Antibodies that block virus attachment to vero cells are a major component of the human neutralizing antibody response against dengue virus type 2

Epidemiological data strongly implicate a role for the host humoral immune response in both protection against and exacerbation of dengue virus-caused disease. In an effort to characterize elements of the normal human immune response against dengue virus we have addressed the issue of antibody-mediated neutralization of dengue virus. We show here the ability of both mouse monoclonal antibody 3H5 and human anti-dengue neutralizing sera to block binding of dengue-2 virus to monkey kidney (Vero) cells. Since Vero cells possess virus receptors but not Fc receptors we conclude that the major effect of host neutralizing antibodies is to block virus attachment to Vero cell dengue virus receptors. Analysis of 61 patient antisera yielded good correlation (Pearson's coefficient = 0.90; P < 0.001) between neutralizing activity and ability to block virus-cell attachment suggesting that antibody-mediated neutralization of dengue virus occurs primarily extracellularly and less by a postat-tachment mechanism as has been described for certain other viruses. © 1995 Wiley-Liss, inc.     

Properdin and factor H production by human dendritic cells modulates their T‐cell stimulatory capacity and is regulated by IFN‐γ

Dendritic cells (DCs) and complement are both key members of the innate and adaptive immune response. Recent experimental mouse models have shown that production of alternative pathway (AP) components by DCs strongly affects their ability to activate and regulate T‐cell responses. In this study we investigated the production and regulation of properdin (fP) and factor H (fH) both integral regulators of the AP, by DCs and tolerogenic DCs (tolDCs). Both fP and fH were produced by DCs, with significantly higher levels of both AP components produced by tolDCs. Upon activation with IFN‐γ both cells increased fH production, while simultaneously decreasing production of fP. IL‐27, a member of the IL‐12 family, increased fH, but production of fP remained unaffected. The functional capacity of fP and fH produced by DCs and tolDCs was confirmed by their ability to bind C3b. Inhibition of fH production by DCs resulted in a greater ability to induce allogenic CD4⁺ T‐cell proliferation. In contrast, inhibition of fP production led to a significantly reduced allostimulatory capacity. In summary, this study shows that production of fP and fH by DCs, differentially regulates their immunogenicity, and that the local cytokine environment can profoundly affect the production of fP and fH.     

Glycogen synthetase kinase 3 inhibition drives MIC-A/B to promote cytokine production by human natural killer cells in Dengue virus type 2 infection

Dengue virus (DENV) is the most widespread arbovirus worldwide and is responsible for major outbreaks. The host's immune response plays a crucial role in controlling this infection but might also contribute to the promotion of viral spread and immunopathology. In response to DENV infection, NK cells preferentially produce cytokines and are cytotoxic in the presence of specific antibodies. Here, we identified that DENV-2 inhibits glycogen synthase kinase 3 (GSK-3) activity to subsequently induce MHC class-1-related chain (MIC) A and MIC-B expression and IL-12 production in monocyte-derived DCs, independently of the STAT-3 pathway. The inhibition of GSK-3 by DENV-2 or small molecules induced MIC-A/B expression on monocyte-derived DCs, resulting in autologous NK cells of a specific increase in IFN-γ and TNF-α production, in the absence of direct cytotoxicity. Together, these findings identified GSK-3 as a regulator of MIC-A/B expression and suggested its role in DENV-2 infection to specifically induce cytokine production by NK cells.     

Contribution of Fcγ receptors to human respiratory syncytial virus pathogenesis and the impairment of T‐cell activation by dendritic cells

Human respiratory syncytial virus (hRSV) is the leading cause of infant hospitalization related to respiratory disease. Infection with hRSV produces abundant infiltration of immune cells into the airways, which combined with an exacerbated pro‐inflammatory immune response can lead to significant damage to the lungs. Human RSV re‐infection is extremely frequent, suggesting that this virus may have evolved molecular mechanisms that interfere with host adaptive immunity. Infection with hRSV can be reduced by administering a humanized neutralizing antibody against the virus fusion protein in high‐risk infants. Although neutralizing antibodies against hRSV effectively block the infection of airway epithelial cells, here we show that both, bone marrow‐derived dendritic cells (DCs) and lung DCs undergo infection with IgG‐coated virus (hRSV‐IC), albeit abortive. Yet, this is enough to negatively modulate DC function. We observed that such a process is mediated by Fcγ receptors (FcγRs) expressed on the surface of DCs. Remarkably, we also observed that in the absence of hRSV‐specific antibodies FcγRIII knockout mice displayed significantly less cellular infiltration in the lungs after hRSV infection, compared with wild‐type mice, suggesting a potentially harmful, IgG‐independent role for this receptor in hRSV disease. Our findings support the notion that FcγRs can contribute significantly to the modulation of DC function by hRSV and hRSV‐IC. Further, we provide evidence for an involvement of FcγRIII in the development of hRSV pathogenesis.     

Plasmacytoid dendritic cells and type 1 interferon promote peripheral expansion of forkhead box protein 3+ regulatory T cells specific for the ubiquitous RNA‐binding nuclear antigen La/Sjögren's syndrome (SS)‐B

RNA‐binding nuclear antigens are a major class of self‐antigen to which immune tolerance is lost in rheumatic diseases. Serological tolerance to one such antigen, La/Sjögren's syndrome (SS)‐B (La), is controlled by CD4⁺ T cells. This study investigated peripheral tolerance to human La (hLa) by tracking the fate of hLa‐specific CD4⁺ T cells expressing the transgenic (Tg) 3B5.8 T cell receptor (TCR) after adoptive transfer into lymphocyte‐replete recipient mice expressing hLa as a neo‐self‐antigen. After initial antigen‐specific cell division, hLa‐specific donor CD4⁺ T cells expressed forkhead box protein 3 (FoxP3). Donor cells retrieved from hLa Tg recipients displayed impaired proliferation and secreted interleukin (IL)?10 in vitro in response to antigenic stimulation. Transfer of highly purified FoxP3‐negative donor cells demonstrated that accumulation of hLa‐specific regulatory T cells (T_reg) was due primarily to expansion of small numbers of donor T_reg. Depletion of recipient plasmacytoid dendritic cells (pDC), but not B cells, severely hampered the accumulation of FoxP3⁺ donor T_reg in hLa Tg recipients. Recipient pDC expressed tolerogenic markers and higher levels of co‐stimulatory and co‐inhibitory molecules than B cells. Adoptive transfer of hLa peptide‐loaded pDC into mice lacking expression of hLa recapitulated the accumulation of hLa‐specific T_reg. Blockade of the type 1 interferon (IFN) receptor in hLa Tg recipients of hLa‐specific T cells impaired FoxP3⁺ donor T cell accumulation. Therefore, peripheral expansion of T_reg specific for an RNA‐binding nuclear antigen is mediated by antigen‐presenting pDC in a type 1 IFN‐dependent manner. These results reveal a regulatory function of pDC in controlling autoreactivity to RNA‐binding nuclear antigens.     

BDCA3 expression is associated with high IFN‐λ production by CD34+‐derived dendritic cells generated in the presence of GM‐CSF,IL‐4, and/or TGF‐β

High BDCA3 expression is associated with a specific human IFN‐λ‐producing dendritic cell (DC) subset. However, BDCA3 has also been detected on other DC subsets. Thus far, development and function of BDCA3 expression on DCs remains poorly understood. Human Langerhans cells (LCs) and interstitial DCs (intDCs) can be generated in vitro by differentiation of CD34⁺ hematopoietic progenitors via distinct precursor DCs (preDCs), CD1a⁺ preDCs, and CD14⁺ preDCs, respectively. Here, we identified BDCA3 expression in this well‐known GM‐CSF/TNF‐α‐driven culture system and described the effect of IL‐4 and/or TGF‐β on induction of BDCA3 expression. In control or TGF‐β cultures, BDCA3 was only detected on CD14⁺ preDC‐derived intDCs. IL‐4 induced BDCA3 expression in both CD14⁺‐derived and CD1a⁺‐derived cultures. TGF‐β and IL‐4 together further increased CD14⁺‐derived and CD1a⁺‐derived BDCA3⁺ DC frequencies, which partly expressed CLEC9A, but were not identical to the BDCA3^highCLEC9A⁺ DC subset in vivo. Importantly, BDCA3⁺ cells, but not BDCA3^? cells, in this system produced high IFN‐λ levels upon polyinosinic:polycytidylic acid (polyI:C) stimulation. This culture system, in which BDCA3 expression is preferentially associated with the intDC lineage and IFN‐λ‐producing capacity, will greatly contribute to further research on the function and regulation of BDCA3 expression and IFN‐λ production by DCs.     

Toll‐like receptor 3 stimulation promotes Ro52/TRIM21 synthesis and nuclear redistribution in salivary gland epithelial cells,partially via type I interferon pathway

Up‐regulated expression of Ro52/tripartite motif‐containing protein 21 (TRIM21), Ro60/TROVE domain family, member 2 (TROVE2) and lupus LA protein/Sjögren's syndrome antigen B (La/SSB) autoantigens has been described in the salivary gland epithelial cells (SGEC) of patients with Sjögren's syndrome (SS). SGECs, the key regulators of autoimmune SS responses, express high levels of surface functional Toll‐like receptor (TLR)‐3, whereas Ro52/TRIM21 negatively regulates TLR‐3‐mediated inflammation. Herein, we investigated the effect of TLR‐3‐signalling on the expression of Ro52/TRIM21, as well as Ro60/TROVE2 and La/SSB autoantigens, by SGECs. The effect of TLR‐3 or TLR‐4 stimulation on autoantigen expression was evaluated by polyI:C or lipopolysaccharide (LPS) treatment, respectively, of SGEC lines (10 from SS patients, 12 from non‐SS controls) or HeLa cells, followed by analysis of mRNA and protein expression. PolyI:C, but not LPS, resulted in a two‐step induction of Ro52/TRIM21 mRNA expression by SGECs, a 12‐fold increment at 6 h followed by a 2·5‐fold increment at 24–48 h, whereas it induced a late two‐fold up‐regulation of Ro60/TROVE2 and La/SSB mRNAs at 48 h. Although protein expression levels were not affected significantly, the late up‐regulation of Ro52/TRIM21 mRNA was accompanied by protein redistribution, from nucleolar‐like pattern to multiple coarse dots spanning throughout the nucleus. These late phenomena were mediated significantly by interferon (IFN)‐β production, as attested by cognate secretion and specific inhibition experiments and associated with IFN regulatory factor (IRF)3 degradation. TLR‐3‐signalling had similar effects on SGECs obtained from SS patients and controls, whereas it did not affect the expression of these autoantigens in HeLa cells. TLR‐3 signalling regulates the expression of autoantigens by SGECs, implicating innate immunity pathways in their over‐expression in inflamed tissues and possibly in their exposure to the immune system.