8) in monocytes and DCs in a dose‐dependent manner within 3 h. However, SAA1 also directly activated monocytes and DCs for signaling and chemotaxis without chemokine interference. SAA1‐induced monocyte migration was nevertheless significantly prevented (60–80% inhibition) in the constant presence of desensitizing exogenous MIP‐1α/CCL3, neutralizing anti‐MIP‐1α/CCL3 antibody, or a combination of CC chemokine receptor 1 (CCR1) and CCR5 antagonists, indicating that this endogenously produced CC chemokine was indirectly contributing to SAA1‐mediated chemotaxis. Further, anti‐IL‐8/CXCL8 antibody neutralized SAA1‐induced monocyte migration, suggesting that endogenous IL‐8/CXCL8 acted in concert with MIP‐1α/CCL3. This explained why SAA1 failed to synergize with exogenously added MIP‐1α/CCL3 or stromal cell‐derived factor‐1α (SDF‐1α)/CXCL12 in monocyte and DC chemotaxis. In addition to direct leukocyte activation, SAA1 induces a chemotactic cascade mediated by expression of cooperating chemokines to prolong leukocyte recruitment to the inflammatory site.     

Pleurotus ferulae water extract enhances the maturation and function of murine bone marrow-derived dendritic cells through TLR4 signaling pathway

Dendritic cells (DCs) play important roles in the regulation of immune system, which link innate and adaptive immune responses. Mature DCs produced interleukin (IL)-12 promote optimal type 1 T helper (Th1) cells and cytotoxic T lymphocytes. The extracts of traditional herbal medicines have been shown to enhance immune responses through promoting the maturation and cytokine production of DCs. Here, we investigated the effects of Pleurotus ferulae water extract (PFWE) on the maturation and function of bone marrow–derived DCs (BM–DCs). Upon PFWE treatment, BM–DCs dose-dependently upregulated the expression of CD40, CD80, CD86 and MHC II and increased the production of IL-12, IL-6 and tumor necrosis factor (TNF)-α but not for IL-10, which is mediated by TLR4 signaling pathway, at least partially. The production of prostaglandin E2 (PGE2) in BM–DCs was decreased by the treatment of PFWE. Moreover, PFWE treatment decreased the expression of active caspase-3 but increased the expression of CCR7. PFWE treated DCs enhanced the proliferation of allogenic CD8⁺ T cells and the capacity of antigen presenting to autologous CD8⁺ T cells. The combination of PFWE and CpG–ODN further enhanced the maturation and function of murine BM–DCs. The results showed that PFWE could enhance the maturation and function of DCs through TLR4 signaling pathway and has additive effect when combined with CpG–ODN, suggesting that PFWE alone or combined with CpG–ODN could be used to enhance the immune responses.     

免疫因素在特发性炎症性肌病中的作用机制

特发性炎症性肌炎包括多发性肌炎,皮肌炎及包涵体肌炎,是一组以肌肉组织的炎细胞浸润,肌无力为主要特点,浸润的炎细胞是以T淋巴细胞和巨噬细胞为主,个别也有B淋巴细胞。本文主要综述了T、B淋巴细胞、DC及其它APCs在IIMs中的病理生理作用。     

Leukemia derived dendritic cell (DCleu) mediated immune response goes along with reduced (leukemia-specific) regulatory T-cells

The blastmodulatory Kit-M, composed of granulocyte-macrophage colony-stimulating-factor (GM-CSF) and Prostaglandin E1 (PGE₁), is known to convert myeloid leukaemic blasts (from AML patients) into leukaemia derived dendritic cells (DC_leu), which activate immunoreactive cells to gain antileukemic/leukaemia-specific activity. In this study we had a special focus on the influence of Kit-M treated, DC/DCleu containing patients’whole blood (WB, n = 16) on the provision of immunosuppressive regulatory T-cells.We could confirm that Kit-M significantly increased frequencies of (mature) dendritic cells (DC) and DC_leu from leukemic whole blood (WB) without induction of blast proliferation. After mixed lymphocyte culture (MLC) with patients’ T-cells we confirmed that DC_leu mediated leukemia-specific responses- going along with activated and leukemia-specific T- and NK-cells in an intracellular cytokine staining assay (ICS) and a degranulation assay (Deg)- resulted in an increased anti-leukemic cytotoxicity (Cytotoxicity Fluorolysis Assay = CTX). We could demonstrate that (leukemia-specific) CD4⁺ and CD8⁺ regulatory T-cell population (T_reg) decreased significantly after MLC compared to controls. We found significant positive correlations of leukemia-specific CD3⁺CD4⁺ cells with frequencies of (mature) DC_leu. Achieved anti-leukemic cytotoxicity correlated significantly positive with leukemia-specific CD3⁺CD8⁺ cells and significantly negatively with (leukemia-specific) T_reg.In summary we demonstrate that immunesuppressive (leukemia-specific) regulatory T-cells are significantly downregulated after Kit-M triggered MLC- going along with a (reinstalled) antileukemic reactivity of the immune system (as demonstrated with functional assays ICS, Deg, CTX).     

Immunological and Regulatory Functions of Uninfected and Virus Infected Immature and Mature Subtypes of Dendritic Cells – a Review

In 1868, dendritic cells (DCs) were discovered in human skin by Paul Langerhans using gold staining. These cells were named Langerhans cells (LCs) after their discoverer who, due to their dendrites, regarded them as neurons. One hundred and eleven years were to pass until it was discovered that in vertebrates these cells originate in the bone marrow as monocytes. In the 1980s, DC research was mostly carried out on DCs that are present in different tissues of mice and humans. These studies revealed that after interaction with foreign antigens, skin LCs/DCs migrate through the lymph vessels to the draining lymph nodes and induce the two arms of the immune response. The isolation of DCs from tissue cell suspensions opened the way to studies on the cells' surface proteins and their ability to stimulate immune responses. During the 1990s, studies revealed the role of DCs in the activation of naïve T cells in the lymph nodes and the regulatory properties of DCs in lymph nodes, thymus, gut, and spleen.Part A of the review deals with the DC system of human and mice and immunological and regulatory functions of subsets of DCs in the skin with reference to migrating and stationary DCs, as well as the connection between DCs and the nervous system. Furthermore, the origin of both follicular DCs that are present in lymphoid tissues and thymic DCs are discussed. Part B is devoted to virus infections of DCs with an emphasis on infections caused by human herpes viruses. Part C presents the modulation of DC gene expression in response to the influenza virus. Contemporary research focuses on the role of DCs in the immune systems of vertebrates. Moreover, studies are being conducted on the regulatory functions of DCs by tissue cells in different organs of vertebrates.     

In situ analysis of lung antigen-presenting cells during murine pulmonary infection with virulent Mycobacterium tuberculosis

Scarce information exists about the role of lung antigen-presenting cells (APCs) in vivo during pulmonary tuberculosis. As APCs activate cellular immunity, following intratracheal inoculation with virulent Mycobacterium tuberculosis, we assessed in situ lung APC recruitment, distribution, granuloma involvement, morphology and mycobacterial burden by using MHC-CII, CD14, scavenger receptor class A (SRA), the murine dendritic cell (DC)-restricted marker CD11c and Ziehl-Neelsen staining. CD11c(+) DC and CD14(+) cell recruitment into lungs appeared by day 14, continuing until day 60. MHC-CII(+) cells increased since day 7, persisting until day 60. Thus, virulent mycobacteria delays (14-21 days) lung APC recruitment compared to model antigens and nonvirulent bacilli (24-48 h). Regarding granuloma constitution, highly bacillary CD14(+) and SRA(+) cells were centrally located. MHC-CII(+) cells were more peripheral, with less mycobacteria. CD11c(+) cells were heterogeneously distributed within granulomas, with scarce bacilli. When labelling lung suspensions for MHC-CII and classifying cells as macrophages or DC, then staining for Ziehl-Neelsen, a remarkable segregation was found regarding bacillary burden. Most macrophage-like cells contained numerous bacilli, while DC had no or scarce mycobacteria. This implies differential APC contributions in situ during pulmonary tuberculosis regarding mycobacterial uptake, granuloma involvement and perhaps bacillary growth.