Citations of BMS216MST
Plasmacytoid Dendritic Cells Are Proportionally Expanded at Diagnosis of Type 1 Diabetes and Enhance Islet Autoantigen Presentation to T-Cells Through Immune Complex Capture
Allen,Jennifer S.; Pang,Karl; Skowera,Ania; Ellis,Richard; Rackham,Chloe; Lozanoska-Ochser,Biliana; Tree,Timothy; Leslie,R.David; Tremble,Jennifer M.; Dayan,Colin M.; Peakman,Mark
Diabetes 2009;58:138-145.
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Abstract
OBJECTIVE--Immune-mediated destruction of {beta}-cells resulting in type 1 diabetes involves activation of proinflammatory, islet autoreactive T-cells, a process under the control of dendritic cells of the innate immune system. We tested the hypothesis that type 1 diabetes development is associated with disturbance of blood dendritic cell subsets that could enhance islet-specific autoimmunity. RESEARCH DESIGN AND METHODS--We examined blood dendritic cells (plasmacytoid and myeloid) in 40 patients with recent-onset diabetes (median duration 28 days) and matched control subjects. We also examined the relative ability of different dendritic cell subsets to process and present soluble or immune complexed islet cell autoantigen (the islet tyrosine phosphatase IA-2) to responder CD4 T-cells. RESULTS--The balance of blood dendritic cells was profoundly disturbed at diabetes diagnosis, with a significantly elevated proportion of plasmacytoid and reduction of myeloid cells compared with control subjects. Dendritic cell subset distribution was normal in long-standing disease and in patients with type 2 diabetes. Both dendritic cell subsets processed and presented soluble IA-2 to CD4 T-cells after short-term culture, but only plasmacytoid dendritic cells enhanced (by as much as 100%) autoantigen presentation in the presence of IA-2+ autoantibody patient serum. CONCLUSIONS--The plasmacytoid subset of dendritic cells is overrepresented in the blood close to diabetes onset and shows a distinctive ability to capture islet autoantigenic immune complexes and enhance autoantigen-driven CD4 T-cell activation. This suggests a synergistic proinflammatory role for plasmacytoid dendritic cells and islet cell autoantibodies in type 1 diabetes
2356
Anti-HIV State but Not Apoptosis Depends on IFN Signature in CD4+ T Cells
Audige,Annette; Urosevic,Mirjana; Schlaepfer,Erika; Walker,Russell; Powell,Doug; Hallenberger,Sabine; Joller,Helen; Simon,Hans Uwe; Dummer,Reinhard; Speck,Roberto F.
The Journal of Immunology 2006;177:6227-6237.
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Abstract
To gain insights into the molecular mechanisms underlying early host responses to HIV in the CD4+ T cell target population, we examined gene expression in CD4+ T cells isolated 24 h after ex vivo HIV infection of lymphocyte aggregate cultures derived from human tonsils. Gene profiling showed a distinct up-regulation of genes related to immune response and response to virus, notably of IFN-stimulated genes (ISGs), irrespective of the coreceptor tropism of the virus. This mostly IFN-{alpha}-dependent gene signature suggested the involvement of plasmacytoid dendritic cells, a principal component of the antiviral immune response. Indeed, depletion of plasmacytoid dendritic cells before HIV inoculation abrogated transcriptional up-regulation of several ISGs and resulted in increased levels of HIV replication. Treatment with a blocking anti-IFN-{alpha}R Ab yielded increased HIV replication; conversely, HIV replication was decreased in pDC-depleted cultures treated with IFN-{alpha}. Among up-regulated ISGs was also TRAIL, indicating a potential role of the IFN signature in apoptosis. However, a blocking anti-TRAIL Ab did not abrogate apoptosis of CD4+ T cells in CXCR4-tropic HIV-infected cultures, suggesting the involvement of pathways other than TRAIL mediated. We conclude that acute HIV infection of lymphoid tissue results in up-regulation of ISGs in CD4+ T cells, which induces an anti-HIV state but not apoptosis
690
TLR9 expression is related to immune activation but is impaired in individuals with chronic immune activation
Ayash-Rashkovsky,Mila; Bentwich,Zvi; Borkow,Gadi
The International Journal of Biochemistry & Cell Biology 2005;37:› Link
Abstract
Millions of individuals in developing countries are infected with helminths and other chronic infectious diseases, such as HIV-1, which lead to persistent immune activation and unbalanced immune state. We have suggested that the capacity of chronically immune activated individuals to protect themselves, cope with infections, and mount protective immunity following vaccination, is highly impaired. Here we examined the expression of toll-like receptor 9 (TLR9), as an essential component in the recognition of immunostimulating bacterial CpG-DNA motifs, in different subsets of human peripheral blood mononuclear cells (PBMC) obtained from chronically immune activated and non-activated individuals. TLR9 expression was correlated to immune cell activation and was upregulated following phytohemagglutinin or anti-CD3 activation. PBMC obtained from chronically immune activated individuals had a different overall pattern of TLR9 expression, including reduced upregulation of this receptor following additional immune activation, and diminished responsiveness to CpG-DNA stimulation, in comparison to non-activated individuals. These differences may partly account for the reduced capacity of chronically immune activated individuals to mount effective immune responses and strengthen the notion that the host immune background should be considered in the design and trial of potential adjuvants and vaccines.
1641
Early and strong immune responses are associated with control of viral replication and recovery in lassa virus-infected cynomolgus monkeys
Baize,S.; Marianneau,P.; Loth,P.; Reynard,S.; Journeaux,A.; Chevallier,M.; Tordo,N.; Deubel,V.; Contamin,H.
J Virol. 2009;83:5890-5903.
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Abstract
Lassa virus causes a hemorrhagic fever endemic in West Africa. The pathogenesis and the immune responses associated with the disease are poorly understood, and no vaccine is available. We followed virological, pathological, and immunological markers associated with fatal and nonfatal Lassa virus infection of cynomolgus monkeys. The clinical picture was characterized by fever, weight loss, depression, and acute respiratory syndrome. Transient thrombocytopenia and lymphopenia, lymphadenopathy, splenomegaly, infiltration of mononuclear cells, and alterations of the liver, lungs, and endothelia were observed. Survivors exhibited fewer lesions and a lower viral load than nonsurvivors. Although all animals developed strong humoral responses, antibodies appeared more rapidly in survivors and were directed against GP(1), GP(2), and NP. Type I interferons were detected early after infection in survivors but only during the terminal stages in fatalities. The mRNAs for CXCL10 (IP-10) and CXCL11 (I-TAC) were abundant in peripheral blood mononuclear cells and lymph nodes from infected animals, but plasma interleukin-6 was detected only in fatalities. In survivors, high activated-monocyte counts were followed by a rise in the total number of circulating monocytes. Activated T lymphocytes circulated in survivors, whereas T-cell activation was low and delayed in fatalities. In vitro stimulation with inactivated Lassa virus induced activation of T lymphocytes from all infected monkeys, but only lymphocytes from survivors proliferated. Thus, early and strong immune responses and control of viral replication were associated with recovery, whereas fatal infection was characterized by major alterations of the blood formula and, in organs, weak immune responses and uncontrolled viral replication
2558
Pegylated interferon-alpha2a kinetics during experimental haemodialysis: impact of permeability and pore size of dialysers
Barril,G.; Quiroga,J.A.; Sanz,P.; Rodriguez-Salvanes,F.; Selgas,R.; Carreno,V.
Alimentary Pharmacology and Therapeutics 2004;20:37-44.
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Abstract
Summary Background : Therapeutics in end-stage renal disease (ESRD) patients undergoing haemodialysis (HD) has to consider potential drug clearance during the dialysis procedure. Pegylated interferon-alpha (PEG-IFN-alpha), a middle-size protein drug active against viral hepatitis, allows convenient once-weekly dosing due to prolonged plasma half-life. Aim : To investigate the impact of permeability and dialyser pore size on PEG-IFN-alpha blood levels during experimental HD. Methods : Polymethylmetacrylate (PMMA) membrane 1.6 m2 dialysers with three different permeabilities/pore sizes were selected. Results : A 40 kDa PEG-IFN-alpha2a (PEGASYS) was not cleared (< 5%) through low-flux/small pore size (25 A;B3A) and high-flux/middle-large pore size (60 A;BKP) dialysers, and was partially (=15%) through intermediate permeability/large pore size (100 A;BKF) dialysers. In contrast, unmodified 17 kDa IFN-alpha2a(Roferon-A) was removed (65%-95%) through BKP or BKF, but not B3A, PMMA dialysers. Moreover, 12 kDaPEG-IFN-alpha2b(PegIntron) was cleared (40%-80%) through PMMA dialysers with pore sizes >= 60 A. When B3A or BKP were replaced every hour PEG-IFN-alpha2a plasma levels remained constant throughout three experimental-HD-sessions, but PEG-IFN-alpha2b was cleared partially every BKP replacement. Porosity differ among high-flux dialysers. Neither PEG-IFN-alpha2a nor PEG-IFN-alpha2b were removed after three HD sessions through (27/31/33 A) pore size polysulphone dialysers. Although PEG-IFN-alpha2a was not cleared through middle pore-size (43 A/AN69ST) polyacrylonitrile dialyser, PEG-IFN-alpha2b was partially removed. Conclusions : The pharmacokinetics of Peg-IFN-alpha may vary in a patient on dialysis
663
Plasmacytoid Dendritic Cells Control TLR7 Sensitivity of Naive B Cells via Type I IFN
Bekeredjian-Ding,Isabelle Beatrice; Wagner,Moritz; Hornung,Veit; Giese,Thomas; Schnurr,Max; Endres,Stefan; Hartmann,Gunther
The Journal of Immunology 2005;174:4043-4050.
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Abstract
Detailed information of human B cell activation via TLR may lead to a better understanding of B cell involvement in autoimmunity and malignancy. In this study we identified a fundamental difference in the regulation of TLR7- and TLR9-mediated B cell stimulation: whereas the induction of polyclonal naive B cell proliferation by the TLR7 ligands resiquimod (R848) and loxoribine required the presence of plasmacytoid dendritic cells (PDCs), activation via the TLR9 ligand CpG was independent of PDCs. We found that PDC-derived type I IFN enhanced TLR7 sensitivity of B cells by selectively up-regulating TLR7 expression. In contrast the expression levels of TLR9 and of other TLRs studied remained unchanged. In the presence of type I IFN, TLR7 ligation triggered polyclonal B cell expansion and B cell differentiation toward Ig-producing plasma cells; notably, this occurred independently of T cell help and B cell Ag. Human B cells did not respond to ligands of other TLRs including TLR2, TLR4 and TLR6 with and without type I IFN. In conclusion, our results reveal a distinct regulation of TLR7 and TLR9 function in human B cells and highlight TLR7 and TLR9 as unique targets for therapeutic intervention in B cell-mediated immunity and disease
411
T Cell-Independent, TLR-Induced IL-12p70 Production in Primary Human Monocytes
Bekeredjian-Ding,Isabelle; Roth,Susanne Ilona; Gilles,Stefanie; Giese,Thomas; Ablasser,Andrea; Hornung,Veit; Endres,Stefan; Hartmann,Gunther
The Journal of Immunology 2006;176:7438-7446.
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Abstract
IL-12p70 is a key cytokine for the induction of Th1 immune responses. IL-12p70 production in myeloid cells is thought to be strictly controlled by T cell help. In this work we demonstrate that primary human monocytes can produce IL-12p70 in the absence of T cell help. We show that human monocytes express TLR4 and TLR8 but lack TLR3 and TLR7 even after preincubation with type I IFN. Simultaneous stimulation of TLR4 and TLR8 induced IL-12p70 in primary human monocytes. IL-12p70 production in peripheral blood myeloid dendritic cells required combined stimulation of TLR7/8 ligands together with TLR4 or with TLR3 ligands. In the presence of T cell-derived IL-4, but not IFN-{gamma}, stimulation with TLR7/8 ligands was sufficient to stimulate IL-12p70 production. In monocytes, type I IFN was required but not sufficient to costimulate IL-12p70 induction by TLR8 ligation. Furthermore, TLR8 ligation inhibited LPS-induced IL-10 in monocytes, and LPS alone gained the ability to stimulate IL-12p70 in monocytes when the IL-10 receptor was blocked. Together, these results demonstrate that monocytes are licensed to synthesize IL-12p70 through type I IFN provided via the Toll/IL-1R domain-containing adaptor inducing IFN-[beta] pathway and the inhibition of IL-10, both provided by combined stimulation with TLR4 and TLR8 ligands, triggering a potent Th1 response before T cell help is established
76
Are interleukin-16 and thrombopoietin new tools for the in vitro generation of dendritic cells?
Bella,Silvia Della; Nicola,Stefania; Timofeeva,Inna; Villa,Maria Luisa; Santoro,Armando; Berardi,Anna C.
Blood 2004;104:4020-4028.
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Abstract
The effects of interleukin 16 (IL-16) on dendritic cell (DC) generation from human CD34+ progenitor cells are not known. Here, we show that IL-16 added to a basal cocktail comprised of granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-4, Flt-3 ligand (Flt3L), and tumor necrosis factor {alpha} (TNF-{alpha}) does induce the CD34+ hematopoietic cells to proliferate in vitro and to differentiate into phenotypically and functionally mature DCs. IL-16 exerts this function more efficiently than stem cell factor (SCF) as a control, thrombopoietin (TPO), or IL-16 plus TPO. Moreover, we show that the combination of IL-16 plus TPO induces the generation of tolerogenic DCs, able to induce an anergic state in T cells that persists when T cells are rechallenged with immunogenic DCs. An altered pattern of cytokine production, a reduced expression of the C-type lectin DC-SIGN, and an increased surface expression of the inhibitory molecules immunoglobulin-like transcript 2 (ILT-2), ILT-3, and ILT-4 may all contribute to confer the tolerogenic properties of these DCs. Generation of tolerogenic DCs may aid the exploration of new therapeutic strategies to promote tolerance to autoantigens and prevent disease development. (Blood. 2004;104:4020-4028)
349
Activation of Human Plasmacytoid Dendritic Cells by TLR9 Impairs Fc{gamma}RII-Mediated Uptake of Immune Complexes and Presentation by MHC Class II
Benitez-Ribas,Daniel; Tacken,Paul; Punt,Cornelis J.A.; de Vries,I.Jolanda; Figdor,Carl G.
The Journal of Immunology 2008;181:5219-5224.
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Abstract
Human plasmacytoid dendritic cells (pDCs)2 exploit Ag uptake receptors like CD32a for internalization of exogenous Ags. Activation of pDC by TLR9 ligand CpG-C induces strong maturation. Surprisingly, we observed that CpG-C-stimulated pDCs showed impaired Ag-specific T cell proliferation whereas the induction of allogeneic T cell proliferation was not affected. We demonstrated that signals from TLR9 caused a rapid down-regulation of the capacity of pDC to take-up Ab-Ag complexes without altering their CD32a expression, thus explaining the reduced Ag presentation. The recent contrasting biological responses that were observed upon TLR9 ligation in pDCs prompted us to study the effect of several TLR9 ligands. We observed that type I IFN-inducer CpG-A, localizing in the early endosomal compartment, did not affect CD32a function, whereas CpGs localizing in the late endosomes and inducing pDC maturation clearly inhibited CD32a-mediated Ag uptake and presentation. We conclude that TLR9 ligands not only determine the type of response, i.e., type I IFN production (innate immunity) or maturation (adaptive immunity), but also directly affect Ag presentation capacity of pDCs. We hypothesize that pDC, once activated via TLR9-ligands reaching the late endosomes, can only present initially sampled Ags and thus are protected from uptake and processing of additional potential self-Ags
2295
Prednisolone Suppresses the Function and Promotes Apoptosis of Plasmacytoid Dendritic Cells
Boor,P.P.C.; Metselaar,H.J.; Mancham,S.; Tilanus,H.W.; Kusters,J.G.; Kwekkeboom,J.
American Journal of Transplantation 2006;6:2332-2341.
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Abstract
Organ transplant recipients are highly susceptible to viral infections early after transplantation. Plasmacytoid dendritic cells (PDC) play a major role in antiviral immunity. Therefore, we determined the numbers of circulating PDC after liver transplantation (LTX) and established the effects of immunosuppressive drugs on PDC survival and function. PDC were determined longitudinally in 13 LTX recipients treated with prednisone and cyclosporin or tacrolimus. Purified PDC were cultured with or without clinically relevant concentrations of cyclosporin, tacrolimus or prednisolone. Apoptosis induction was monitored by determination of active caspase-3, nuclear condensation and annexin-V/7AAD staining. After LTX, a 4-fold reduction in the number of circulating PDC was observed (p < 0.01), which recovered partially after discontinuation of prednisone treatment. In vitro, prednisolone induced apoptosis in PDC, while cyclosporin and tacrolimus did not. Higher doses of prednisolone were needed to induce apoptosis in Toll-like receptor (TLR)-stimulated PDC. However, non-apoptosis inducing concentrations of prednisolone suppressed interferon-alpha production, upregulation of co-stimulatory molecules and allo-stimulatory capacity of TLR-stimulated PDC. In conclusion, prednisolone induces apoptosis in PDC, which explains the decline in circulating PDC numbers after transplantation. Moreover, prednisolone suppresses the functions of TLR-stimulated PDC. Therefore, corticosteroid-free immunosuppressive therapy may reduce the number and severity of viral infections after transplantation
1342
Attenuated Expression of A20 Markedly Increases the Efficacy of Double-Stranded RNA-Activated Dendritic Cells As an Anti-Cancer Vaccine
Breckpot,Karine; erts-Toegaert,Cindy; Heirman,Carlo; Peeters,Uschi; Beyaert,Rudi; Aerts,Joeri L.; Thielemans,Kris
The Journal of Immunology 2009;182:860-870.
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Abstract
A20 is a zinc finger protein with ubiquitin-modifying activity. A20 has been described as negatively regulating signaling induced by the TNF receptor and TLR family in a number of cell types, including mouse bone marrow-derived dendritic cells (DCs). However, the expression and effect of A20 in activated human monocyte-derived DCs have not been previously evaluated. We report that DCs activated with the TLR3 ligand poly(I:C) up-regulate A20. Down-regulating A20 demonstrated its role in the functional activation of DCs. A20 down-regulated DCs showed higher activation of the transcription factors NF-{kappa}B and activator protein-1, which resulted in increased and sustained production of IL-6, IL-10, and IL-12p70. We additionally silenced the immunosuppressive cytokine IL-10 and demonstrated that IL-10 inhibits T cell proliferation. We further demonstrated that A20 down-regulated DCs skew naive CD4+ T cells toward IFN-{gamma} producing Th1 cells, a process which is dependent on IL-12p70 and which is unaffected by IL-10. Furthermore, A20 and/or IL-10 down-regulated DCs had an enhanced capacity to prime Melan-A/MART-1 specific CD8+ T cells. Finally, we demonstrated that potent T cell stimulatory DCs are generated by the simultaneous delivery of poly(I:C12U), A20, or A20/IL-10 small interfering RNA and Ag-encoding mRNA, introducing a one step approach to improve DC-based vaccines. Together these findings demonstrate that A20 negatively regulates NF-{kappa}B and activator protein-1 in DCs and that down-regulation of A20 results in DCs with enhanced T cell stimulatory capacity
2524
Plasmacytoid dendritic cell-specific receptor ILT7-Fc{varepsilon}RI{gamma} inhibits Toll-like receptor-induced interferon production
Cao,Wei; Rosen,David B.; Ito,Tomoki; Bover,Laura; Bao,Musheng; Watanabe,Gokuran; Yao,Zhengbin; Zhang,Li; Lanier,Lewis L.; Liu,Yong Jun
Journal of Experimental Medicine 2006;203:1399-1405.
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Abstract
Immunoglobulin-like transcripts are a family of inhibitory and stimulatory cell surface immune receptors. Transcripts for one member of this family, ILT7, are selectively expressed in human plasmacytoid dendritic cells (pDCs). We demonstrate here that ILT7 protein associates with the signal adapter protein Fc{varepsilon}RI{gamma} to form a receptor complex. Using an anti-ILT7 monoclonal antibody, we show that ILT7 is expressed specifically on human pDCs, but not on myeloid dendritic cells or other peripheral blood leukocytes. Cross-linking of ILT7 resulted in phosphorylation of Src family kinases and Syk kinase and induced a calcium influx in freshly isolated pDCs, which was blocked by Src family and Syk kinases inhibitors, thus indicating the activation of an immunoreceptor-based tyrosine activation motif-mediated signaling pathway. ILT7 cross-linking on CpG or influenza virus-stimulated primary pDCs inhibited the transcription and secretion of type I interferon and other cytokines. Therefore, the ILT7-Fc{varepsilon}RI{gamma} receptor complex negatively regulates the innate immune functions of human pDCs
698
BDCA2/Fc[RI¦ Complex Signals through a Novel BCR-Like Pathway in Human Plasmacytoid Dendritic Cells
Cao,Wei; Zhang,Li; Rosen,David B.; Bover,Laura; Watanabe,Gokuran; Bao,Musheng; Lanier,Lewis L.; Liu,Yong Jun
PLoS Biology 2007;5:e248› Link
Abstract
Dendritic cells are equipped with lectin receptors to sense the extracellular environment and modulate cellular responses. Human plasmacytoid dendritic cells (pDCs) uniquely express blood dendritic cell antigen 2 (BDCA2) protein, a C-type lectin lacking an identifiable signaling motif. We demonstrate here that BDCA2 forms a complex with the transmembrane adapter Fc[RI¦. Through pathway analysis, we identified a comprehensive signaling machinery in human pDCs, similar to that which operates downstream of the B cell receptor (BCR), which is distinct from the system involved in T cell receptor (TCR) signaling. BDCA2 crosslinking resulted in the activation of the BCR-like cascade, which potently suppressed the ability of pDCs to produce type I interferon and other cytokines in response to Toll-like receptor ligands. Therefore, by associating with Fc[RI¦, BDCA2 activates a novel BCR-like signaling pathway to regulate the immune functions of pDCs
2621
Altered T helper 1 reaction but not increase of virus load in patients with dengue hemorrhagic fever
Chen,R.F.; Liu,J.W.; Yeh,W.T.; Wang,L.; Chang,J.C.; Yu,H.R.; Cheng,J.T.; Yang,K.D.
FEMS Immunology and Medical Microbiology 2005;44:43-50.
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Abstract
To investigate whether dengue-2 patients with and without dengue hemorrhagic fever had different virus load, immune mediators, or T helper (Th) reaction, we simultaneously measured virus load, immune mediators and the Th1/Th2 transcription factors T-bet/GATA-3 mRNA expression in a large outbreak of dengue-2 infections in Southern Taiwan. Results showed that virus load was not significantly different between patients with and without dengue hemorrhagic fever. Patients with dengue fever had higher IFN-? levels, but patients with dengue hemorrhagic fever had significantly higher IL-10 levels. Further studies showed that patients with dengue hemorrhagic fever had a significantly lower T-bet than those with dengue fever, but GATA-3 mRNA expression in peripheral blood leukocytes was not significant difference between both groups. In conclusion, altered Th1 reaction as reflected by lower T-bet mRNA expression associated with higher IL-10 levels might be involved in the pathogenesis of dengue hemorrhagic fever.
918
Altered T helper 1 reaction but not increase of virus load in patients with dengue hemorrhagic fever
Chen,Rong Fu; Liu,Jien Wei; Yeh,Wen Ting; Wang,Lin; Chang,Jen Chieh; Yu,Hong Ren; Cheng,Jiin Tsuey; Yang,Kuender D.
FEMS Immunology and Medical Microbiology 2005;44:43-50.
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Abstract
Abstract To investigate whether dengue-2 patients with and without dengue hemorrhagic fever had different virus load, immune mediators, or T helper (Th) reaction, we simultaneously measured virus load, immune mediators and the Th1/Th2 transcription factors T-bet/GATA-3 mRNA expression in a large outbreak of dengue-2 infections in Southern Taiwan. Results showed that virus load was not significantly different between patients with and without dengue hemorrhagic fever. Patients with dengue fever had higher IFN-gamma levels, but patients with dengue hemorrhagic fever had significantly higher IL-10 levels. Further studies showed that patients with dengue hemorrhagic fever had a significantly lower T-bet than those with dengue fever, but GATA-3 mRNA expression in peripheral blood leukocytes was not significant difference between both groups. In conclusion, altered Th1 reaction as reflected by lower T-bet mRNA expression associated with higher IL-10 levels might be involved in the pathogenesis of dengue hemorrhagic fever
652
SAGE library screening reveals ILT7 as a specific plasmacytoid dendritic cell marker that regulates type I IFN production
Cho,Minkwon; Ishida,Koji; Chen,Jingtao; Ohkawa,Jun; Chen,Wei; Namiki,Sahori; Kotaki,Ayumi; Arai,Naoko; Arai,Ken ichi; Kamogawa-Schifter,Yumiko
International Immunology 2008;20:155-164.
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Abstract
Plasmacytoid dendritic cells (pDCs) link innate to acquired immune responses by producing high levels of type I IFN upon infection. In order to identify the specific genes that control pDC, we compared serial analysis of gene expression libraries from human pDCs, herpes simplex virus-stimulated pDCs and monocytes. We found that Ig-like transcript ILT7 is specifically expressed on pDC cell surfaces and is down-regulated when pDC mature in response to viral or bacterial stimulation. ILT7 expression on the cell surface required association with the Fc{epsilon}RI{gamma} adaptor molecule. Although treatment with one anti-ILT7-specific mAb suppressed type I IFN production in response to cytosine-phosphate-guanosice (CpG) stimulation, another anti-ILT7 mAb up-regulated type I IFN production. We conclude that ILT7 is a key regulator of human pDC function
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TLR9 Activation Is Defective in Common Variable Immune Deficiency
Cunningham-Rundles,Charlotte; Radigan,Lin; Knight,Adina K.; Zhang,Li; Bauer,Laura; Nakazawa,Atsushi
The Journal of Immunology 2006;176:1978-1987.
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Abstract
Common variable immune deficiency (CVID) is a primary immune deficiency characterized by low levels of serum immune globulins, lack of Ab, and reduced numbers of CD27+ memory B cells. Although T, B, and dendritic cell defects have been described, for the great majority, genetic causes have not been identified. In these experiments, we investigated B cell and plasmacytoid dendritic cell activation induced via TLR9, an intracellular recognition receptor that detects DNA-containing CpG motifs from viruses and bacteria. CpG-DNA activates normal B cells by the constitutively expressed TLR9, resulting in cytokine secretion, IgG class switch, immune globulin production, and potentially, the preservation of long-lived memory B cells. We found that CpG-DNA did not up-regulate expression of CD86 on CVID B cells, even when costimulated by the BCR, or induce production of IL-6 or IL-10 as it does for normal B cells. TLR9, found intracytoplasmically and on the surface of oligodeoxynucleotide-activated normal B cells, was deficient in CVID B cells, as was TLR9 mRNA. TLR9 B cell defects were not related to proportions of CD27+ memory B cells. CpG-activated CVID plasmacytoid dendritic cells did not produce IFN-{alpha} in normal amounts, even though these cells contained abundant intracytoplasmic TLR9. No mutations or polymorphisms of TLR9 were found. These data show that there are broad TLR9 activation defects in CVID which would prevent CpG-DNA-initiated innate immune responses; these defects may lead to impaired responses of plasmacytoid dendritic cells and loss of B cell function
484
Functional repertoire of dendritic cells generated in granulocyte macrophage-colony stimulating factor and interferon-{alpha}
Della Bella,Silvia; Nicola,Stefania; Riva,Antonio; Biasin,Mara; Clerici,Mario; Villa,Maria Luisa
Journal of Leukocyte Biology 2003;jlb› Link
Abstract
Monocyte-derived dendritic cells (DCs) generated in granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-4 (IL-4-DCs) are used to enhance antitumor immunity in cancer patients, although recent evidence suggests that their functional repertoire may be incomplete; in particular, IL-4-DCs appear unable to induce type 2 cytokine-producing T helper (Th) cells. To assess whether type 1 interferon (IFN) could replace IL-4 and generate DCs with a more complete repertoire, we characterized in detail DCs generated from human monocytes cultured with GM-CSF and IFN-{alpha} (IFN-DCs). We found that IFN-{alpha} induces DC differentiation more efficiently than IL-4, yielding similar numbers of DCs in a shorter time and that this differentiation persists upon removal of cytokines. Although IFN-DCs had a more mature immunophenotype than IL-4-DCs, showing higher expression of CD80, CD86, and CD83, they still preserved comparable endocytic and phagocytic capacities and responsiveness to maturation stimuli. IFN-DCs had strong antigen-presenting capacity, inducing intense proliferation of T cells to alloantigens or influenza virus. Moreover, IFN-DCs produced lower levels of IL-12p70 and higher levels of IFN-{alpha}, IL-4, and IL-10 than IL-4-DCs. As a consequence of this different pattern of cytokine secretion, IFN-DCs induced T cells to produce type 1 (IFN-{gamma}) and type 2 (IL-4 and IL-10) cytokines, and as expected, IL-4-DCs induced only Th1 differentiation. As immune responses with extreme Th1 bias are considered inadequate for the induction of optimal, systemic antitumor immunity, the ability of IFN-DCs to promote more balanced cytokine responses may suggest the advisability to consider these cells in the development of future, DC-based immunotherapy trials
1490
Modifications in Small Interfering RNA That Separate Immunostimulation from RNA Interference
Eberle,Florian; Giessler,Kerstin; Deck,Christopher; Heeg,Klaus; Peter,Mirjam; Richert,Clemens; Dalpke,Alexander H.
The Journal of Immunology 2008;180:3229-3237.
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Abstract
Synthetic small interfering RNA (siRNA) can suppress the expression of endogenous mRNA through RNA interference. It has been reported that siRNA can induce type I IFN production from plasmacytoid dendritic cells, leading to off-target effects. To separate immunostimulation from the desired gene-specific inhibitory activity, we designed RNA strands with chemical modifications at strategic positions of the ribose or nucleobase residues. Substitution of uridine residues by 2'-deoxyuridine or thymidine residues was found to decrease type I IFN production upon in vitro stimulation of human PBMC. Thymidine residues in both strands of a siRNA duplex further decreased immunostimulation. Fortunately, the thymidine residues did not affect gene-silencing activity. In contrast, 2'-O-methyl groups at adenosine and uridine residues reduced both IFN-{alpha} secretion and gene-silencing activity. Oligoribonucleotides with 2'-O-methyladenosine residues actively inhibited IFN-{alpha} secretion induced by other immunostimulatory RNAs, an effect not observed for strands with 2'-deoxynucleosides. Furthermore, neither 5-methylcytidine nor 7-deazaguanosine residues in the stimulatory strands affected IFN-{alpha} secretion, suggesting that recognition does not involve sites in the major groove of duplex regions. The activity data, together with structure prediction and exploratory UV-melting analyses, suggest that immunostimulatory sequences adopt folded structures. The results show that immunostimulation can be suppressed by suitable chemical modifications without losing siRNA potency by introducing seemingly minor structural changes
2544
Plasmacytoid dendritic cells activate allergen-specific TH2 memory cells: Modulation by CpG oligodeoxynucleotides
Farkas,L.; Kvale,E.O.; Johansen,F.E.; Jahnsen,F.L.; Lund-Johansen,F.
Journal of Allergy and Clinical Immunology 2004;114:436-443.
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Abstract
Background Plasmacytoid dendritic cells (PDCs) accumulate in the nasal mucosa of allergic rhinitis patients, but their function in upper airway allergy has not been determined. CpG oligodeoxynucleotides, potent adjuvants in immunotherapeutic strategies in animal models, are especially effective at activating PDCs. These cells are therefore potential targets for immunomodulation in humans. Objective In this study, PDCs were compared with CD11c+ dendritic cells (DCs), a very potent antigen-presenting cell type, for their capacity to induce allergen-dependent activation of TH2 memory cells. Then, we investigated whether CpG-activated PDCs were able to modulate the allergen-specific TH2 memory response. Methods DCs were isolated from patients with upper airway allergy and cocultured with autologous CD4+ T cells with or without grass pollen extract and CpG. In some experiments cells were restimulated with allergen-pulsed monocyte-derived DCs. T-cell activation was measured by their proliferative response and cytokine production. Results PDCs stimulated allergen-dependent T-cell proliferation and TH2 cytokine production as efficiently as CD11c+ DCs. CpG-activated PDCs inhibited allergen-dependent proliferation of TH2 memory cells and markedly increased IFN-ã production in PDC/T-cell cocultures; the former effect depended on the CpG-induced IFN-a/ß production by the PDCs. Conclusion Our results demonstrated that PDCs efficiently drive allergen-dependent TH2 memory responses, suggesting that they play an active role in the allergic reaction. However, in the presence of CpG, PDCs were responsible for increased production of the TH1-related cytokines IFN-a and IFN-ã, indicating that mucosal PDCs may be targets for CpG-based immunotherapeutic strategies against airway allergy.
945
Plasmacytoid dendritic cells induce a distinct cytokine pattern in virus-specific CD4+ memory T cells that is modulated by CpG oligodeoxynucleotides
Farkas,L.; Kvale,E.O.; Lund-Johansen,F.; Jahnsen,F.L.
Scand.J.Immunol. 2006;64:404-411.
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Abstract
Inherent properties of dendritic cell (DC) subsets are important in the regulation of naive T-cell differentiation (e.g. Th1 versus Th2 cells), whereas effector memory T cells are believed to produce a fixed cytokine repertoire independent of the type of antigen presenting cell. Here we show that two distinct human DC subsets, plasmacytoid DC (PDC) and myeloid CD11c+ DC, induced autologous mumps virus-specific memory CD4(+) T cells to produce markedly different cytokine patterns upon antigen stimulation. PDC stimulated the T cells to produce gamma-interferon (IFN-gamma) and interleukin-(IL)-10, whereas CD11c+ DC induced lower levels of IFN-gamma, virtually no IL-10, but significant levels of IL-5. Analysis of intracellular cytokine production showed simultaneous production of IL-10 and IFN-gamma in mumps-specific T cells activated by PDC, a cytokine pattern similar to that described for Th1-like regulatory cells. Introduction of CpG oligodeoxynucleotides in PDC/T-cell co-cultures had synergistic effect on virus-dependent IFN-gamma production, whereas the other cytokines remained unchanged. Together, our results show that the type of DC involved in reactivation of previously primed T cells may have significant impact on the resulting cytokine response and suggest that targeting of viral antigens and adjuvant to specific DC subsets should be considered in the design of therapeutic antiviral vaccines
1456
Functional defects of dendritic cells in patients with CD40 deficiency
Fontana,Stefania; Moratto,Daniele; Mangal,Surinder; De Francesco,Maria; Vermi,William; Ferrari,Simona; Facchetti,Fabio; Kutukculer,Necil; Fiorini,Claudia; Duse,Marzia; Das,Pranab K.; Notarangelo,Luigi D.; Plebani,Alessandro; Badolato,Raffaele
Blood 2003;102:4099-4106.
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Abstract
We have recently identified 2 patients with a rare autosomal recessive form of hyper IgM disease, known as HIGM3, caused by mutations in the CD40 gene. These patients had opportunistic infections observed on X-linked hyper IgM syndrome (HIGM), suggesting that the CD40-CD40 ligand interaction is important for promoting T-cell-mediated immunity. To evaluate whether innate immunity signals may substitute CD154 for inducing the maturation of dendritic cells (DCs), we analyzed monocyte-derived DCs in these patients. Monocyte-derived DCs of HIGM3 subjects on ex vivo stimulation with tumor necrosis factor-{alpha} (TNF-{alpha}) or lipopolysaccharide (LPS) combined with interferon-{gamma} (IFN-{gamma}) normally express all the markers of mature DCs, such as CD83 and DC-LAMP. However, cell surface levels of HLA-DR in mature DCs are reduced, as is costimulatory activity of these cells for allogeneic naive T cells. In addition, CD40-deficient DCs secrete lower amounts of interleukin-12 (IL-12) but larger quantities of IL-10 than control subjects. Finally, analysis of circulating plasmacytoid DCs demonstrates a normal percentage of this subset in CD40-deficient cells, but IFN-{alpha} secretion in response to herpes simplex virus 1 (HSV-1) infection is severely reduced in patients. These observations suggest that the severe impairment of DC maturation may contribute to the defect of T-cell-mediated immunity observed in HIGM3 patients. (Blood. 2003;102:
1496
Anti-BDCA-4 (neuropilin-1) antibody can suppress virus-induced IFN-alpha production of plasmacytoid dendritic cells
Grage-Griebenow,Evelin; Loseke,Stefan; Kauth,Marion; Gehlhar,Kirsten; Zawatzky,Rainer; Bufe,Albrecht
Immunol Cell Biol 2007;85:383-390.
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Abstract
Plasmacytoid dendritic cells (PDC) in human blood are the main source of virus-induced interferon (IFN)-alpha. They exhibit a lineage-negative phenotype but all express BDCA-4, which is homologous to the neuronal receptor neuropilin-1. Specific staining with anti-BDCA-4 antibody is used for positive isolation of PDC from blood by magnetic cells sorting. Here, it is demonstrated that these positively selected PDC showed reduced or completely abolished IFN-alpha release compared to unstained PDC, which were negatively selected by magnetic depletion of lineage-positive blood mononuclear cells. In addition, treatment of these unstained PDC with anti-BDCA-4 mAb also resulted in at least two-fold lower or reduced virus-induced IFN-alpha production. It is shown that the antibody not only affects cell survival or block virus attachment but also reduces IFN-alpha release induced by non-viral CpG oligodeoxynucleotides. In conclusion, data suggest an immunoregulatory role for BDCA-4 on PDC as demonstrated for IFN-alpha response to virus.
1196
Pegylated interferon-[alpha] protects type 1 pneumocytes against SARS coronavirus infection in macaques
Haagmans,Bart; Kuiken,Thijs; Martina,Byron; Fouchier,Ron; Rimmelzwaan,Guus; van Amerongen,Geert; van Riel,Debby; de Jong,Ton; Itamura,Shigeyuki; Chan,Kwok Hung; Tashiro,Masato; Osterhaus,Albert
Nat Med 2004;10:290-293.
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Abstract
The primary cause of severe acute respiratory syndrome (SARS) is a newly discovered coronavirus. Replication of this SARS coronavirus (SCV) occurs mainly in the lower respiratory tract, and causes diffuse alveolar damage. Lack of understanding of the pathogenesis of SARS has prevented the rational development of a therapy against this disease. Here we show extensive SCV antigen expression in type 1 pneumocytes of experimentally infected cynomolgus macaques (Macaca fascicularis) at 4 d postinfection (d.p.i.), indicating that this cell type is the primary target for SCV infection early in the disease, and explaining the subsequent pulmonary damage. We also show that prophylactic treatment of SCV-infected macaques with the antiviral agent pegylated interferon- (IFN-) significantly reduces viral replication and excretion, viral antigen expression by type 1 pneumocytes and pulmonary damage, compared with untreated macaques. Postexposure treatment with pegylated IFN- yielded intermediate results. We therefore suggest that pegylated IFN- protects type 1 pneumocytes from SCV infection, and should be considered a candidate drug for SARS therapy
1198
Identification and Functional Analysis of Tumor-Infiltrating Plasmacytoid Dendritic Cells in Head and Neck Cancer
Hartmann,Evelyn; Wollenberg,Barbara; Rothenfusser,Simon; Wagner,Moritz; Wellisch,Daniela; Mack,Brigitte; Giese,Thomas; Gires,Olivier; Endres,Stefan; Hartmann,Gunther
Cancer Research 2003;63:6478-6487.
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Abstract
The antitumor activity of IFN-{alpha} is well established. However, the role of the plasmacytoid dendritic cell (PDC), the major producer of IFN-{alpha} upon viral infection, in tumor biology is unknown. We sought to study the presence and function of PDC in a human solid tumor. Here, we demonstrate that PDCs infiltrate tumor tissue of patients with head and neck squamous cell carcinoma (HNSCC). Functional activity of PDC was examined by using CpG motif containing oligonucleotides, a defined microbial stimulus for PDCs (recognized via toll-like receptor 9). We found that HNSCC diminished the ability of PDC to produce IFN-{alpha} in response to CpG motif containing oligonucleotide. Tumor-induced down-regulation of toll-like receptor 9 was identified as one mechanism likely contributing to impaired PDC function within the tumor environment. In tumor-draining lymph nodes, suppression of CpG-induced IFN-{alpha} production was less pronounced than in single-cell suspensions of primary tumor tissue. In these lymph nodes, CpG-induced IFN-{alpha} production was associated with increased levels of interferon-induced protein 10 and IFN-{gamma} and activation of CD4 and CD8 T cells. These results show for the first time the presence of PDCs in human solid tumor tissue and that tumors suppress the capacity of PDCs to produce IFN-{alpha}. PDCs, which in the absence of appropriate stimulation are reported to promote regulatory CD8 T cells, may contribute to an impaired T-cell-mediated immune response in HNSCC
22
Differential inhibition of human cytomegalovirus (HCMV) by toll-like receptor ligands mediated by interferon-beta in human foreskin fibroblasts and cervical tissue
Harwani,Sailesh C.; Lurain,Nell S.; Zariffard,M.Reza; Spear,Gregory T.
Virology Journal 2007;4:133› Link
Abstract
Human cytomegalovirus (HCMV) can be acquired sexually and is shed from the genital tract. Cross-sectional studies in women show that changes in genital tract microbial flora affect HCMV infection and/or shedding. Since genital microbial flora may affect HCMV infection or replication by stimulating cells through Toll-like receptors (TLR), we assessed the effects of defined TLR-ligands on HCMV replication in foreskin fibroblasts and ectocervical tissue. Poly I:C (a TLR3-ligand) and lipopolysaccharide (LPS, a TLR4-ligand) inhibited HCMV and induced secretion of IL-8 and Interferon-beta (IFN¦) in both foreskin fibroblasts and ectocervical tissue. The anti-HCMV effect was reversed by antibody to IFN¦. CpG (TLR9 ligand) and lipoteichoic acid (LTA, TLR2 ligand) also inhibited HCMV infection in ectocervical tissue and this anti-HCMV effect was also reversed by anti-IFN¦ antibody. In contrast, LTA and CpG did not inhibit HCMV infection in foreskin fibroblasts. This study shows that TLR ligands induce an HCMV-antiviral effect that is mediated by IFN¦ suggesting that changes in genital tract flora may affect HCMV infection or shedding by stimulating TLR. This study also contrasts the utility of two models that can be used for assessing the interaction of microbial flora with HCMV in the genital tract. Clear differences in the response to different TLR ligands suggests the explant model more closely reflects in vivo responses to genital infections
2645
CD4 Binding Affinity Determines Human Immunodeficiency Virus Type 1-Induced Alpha Interferon Production in Plasmacytoid Dendritic Cells
Haupt,Sabrina; Donhauser,Norbert; Chaipan,Chawaree; Schuster,Philipp; Puffer,Bridget; Daniels,Rod S.; Greenough,Thomas C.; Kirchhoff,Frank; Schmidt,Barbara
Journal of Virology 2008;82:8900-8905.
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Abstract
Plasmacytoid dendritic cells (PDC) are major producers of type I interferons (IFN) in response to human immunodeficiency virus type 1 (HIV-1) infection. To better define the underlying mechanisms, we studied the magnitude of alpha IFN (IFN-{alpha}) induction by recombinant viruses containing changes in the Env protein that impair or disrupt CD4 binding or expressing primary env alleles with differential coreceptor tropism. We found that the CD4 binding affinity but not the viral coreceptor usage is critical for the attachment of autofluorescing HIV-1 to PDC and for subsequent IFN-{alpha} induction. Our results illustrate the importance of the gp120-CD4 interaction in determining HIV-1-induced immune stimulation via IFN-{alpha} production
2414
Cell-Cell Fusion Induced by Measles Virus Amplifies the Type I Interferon Response
Herschke,F.; Plumet,S.; Duhen,T.; Azocar,O.; Druelle,J.; Laine,D.; Wild,T.F.; Rabourdin-Combe,C.; Gerlier,D.; Valentin,H.
Journal of Virology 2007;81:12859-12871.
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Abstract
Measles virus (MeV) infection is characterized by the formation of multinuclear giant cells (MGC). We report that beta interferon (IFN-{beta}) production is amplified in vitro by the formation of virus-induced MGC derived from human epithelial cells or mature conventional dendritic cells. Both fusion and IFN-{beta} response amplification were inhibited in a dose-dependent way by a fusion-inhibitory peptide after MeV infection of epithelial cells. This effect was observed at both low and high multiplicities of infection. While in the absence of virus replication, the cell-cell fusion mediated by MeV H/F glycoproteins did not activate any IFN-{alpha}/{beta} production, an amplified IFN-{beta} response was observed when H/F-induced MGC were infected with a nonfusogenic recombinant chimerical virus. Time lapse microscopy studies revealed that MeV-infected MGC from epithelial cells have a highly dynamic behavior and an unexpected long life span. Following cell-cell fusion, both of the RIG-I and IFN-{beta} gene deficiencies were trans complemented to induce IFN-{beta} production. Production of IFN-{beta} and IFN-{alpha} was also observed in MeV-infected immature dendritic cells (iDC) and mature dendritic cells (mDC). In contrast to iDC, MeV infection of mDC induced MGC, which produced enhanced amounts of IFN-{alpha}/{beta}. The amplification of IFN-{beta} production was associated with a sustained nuclear localization of IFN regulatory factor 3 (IRF-3) in MeV-induced MGC derived from both epithelial cells and mDC, while the IRF-7 up-regulation was poorly sensitive to the fusion process. Therefore, MeV-induced cell-cell fusion amplifies IFN-{alpha}/{beta} production in infected cells, and this indicates that MGC contribute to the antiviral immune response
2552
Sequence-specific potent induction of IFN-[alpha] by short interfering RNA in plasmacytoid dendritic cells through TLR7
Hornung,Veit; Guenthner-Biller,Margit; Bourquin,Carole; Ablasser,Andrea; Schlee,Martin; Uematsu,Satoshi; Noronha,Anne; Manoharan,Muthiah; Akira,Shizuo; de Fougerolles,Antonin; Endres,Stefan; Hartmann,Gunther
Nat Med 2005;11:263-270.
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Abstract
Short interfering RNA (siRNA) is used in RNA interference technology to avoid non-target-related induction of type I interferon (IFN) typical for long double-stranded RNA. Here we show that in plasmacytoid dendritic cells (PDC), an immune cell subset specialized in the detection of viral nucleic acids and production of type I IFN, some siRNA sequences, independent of their GU content, are potent stimuli of IFN- production. Localization of the immunostimulatory motif on the sense strand of a potent IFN--inducing siRNA allowed dissection of immunostimulation and target silencing. Injection into mice of immunostimulatory siRNA, when complexed with cationic liposomes, induced systemic immune responses in the same range as the TLR9 ligand CpG, including IFN- in serum and activation of T cells and dendritic cells in spleen. Immunostimulation by siRNA was absent in TLR7-deficient mice. Thus sequence-specific TLR7-dependent immune recognition in PDC needs to be considered as an additional biological activity of siRNA, which then should be termed immunostimulatory RNA (isRNA).
1203
Replication-Dependent Potent IFN-{alpha} Induction in Human Plasmacytoid Dendritic Cells by a Single-Stranded RNA Virus
Hornung,Veit; Schlender,Jorg; Guenthner-Biller,Margit; Rothenfusser,Simon; Endres,Stefan; Conzelmann,Karl Klaus; Hartmann,Gunther
The Journal of Immunology 2004;173:5935-5943.
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Abstract
Plasmacytoid dendritic cells sense viral ssRNA or its degradation products via TLR7/8 and CpG motifs within viral DNA via TLR9. Although these two endosomal pathways operate independently of viral replication, little is known about the detection of actively replicating viruses in plasmacytoid dendritic cell (PDC). Replication and transcription of the viral genome of ssRNA viruses as well as many DNA viruses lead to the formation of cytosolic dsRNA absent in noninfected cells. In this study, we used human respiratory syncytial virus (HRSV) encoding a fusion (F) protein for direct cytosolic entry. Both HRSV infection and cytosolic delivery of a 65-nt dsRNA led to potent IFN-{alpha} induction in PDC, but not in myeloid dendritic cells. Inactivation of HRSV by UV irradiation abrogated IFN-{alpha} induction in PDC. The comparison of two respiratory syncytial virus (RSV) constructs carrying either the HRSV or the bovine RSV F protein revealed that F-mediated cytosolic entry of RSV was absolutely required for IFN-{alpha} induction in PDC. HRSV-induced IFN-{alpha} production was independent of endosomal acidification and of protein kinase R (PKR) kinase activity, as demonstrated with chloroquine and the PKR inhibitor 2-aminopurine, respectively. In contrast, the induction of IFN-{alpha} by the TLR7/8 ligand R848, by the TLR9 ligand CpG-A ODN 2216, and by inactivated influenza virus (TLR7/8 dependent) was completely blocked by 2-aminopurine. IFN-{alpha} induction by mouse pathogenic Sendai virus was not affected in PKR- and MyD88-deficient mice, confirming that a ssRNA virus, which is able to directly enter host cells via fusion at the plasma membrane, can be detected by PDC independently of PKR, TLR7/8, and TLR9
355
Recombinant Newcastle disease virus (NDV) with inserted gene coding for GM-CSF as a new vector for cancer immunogene therapy
Janke,M.; Peeters,B.; de Leeuw,O.; Moorman,R.; Arnold,A.; Fournier,P.; Schirrmacher,V.
Gene Ther 2007;14:1639-1649.
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Abstract
This is the first report describing recombinant (rec) Newcastle disease virus (NDV) as vector for gene therapy of cancer. The gene encoding granulocyte/macrophage colony-stimulating factor (GM-CSF) was inserted as an additional transcription unit at two different positions into the NDV genome. The rec virus with the strongest production of the gene product (rec(GM-CSF)) was selected for our study. The insertion of the new foreign gene did neither affect the main features of NDV replication nor its tumor selectivity. The gene product was biologically active and stable. Tumor vaccine cells infected by rec(GM-CSF) stimulated human peripheral blood mononuclear cells (PBMC) to exert antitumor bystander effects in vitro in a tumor neutralization assay. These effects were significantly increased when compared to vaccine infected by rec(-) virus. Furthermore, rec(GM-CSF) led to a much higher interferon- (IFN-) production than rec(-) when added as virus or as virus-modified vaccine to PBMC. Two distinct cell types, monocytes and plasmacytoid dendritic cells were shown to contribute to the augmented IFN- response of PBMC. In conclusion, the already inherent anti-neoplastic and immunostimulatory properties of NDV could be further augmented by the introduction of a therapeutic gene whose product initiates a broad cascade of immunological effects in the microenvironment of the vaccine.
2262
Enumeration and phenotypical analysis of distinct dendritic cell subsets in psoriatic arthritis and rheumatoid arthritis
Jongbloed,Sarah L.; Lebre,M.Cristina; Fraser,Alasdair R.; Gracie,J.Alastair; Sturrock,Roger D.; Tak,Paul P.; McInnes,Iain B.
Arthritis Research & Therapy 2005;8:R15› Link
Abstract
Dendritic cells (DCs) comprise heterogeneous subsets of professional antigen-presenting cells, linking innate and adaptive immunity. Analysis of DC subsets has been hampered by a lack of specific DC markers and reliable quantitation assays. We characterised the immunophenotype and functional characteristics of psoriatic arthritis (PsA)-derived and rheumatoid arthritis (RA)-derived myeloid DCs (mDCs) and plasmacytoid DCs (pDCs) to evaluate their potential role in arthritis. Circulating peripheral blood (PB) pDC numbers were significantly reduced in PsA patients ( P = 0.0098) and RA patients ( P = 0.0194), and mDCs were significantly reduced in RA patients ( P = 0.0086) compared with healthy controls. The number of circulating mDCs in RA PB was significantly inversely correlated to C-reactive protein ( P = 0.021). The phenotype of both DC subsets in PsA PB and RA PB was immature as compared with healthy controls. Moreover, CD62L expression was significantly decreased on both mDCs (PsA, P = 0.0122; RA, P = 0.0371) and pDCs (PsA, P = 0.0373; RA, P = 0.0367) in PB. Both mDCs and pDCs were present in PsA synovial fluid (SF) and RA SF, with the mDC:pDC ratio significantly exceeding that in matched PB (PsA SF, P = 0.0453; RA SF, P = 0.0082). pDCs isolated from RA SF and PsA SF displayed an immature phenotype comparable with PB pDCs. RA and PsA SF mDCs, however, displayed a more mature phenotype (increased expression of CD80, CD83 and CD86) compared with PB mDCs. Functional analysis revealed that both SF DC subsets matured following toll-like receptor stimulation. pDCs from PB and SF produced interferon alpha and tumour necrosis factor alpha on TLR9 stimulation, but only SF pDCs produced IL-10. Similarly, mDCs from PB and SF produced similar tumour necrosis factor alpha levels to TLR2 agonism, whereas SF mDCs produced more IL-10 than PB controls. Circulating DC subset numbers are reduced in RA PB and PsA PB with reduced CD62L expression. Maturation is incomplete in the inflamed synovial compartment. Immature DCs in SF may contribute to the perpetuation of inflammation via sampling of the inflamed synovial environment, and in situ presentation of arthritogenic antigen
973
Interleukin-18 attracts plasmacytoid dendritic cells (DC2) and promotes Th1 induction by DC2 through IL-18 receptor expression
Kaser,Arthur; Kaser,Susanne; Kaneider,Nicole C.; Enrich,Barbara; Wiedermann,Christian J.; Tilg,Herbert
Blood 2003;2002-2007.
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Abstract
In vivo evidence suggests that interleukin-18 (IL-18) shapes the development of adaptive immunity toward T helper cell type 1 (Th1) responses. Monocyte-derived dendritic cells 1 (DC1) preferentially induce a Th1 response, while plasmacytoid DC-derived DC2 have been linked to a Th2 response. We analyzed the role of IL-18 during the initiation phase of a Th response in vitro to elucidate the basis of the aforementioned in vivo observations. IL-18 was constitutively released from DC1, but not DC2. Neutralization of IL-18 in co-culture experiments of DC1 with allogeneic naive T lymphocytes did not alter the Th1/Th2 phenotype, while anti-IL-12 efficiently downregulated the Th1 response. Unexpectedly, IL-18R {alpha} and {beta} chain were expressed on DC2 lineage. IL-18R expression was functional as IL-18 induced chemotaxis in plasmacytoid DC (pre-DC2), and enhanced the allostimulatory capacity of IL-3-differentiated DC2. Pre-DC2 exposed to IL-18 skewed the development of Th cells toward Th1 in co-culture experiments of DC2 and allogeneic naive T cells, which was inhibited by IL-12 p70 neutralization. IL-18 might have a profound role during the initiation phase of an immune response by recruiting pre-DC2 and modulating the function of DC2
25
Immunostimulatory properties of CpG-oligonucleotides are enhanced by the use of protamine nanoparticles
Kerkmann,M.; Lochmann,D.; Weyermann,J.; Marschner,A.; Poeck,H.; Wagner,M.; Battiany,J.; Zimmer,A.; Endres,S.; Hartmann,G.
Oligonucleotides. 2006;16:313-322.
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Abstract
The aim of this paper was to investigate if the immunostimulatory effects of CpG-oligonucleotides (CpG-ODN) can be enhanced by the use of biodegradable protamine nanoparticles (proticles). We analyzed size, surface charge, and morphology of protamine nanoparticles containing CpG-ODN with photon correlation spectroscopy and transmission electron microscopy. Immunostimulatory effects of these nanoparticles on B cells, plasmacytoid dendritic cells (PDC), peripheral blood mononuclear cells, and whole blood were studied. Cytokine production, activation of the cells in terms of upregulation of surface molecules and uptake of nanoparticles were examined. We found that the use of protamine nanoparticles significantly increased (20-fold) CpG-ODN mediated interferon (IFN)-alpha production of PDC. ODN uptake in PDC was only marginally enhanced. CpG-ODN mediated IP-10 production in whole blood was strongly enhanced by the use of nanoparticles. Apart from a slight increase in CpG-ODN-induced interleukin (IL)-6 production in B cells, other parameters like the CpG-mediated activation of B cells and PDC as well as tumor necrosis factor (TNF)-alpha production of PDC remained largely unchanged. The use of control ODN indicated that the protamine nanoparticles themselves have no immunostimulatory properties. These results strongly support the use of particulate delivery systems like biodegradable protamine nanoparticles for the development of CpG-ODN-based therapeutics
2741
Spontaneous formation of nucleic acid-based nanoparticles is responsible for high IFN-a induction by CpG-A in plasmacytoid dendritic cells
Kerkmann,Miren; Costa,Lilian T.; Richter,Christine; Rothenfusser,Simon; Battiany,Julia; Hornung,Veit; Johnson,Judith; Englert,Steffen; Ketterer,Thomas; Heckl,Wolfgang; Thalhammer,Stefan; Endres,Stefan; Hartmann,Gunther
Journal of Biological Chemistry 2004;280:M410868200› Link
Abstract
Plasmacytoid dendritic cells (PDC) represent a highly specialized immune cell subset that produces large quantities of the anti-viral cytokines type I interferons (IFN-a and IFN-{beta}) upon viral infection. PDC employ a member of the family of toll-like receptors, TLR9, to detect CpG motifs (unmethylated CG dinucleotides in certain base context) present in viral DNA. A certain group of CpG motif containing oligodeoxynucleotides (CpG ODN), CpG-A, was the first synthetic stimulus available that induced large amounts of IFN-a in PDC. However, the mechanism responsible for this activity remained elusive. CpG-A is characterized by a central palindrome and poly G at the 5{acute} and 3{acute}end. Here we demonstrate that CpG-A self-assembles to higher-order tertiary structures via G-tetrad formation of their poly G motifs. Spontaneous G-tetrad formation of CpG-A required the palindrome sequence allowing structure formation in a physiological environment. Once formed, G-tetrad-linked structures were stable even under denaturing conditions. Atomic force microscopy revealed that the tertiary structures formed by CpG-A represent nucleic-acid-based nanoparticles in the size range of viruses. Similarly-sized preformed polystyrene nanoparticles, loaded with a CpG ODN that is otherwise weak at inducing IFN-a (CpG-B), gained the potency of CpG-A to induce IFN-a. Higher ODN uptake in PDC was not responsible for the higher IFN-a-inducing activity of CpG-A or of CpG-B-coated nanoparticles as compared to CpG-B. Based on these results we propose a model in which the spatial configuration of CpG motifs as particle is responsible for the virus-like potency of CpG-A to induce IFN-a in PDC
350
Activation with CpG-A and CpG-B Oligonucleotides Reveals Two Distinct Regulatory Pathways of Type I IFN Synthesis in Human Plasmacytoid Dendritic Cells
Kerkmann,Miren; Rothenfusser,Simon; Hornung,Veit; Towarowski,Andreas; Wagner,Moritz; Sarris,Anja; Giese,Thomas; Endres,Stefan; Hartmann,Gunther
The Journal of Immunology 2003;170:4465-4474.
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Abstract
Two different CpG oligonucleotides (ODN) were used to study the regulation of type I IFN in human plasmacytoid dendritic cells (PDC): ODN 2216, a CpG-A ODN, known to induce high amounts of IFN-{alpha} in PDC, and ODN 2006, a CpG-B ODN, which is potent at stimulating B cells. CpG-A ODN showed higher and prolonged kinetics of type I IFN production compared with that of CpG-B ODN. In contrast, CpG-B ODN was more active than CpG-A ODN in stimulating IL-8 production and increasing costimulatory and Ag-presenting molecules, suggesting that CpG-A and CpG-B trigger distinct regulatory pathways in PDC. Indeed, CpG-A ODN, but not CpG-B ODN, activated the type I IFNR-mediated autocrine feedback loop. PDC were found to express high constitutive levels of IFN regulatory factor (IRF)7. IRF7 and STAT1, but not IRF3, were equally up-regulated by both CpG-A and CpG-B. CD40 ligand synergistically increased CpG-B-induced IFN-{alpha} independent of the IFNR but did not affect CpG-B-induced IFN-{beta}. In conclusion, our studies provide evidence for the existence of two distinct regulatory pathways of type I IFN synthesis in human PDC, one dependent on and one independent of the IFNR-mediated feedback loop. The alternate use of these pathways is based on the type of stimulus rather than the quantity of IFN-{alpha}{beta} available to trigger the IFNR. Constitutive expression of IRF7 and the ability to produce considerable amounts of IFN-{alpha} independent of the IFNR seem to represent characteristic features of PDC
56
Impaired Plasmacytoid Dendritic Cell Innate Immune Responses in Patients with Herpes Virus-Associated Acute Retinal Necrosis
Kittan,Nicolai A.; Bergua,Antonio; Haupt,Sabrina; Donhauser,Norbert; Schuster,Philipp; Korn,Klaus; Harrer,Thomas; Schmidt,Barbara
The Journal of Immunology 2007;179:4219-4230.
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Abstract
Plasmacytoid dendritic cells (PDC), the main producers of type I IFNs in the blood, are important for the recognition and control of viral and bacterial infections. Because several viruses induce IFN-{alpha} production, severe courses of herpes virus infections in nonimmunocompromised patients may be related to numerical or functional PDC deficits. To evaluate this hypothesis, PBMC and PDC were repeatedly isolated from nine patients with acute retinal necrosis (ARN), caused by herpes simplex or varicella zoster virus. The patients experienced meningitis/encephalitis and frequent infections in childhood (n = 2), recurrent herpes virus infections at unusual localizations (n = 2), ocular surgery (n = 1), infections (n = 4), and stress around ARN (n = 6). The median percentage of isolated PDC was significantly lower in patients compared with 18 age-matched healthy controls (p < 0.001), confirmed by FACS analysis using peripheral blood, and was extremely low during acute disease. PDC counts dropped in five controls suffering from respiratory infections or diarrhea. IFN-{alpha} production in PDC and PBMC exposed to different stimuli was significantly lower in patients than in controls (p < 0.05). Anergy to these stimuli was observed on four occasions, in particular during acute disease. PDC of patients showed up-regulated IFN regulatory factor-7 mRNA levels and evidence of in vivo activation (CD80) and maturation (CD83) (p < 0.05). CD8+ cell responses were significantly lower in patients vs controls (p = 0.04). These data support a risk factor model in which numerical and functional deficits in PDC-mediated innate immune responses contribute to an impaired control of latent herpes virus infections and subsequent development of ARN
2488
Human recombinant interferon alfa-2a for the treatment of Behcet's disease with sight threatening posterior or panuveitis
Kotter,I.; Zierhut,M.; Eckstein,A.K.; Vonthein,R.; Ness,T.; Gunaydin,I.; Grimbacher,B.; Blaschke,S.; Meyer-Riemann,W.; Peter,H.H.; Stubiger,N.
British Journal of Ophthalmology 2003;87:423-431.
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Abstract
Background: Behcet's disease is a multisystem vasculitis of unknown origin. Standard treatment mainly comprises systemic immunosuppressive agents. Ocular involvement, mostly posterior uveitis with retinal vasculitis, leads to blindness in 20-50% of the involved eyes within 5 years. The efficacy of interferon alfa-2a was studied in patients with sight threatening posterior uveitis or retinal vasculitis. Methods: 50 patients were included in this open, non-randomised, uncontrolled prospective study. Recombinant human interferon alfa-2a (rhIFN{alpha}-2a) was applied at a dose of 6 million units subcutaneously daily. Dose reduction was performed according to a decision tree until discontinuation. Disease activity was evaluated every 2 weeks by the Behcet's disease activity scoring system and the uveitis scoring system. Results: Response rate of the ocular manifestations was 92% (three non-responder, one incomplete response). Mean visual acuity rose significantly from 0.56 to 0.84 at week 24 (p<0.0001). Posterior uveitis score of the affected eyes fell by 46% every week (p<0.001). Remission of retinal inflammation was achieved by week 24. Mean Behcet's disease activity score fell from 5.8 to 3.3 at week 24 and further to 2.8 at week 52. After a mean observation period of 36.4 months (range 12-72), 20 patients (40%) are off treatment and disease free for 7-58 months (mean 29.5). In the other patients maintenance IFN dosage is three million units three times weekly. Conclusions: rhIFN{alpha}-2a is effective in ocular Behcet's disease, leading to significant improvement of vision and complete remission of ocular vasculitis in the majority of the patients
65
CD11c+ dendritic cells and plasmacytoid DCs are activated by human cytomegalovirus and retain efficient T cell-stimulatory capability upon infection
Kvale,Espen O.; Dalgaard,Jakob; Lund-Johansen,Fridtjof; Rollag,Halvor; Farkas,Lorant; Midtvedt,Karsten; Jahnsen,Frode L.; Brinchmann,Jan E.; Olweus,Johanna
Blood 2006;107:2022-2029.
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Abstract
It has been suggested that human cytomegalovirus (HCMV) evades the immune system by infecting and paralyzing antigen-presenting cells. This view is based mainly on studies of dendritic cells (DCs) obtained after culture of monocytes (moDCs). It is contradicted by the asymptomatic course of HCMV infection in healthy persons, indicating that other key antigen-presenting cells induce an efficient immune response. Here we show that HCMV activates CD11c+ DCs and plasmacytoid DCs (PDCs). In contrast to moDCs, CD11c+ DCs and PDCs produced interferon (IFN) type 1 when exposed to HCMV. Autocrine IFN type 1 partially protected CD11c+ DCs against infection, whereas PDCs were resistant to HCMV even when IFN type 1 activity was inhibited. HCMV exposure induced the maturation of CD11c+ DCs by IFN type 1-dependent and -independent mechanisms. Importantly, CD11c+ DCs infected by inhibiting IFN type 1 activity retained full capacity to stimulate T cells. Renal transplant recipients receiving immunosuppressive treatment had lower frequencies of CD11c+ DCs and PDCs in blood than did healthy controls. The results show that HCMV activates the immune system by interacting with CD11c+ DCs and PDCs and that recipients of renal transplants have low frequencies of these cell types in blood
31
Plasmacytoid DCs regulate recall responses by rapid induction of IL-10 in memory T cells
Kvale,Espen O.; Floisand,Yngvar; Lund-Johansen,Fridtjof; Rollag,Halvor; Farkas,Lorant; Ghanekar,Smita; Brandtzaeg,Per; Jahnsen,Frode L.; Olweus,Johanna
Blood 2007;109:3369-3376.
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Abstract
Dendritic cells (DCs) are believed to regulate T cell-mediated immunity primarily by directing differentiation of naive T cells. Here, we show that a large fraction of CD4+ memory cells produce IL-10 within the first hours after interaction with plasmacytoid DCs (PDCs). In contrast, CD11c+ DCs induce IFN-{gamma} and little IL-10. IL-10-secreting T cells isolated after 36 hours of culture with PDCs suppressed antigen-induced T-cell proliferation by an IL-10-dependent mechanism, but were distinct from natural and type 1 regulatory T cells. They proliferated strongly and continued to secrete IL-10 during expansion with PDCs, and after restimulation with immature monocyte-derived DCs or CD11c+ DCs. The IL-10-producing T cells acquired the ability to secrete high levels of IFN-{gamma} after isolation and subsequent coculture with PDCs or CD11c+ DCs. Compared to CD11c+ DCs, PDCs were superior in their ability to selectively expand T cells that produced cytokines on repeated antigenic challenge. The DC-dependent differences in cytokine profiles were observed with viral recall antigen or staphylococcal enterotoxin B and were independent of extracellular type I interferon or IL-10. Our results show that DCs can regulate memory responses and that PDCs rapidly induce regulatory cytokines in effector T cells that can suppress bystander activity
825
Dysfunctional interferon-¦ production by peripheral plasmacytoid dendritic cells upon Toll-like receptor-9 stimulation in patients with systemic lupus erythematosus
Kwok,Seung Ki; Lee,June Yong; Park,Se Ho; Cho,Mi La; Min,So Youn; Park,Sung Hwan; Kim,Ho Youn; Cho,Young Gyu
Arthritis Research & Therapy 2008;10:R29› Link
Abstract
It is well known that interferon (IFN)-¦ is important to the pathogenesis of systemic lupus erythematosus (SLE). However, several reports have indicated that the number of IFN-¦ producing cells are decreased or that their function is defective in patients with SLE. We studied the function of plasmacytoid dendritic cells (pDCs) under persistent stimulation of Toll-like receptor (TLR)9 via a TLR9 ligand (CpG ODN2216) or SLE serum. Methods The concentrations of IFN-¦ were determined in serum and culture supernatant of peripheral blood mononuclear cells (PBMCs) from SLE patients and healthy controls after stimulation with CpG ODN2216 or SLE serum. The numbers of circulating pDCs were analyzed by fluoresence-activated cell sorting analysis. pDCs were treated with CpG ODN2216 and SLE serum repeatedly, and levels of produced IFN-¦ were measured. The expression of IFN-¦ signature genes and inhibitory molecules of TLR signaling were examined in PBMCs from SLE patients and healthy control individuals. Results Although there was no significant difference in serum concentration of IFN-¦ and number of circulating pDCs between SLE patients and healthy control individuals, the IFN-¦ producing capacity of PBMCs was significantly reduced in SLE patients. Interestingly, the degree which TLR9 ligand-induced IFN-¦ production in SLE PBMCs was inversely correlated with the SLE serum-induced production of IFN-¦ in healthy PMBCs. Because repeated stimulation pDCs with TLR9 ligands showed decreased level of IFN-¦ production, continuous TLR9 stimulation may lead to decreased production of IFN-¦ in SLE PBMCs. In addition, PBMCs isolated from SLE patients exhibited higher expression of IFN-¦ signature genes and inhibitory molecules of TLR signaling, indicating that these cells had already undergone IFN-¦ stimulation and had become desensitized to TLR signaling. Conclusion We suggest that the persistent presence of endogenous IFN-¦ inducing factors induces TLR tolerance in pDCs of SLE patients, leading to impaired production of IFN-¦
2664
Circulating dendritic cells and interferon-alpha production in patients with tuberculosis: correlation with clinical outcome and treatment response
Lichtner,M.; Rossi,R.; Mengoni,F.; Vignoli,S.; Colacchia,B.; Massetti,A.P.; Kamga,I.; Hosmalin,A.; Vullo,V.; Mastroianni,C.M.
Clinical and Experimental Immunology 2006;143:329-337.
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Abstract
Summary Dendritic cells (DC) have been characterized recently as having an important role in the initiation and control of immunological response to Mycobacterium tuberculosis infection. Blood DC have been subdivided into myeloid (mDC) and plasmacytoid (pDC) subsets, on the basis of differences in phenotype markers and function. Little is known about the enumeration and functional evaluation of circulating DC in patients with tuberculosis and their correlation with clinical outcome during the course of anti-tuberculous treatment. We assessed circulating mDC and pDC counts measured by a newly developed single-platform flow cytometric assay based on TruCOUNT, as well as the production of interferon (IFN)-alpha after in vitro stimulation by herpes simplex virus (HSV-1) in 24 patients with active tuberculosis (TB) and 37 healthy donors. Absolute numbers of both DC subsets were decreased significantly in patients with active TB compared to controls. Similarly, the production of IFN-alpha was highly impaired. In 13 patients these parameters were assessed longitudinally, before and after the specific anti-microbial treatment. Most interestingly, in all nine patients with successful anti-tuberculous therapy there was a significant and marked increase of pDC counts and IFN-alpha production. In contrast, no significant longitudinal variations in DC counts and IFN-alpha production were observed in four patients with lack of response to specific treatment. In conclusion, active TB is associated with a defect in blood DC numbers and IFN-alpha production that is restored after bacterial clearance and clinical improvement, as a result of effective anti-tuberculous treatment
633
Induction of antigen-specific CD8+ cytotoxic T cells by dendritic cells co-electroporated with a dsRNA analogue and tumor antigen mRNA
Michiels,A.; Breckpot,K.; Corthals,J.; Tuyaerts,S.; Bonehill,A.; Heirman,C.; Thielemans,K.; Aerts,J.
Gene Ther 2006;13:1027-1036.
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Abstract
The maturation state of dendritic cells (DCs) is an important determinant for the initiation and regulation of adaptive immune responses. In this study, we wanted to assess whether functional activation of human monocyte-derived DCs can be achieved by electroporation of an activation signal in the form of double-stranded (ds) RNA and whether simultaneous electroporation of the dsRNA with tumor antigen encoding mRNA can lead to the induction of a cytotoxic T-lymphocyte (CTL) response. Electroporation of immature DCs with poly(I:C12U), a dsRNA analogue, resulted in phenotypic as well as functional changes, indicative of DC maturation. Co-electroporation of DCs with both poly(I:C12U) and Melan-A/MART-1 encoding mRNA induced strong anti-Melan-A/MART-1 CD8+ T-cell responses in vitro. Higher numbers of Melan-A/MART-1-specific CTLs were consistently obtained with poly(I:C12U)-activated DCs compared to DCs matured in the presence of an inflammatory cytokine cocktail. These results indicate that DC co-electroporation with both dsRNA and tumor antigen encoding mRNA induces fully activated and antigen-loaded DCs that promote antigen-specific CTL responses and may provide the basis for future immunotherapeutic strategies.
1223
Electroporation of immature and mature dendritic cells: implications for dendritic cell-based vaccines
Michiels,A.; Tuyaerts,S.; Bonehill,A.; Corthals,J.; Breckpot,K.; Heirman,C.; van Meirvenne,S.; Dullaers,M.; Allard,S.; Brasseur,F.; van der Bruggen,P.; Thielemans,K.
Gene Therapy 2005;12:772-782.
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Abstract
Until now, studies utilizing mRNA electroporation as a tool for the delivery of tumor antigens to human monocyte-derived dendritic cells (DC) have focused on DC electroporated in an immature state. Immature DC are considered to be specialized in antigen capture and processing, whereas mature DC present antigen and have an increased T-cell stimulatory capacity. Therefore, the consensus has been to electroporate DC before maturation. We show that the transfection efficiency of DC electroporated either before or after maturation was similarly high. Both immature and mature electroporated DC, matured in the presence of an inflammatory cytokine cocktail, expressed mature DC surface markers and preserved their capacity to secrete cytokines and chemokines upon CD40 ligation. In addition, both immature and mature DC can be efficiently cryopreserved before or after electroporation without deleterious effects on viability, phenotype or T-cell stimulatory capacity including in vitro antigen-specific T-cell activation. However, DC electroporated after maturation are more efficient in in vitro migration assays and at least as effective in antigen presentation as DC electroporated before maturation. These results are important for vaccination strategies where an optimal antigen presentation by DC after migration to the lymphoid organs is crucial.
1222
Impact of plasmacytoid dendritic cells on outcome after reduced-intensity conditioning allogeneic stem cell transplantation
Mohty,M.; Blaise,D.; Faucher,C.; Bardou,J.; Gastaut,A.; Viens,P.; Olive,D.; Gaugler,B.
Leukemia 2004;19:1-6.
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Abstract
The reconstitution of the plasmacytoid dendritic cells (PDCs) compartment might influence outcome after allogeneic stem cell transplantation (allo-SCT). Thus, we investigated the impact of blood PDCs measured at the third month after reduced-intensity conditioning (RIC) in 54 patients who received an HLA-identical sibling allo-SCT. The absence of grade II-IV acute graft-versus-host-disease (GVHD) was associated with an improved PDC count at 3 months after RIC-allo-SCT (P=0.003; OR=6.4; 95% CI, 1.9-22). The CD34+ stem cell dose and other lymphoid subsets infused with the allograft did not affect PDC recovery. Although PDC count could not predict death from progression or relapse, patients with a 'high' PDC recovery profile had an improved overall survival (OS; P=0.03), in contrast to patients with a 'low' PDC recovery profile who had an increased incidence of nonrelapse mortality (GVHD, infections) (P=0.03). The overall incidence of late infections (viral, fungal and bacterial) was significantly higher in the 'low' PDC recovery group as compared to the 'high' PDC recovery group (59 vs 19%; P=0.002). In a multivariate analysis, only a 'high' PDC count was significantly predictive of a decreased risk of death (P=0.04; RR=0.34; 95% CI, 0.12-0.96). Monitoring of PDCs at 3 months after RIC-allo-SCT may be a useful indicator predictor of long-term outcome.
1225
High Mobility Group B1 Protein Suppresses the Human Plasmacytoid Dendritic Cell Response to TLR9 Agonists
Popovic,Petar J.; DeMarco,Richard; Lotze,Michael T.; Winikoff,Steven E.; Bartlett,David L.; Krieg,Arthur M.; Guo,Z.Sheng; Brown,Charles K.; Tracey,Kevin J.; Zeh,Herbert J.,III
The Journal of Immunology 2006;177:8701-8707.
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Abstract
Plasmacytoid dendritic cells (PDC) are innate immune effector cells that are recruited to sites of chronic inflammation, where they modify the quality and nature of the adaptive immune response. PDCs modulate adaptive immunity in response to signals delivered within the local inflammatory milieu by pathogen- or damage-associated molecular pattern, molecules, and activated immune cells (including NK, T, and myeloid dendritic cells). High mobility group B1 (HMGB1) is a recently identified damage-associated molecular pattern that is released during necrotic cell death and also secreted from activated macrophages, NK cells, and mature myeloid dendritic cells. We have investigated the effect of HMGB1 on the function of PDCs. In this study, we demonstrate that HMGB1 suppresses PDC cytokine secretion and maturation in response to TLR9 agonists including the hypomethylated oligodeoxynucleotide CpG- and DNA-containing viruses. HMGB1-inhibited secretion of several proinflammatory cytokines including IFN-{alpha}, IL-6, TNF-{alpha}, inducible protein-10, and IL-12. In addition, HMGB1 prevented the CpG induced up-regulation of costimulatory molecules on the surface of PDC and potently suppressed their ability to drive generation of IFN-{gamma}-secreting T cells. Our observations suggest that HMGB1 may play a critical role in regulating the immune response during chronic inflammation and tissue damage through modulation of PDC function
748
Toll-Like Receptor 9-Dependent Immune Activation by Unmethylated CpG Motifs in Aspergillus fumigatus DNA
Ramirez-Ortiz,Zaida G.; Specht,Charles A.; Wang,Jennifer P.; Lee,Chrono K.; Bartholomeu,Daniella C.; Gazzinelli,Ricardo T.; Levitz,Stuart M.
Infection and Immunity 2008;76:2123-2129.
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Abstract
Phagocytic defenses are critical for effective host defenses against the opportunistic fungal pathogen Aspergillus fumigatus. Previous studies found that following challenge with A. fumigatus, Toll-like receptor 9 (TLR9) knockout mice survived longer than wild-type mice. However, the mechanism responsible was not defined. Here we demonstrate that A. fumigatus contains unmethylated CpG sequences, the natural ligands for TLR9. A. fumigatus DNA and synthetic CpG-rich oligodeoxynucleotides (ODNs) containing sequences found in the A. fumigatus genome potently stimulated the production of proinflammatory cytokines in mouse bone marrow-derived dendritic cells (BMDCs) and human plasmacytoid dendritic cells. The response was decreased when the fungal DNA was treated with a CpG methylase or with CpG-specific endonucleases. A role for TLR9 was demonstrated as cytokine production was abolished in BMDCs from TLR9-deficient mice. Moreover, transfection of HEK293 cells with human TLR9 conferred responsiveness to synthetic CpG-rich ODNs containing sequences found in A. fumigatus DNA. Taken together, these data demonstrate that TLR9 detects A. fumigatus DNA, resulting in the secretion of proinflammatory cytokines, which may contribute to the immune response to the pathogen
2432
TLR7/8 Triggering Exerts Opposing Effects in Acute versus Latent HIV Infection
Schlaepfer,Erika; Audige,Annette; Joller,Helene; Speck,Roberto F.
The Journal of Immunology 2006;176:2888-2895.
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Abstract
TLRs trigger innate immunity by recognizing conserved motifs of microorganisms. Recently, ssRNAs from HIV and influenza virus were shown to trigger TLR7 and 8. Thus, we hypothesized that HIV ssRNA, by triggering TLR7/8, affects HIV pathogenesis. Indeed, HIV ssRNA rendered human lymphoid tissue of tonsillar origin or PBMC barely permissive to HIV replication. The synthetic compound R-848, which also triggers TLR7/8, showed similar anti-HIV activity. Loss of R-848's activity in lymphoid tissue depleted of B cells suggested a role for B cells in innate immunity. TLR7/8 triggering appears to exert antiviral effects through soluble factors: conditioned medium reduced HIV replication in indicator cells. Although a number of cytokines and chemokines were increased upon adding R-848 to lymphoid tissue, blocking those cytokines/chemokines (i.e., IFN-{alpha} receptor, IFN-{gamma}, MIP-1{alpha}, -1[beta], RANTES, and stromal cell-derived factor-1) did not result in the reversal of R-848's anti-HIV activity. Thus, the nature of this soluble factor(s) remains unknown. Unlike lymphoid tissue acutely infected with HIV, triggering latently infected promonocytic cells induced the release of HIV virions. The anti-HIV effects of triggering TLR7/8 may inhibit rapid killing, while pro-HIV effects may guarantee a certain replication level. Compounds triggering TLR7/8 may be attractive drug candidates to purge latent HIV while preventing new infections
519
CpG Oligodeoxynucleotides Block Human Immunodeficiency Virus Type 1 Replication in Human Lymphoid Tissue Infected Ex Vivo
Schlaepfer,Erika; Audige,Annette; von Beust,Barbara; Manolova,Vania; Weber,Markus; Joller,Helene; Bachmann,Martin F.; Kundig,Thomas M.; Speck,Roberto F.
Journal of Virology 2004;78:12344-12354.
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Abstract
Oligodeoxynucleotides (ODNs) with immunomodulatory motifs control a number of microbial infections in animal models, presumably by acting through toll-like receptor 9 (TLR9) to induce a number of cytokines (e.g., alpha interferon and tumor necrosis factor alpha). The immunomodulatory motif consists of unmethylated sequences of cytosine and guanosine (CpG motif). ODNs without CpG motifs do not trigger TLR9. We hypothesized that triggering of TLR9 generates a cellular environment unfavorable for human immunodeficiency virus (HIV) replication. We tested this hypothesis in human lymphocyte cultures and found that phosphorothioate-modified ODN CpG2006 (type B ODNs) inhibited HIV replication nearly completely and prevented the loss of CD4+ T cells. ODNs CpG2216 and CpG10 (type A ODNs) were less effective. CpG2006 blocked HIV replication in purified CD4+ T cells and T-cell lines; CpG10 was ineffective in this setting, indicating that type A ODNs may inhibit HIV replication in CD4+ T-cell lines indirectly through a separate cell subset. However, control ODNs without CpG motifs also showed anti-HIV effects, indicating that these effects are nonspecific and not due to TLR9 triggering. The mechanism of action is not clear. CpG2006 and its control ODN blocked syncytium formation in a cell fusion-based assay, but CpG10, CpG2216, and their control ODNs did not. The latter types interfered with the HIV replication cycle during disassembly or reverse transcription. In contrast, CpG2006 and CpG2216 specifically induced cytokines critical to initiation of the innate immune response. In summary, the nonspecific anti-HIV activity of CpG ODNs, their ability to stimulate HIV replication in latently infected cells, potentially resulting in their elimination, and their documented ability to link the innate and adaptive immune responses make them attractive candidates for further study as anti-HIV drugs
441
Inhibition of Toll-Like Receptor 7- and 9-Mediated Alpha/Beta Interferon Production in Human Plasmacytoid Dendritic Cells by Respiratory Syncytial Virus and Measles Virus
Schlender,Jorg; Hornung,Veit; Finke,Stefan; Gunthner-Biller,Margit; Marozin,Sabrina; Brzozka,Krzysztof; Moghim,Sharareh; Endres,Stefan; Hartmann,Gunther; Conzelmann,Karl Klaus
Journal of Virology 2005;79:5507-5515.
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Abstract
Human plasmacytoid dendritic cells (PDC) are key sentinels alerting both innate and adaptive immune responses through production of huge amounts of alpha/beta interferon (IFN). IFN induction in PDC is triggered by outside-in signal transduction pathways through Toll-like receptor 7 (TLR7) and TLR9 as well as by recognition of cytosolic virus-specific patterns. TLR7 and TLR9 ligands include single-stranded RNA and CpG-rich DNA, respectively, as well as synthetic derivatives thereof which are being evaluated as therapeutic immune modulators promoting Th1 immune responses. Here, we identify the first viruses able to block IFN production by PDC. Both TLR-dependent and -independent IFN responses are abolished in human PDC infected with clinical isolates of respiratory syncytial virus (RSV), RSV strain A2, and measles virus Schwarz, in contrast to RSV strain Long, which we previously identified as a potent IFN inducer in human PDC (Hornung et al., J. Immunol. 173:5935-5943, 2004). Notably, IFN synthesis of PDC activated by the TLR7 and TLR9 agonists resiquimod (R848) and CpG oligodeoxynucleotide 2216 is switched off by subsequent infection by RSV A2 and measles virus. The capacity of RSV and measles virus of human PDC to shut down IFN production should contribute to the characteristic features of these viruses, such as Th2-biased immune pathology, immune suppression, and superinfection
405
Human Cytomegalovirus Impairs the Function of Plasmacytoid Dendritic Cells in Lymphoid Organs
Schneider,Kerstin; Meyer-Koenig,Ursula; Hufert,Frank T.
PLoS ONE 2008;3:› Link
Abstract
Human dendritic cells (DCs) are the main antigen presenting cells (APC) and can be divided into two main populations, myeloid and plasmacytoid DCs (pDCs), the latter being the main producers of Type I Interferon. The vast majority of pDCs can be found in lymphoid organs, where the main pool of all immune cells is located, but a minority of pDCs also circulate in peripheral blood. Human cytomegalovirus (HCMV) employs multiple mechanisms to evade the immune system. In this study, we could show that pDCs obtained from lymphoid organs (tonsils) (tpDCs) and from blood (bpDCs) are different subpopulations in humans. Interestingly, these populations react in opposite manner to HCMV-infection. TpDCs were fully permissive for HCMV. Their IFN-¦ production and the expression of costimulatory and adhesion molecules were altered after infection. In contrast, in bpDCs HCMV replication was abrogated and the cells were activated with increased IFN-¦ production and upregulation of MHC class I, costimulatory, and adhesion molecules. HCMV-infection of both, tpDCs and bpDCs, led to a decreased T cell stimulation, probably mediated through a soluble factor produced by HCMV-infected pDCs. We propose that the HCMV-mediated impairment of tpDCs is a newly discovered mechanism selectively targeting the host's major population of pDCs residing in lymphoid organs
2698
Correlation of in vivo and in vitro release data for rh-INF[alpha] lipid implants
Schwab,M.; Kessler,B.; Wolf,E.; Jordan,G.; Mohl,S.; Winter,G.
European Journal of Pharmaceutics and Biopharmaceutics 2008;70:690-694.
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Abstract
Previous in vitro experiments had shown that rh-INF[alpha] releasing tristearin implants feature promising properties making them an excellent tool for the delivery of therapeutic proteins. Sustained release for periods up to one month could be achieved, associated with high protein stabilization. The objective of this study was to investigate for the first time the in vivo release properties of these implants in rabbits and to establish an in vivo-in vitro correlation. Computer modeling was used to simulate rh-INF[alpha] serum levels based on pharmacokinetic data. Protein serum concentrations on therapeutically relevant nearly constant levels could be detected for 9 days. Modeling revealed that in vivo release correlated closely with the release monitored in vitro.
532
Impaired maturation and altered regulatory function of plasmacytoid dendritic cells in multiple sclerosis
Stasiolek,Mariusz; Bayas,Antonios; Kruse,Niels; Wieczarkowiecz,Anja; Toyka,Klaus V.; Gold,Ralf; Selmaj,Krzysztof
Brain 2006;129:1293-1305.
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Abstract
Plasmacytoid dendritic cells (pDCs) represent a DC subtype that exerts divergent functions in innate and adoptive immunity including the immediate reaction to microbial factors and the induction of immunoregulatory responses. It is thought that different DC subtypes may be critically involved in the pathogenesis of multiple sclerosis (MS). In our study we assessed the phenotype, maturation and functional properties of peripheral blood pDCs from 35 clinically stable, untreated multiple sclerosis patients, 30 healthy controls and 9 patients with pneumonia, which was used as a non-specific inflammatory condition (NIC). Ex vivo expression of CD86 and 4-1BBL was significantly lower on pDCs from multiple sclerosis patients than from controls and patients with NIC (22 versus 47 versus 41% and 12 versus 35 versus 32%, respectively). When stimulated with IL-3 and CD40L, pDCs of multiple sclerosis patients showed inefficient maturation as demonstrated by significantly lower or delayed upregulation of CD86, 4-1BBL, CD40 and CD83. Additionally, in multiple sclerosis, stimulation of pDCs by unmethylated cytosine-phosphate-guanosine oligodeoxynucleotides (CpG ODN) resulted in a significantly lower interferon (IFN) alpha secretion than in controls. In multiple sclerosis, but not in controls, pDCs failed to upregulate proliferative responses and IFN-gamma secretion of autologous peripheral blood mononuclear cells (PBMC) in a co-culture system. Moreover, depletion of pDCs in multiple sclerosis patients, but not in controls, had no effect on generation of CD4+Foxp3+ regulatory T cells. We also provide data showing that glatiramer acetate (GA) treatment partially restores phenotype and function of pDCs in multiple sclerosis patients. These findings suggest functional abnormalities of pDCs in these patients, which might be of importance in the understanding of the development of immune dysregulation in this disease
10
Increased Expression of Leukocyte Ig-Like Receptor-1 and Activating Role of UL18 in the Response to Cytomegalovirus Infection
Wagner,Claudia S.; Riise,Gerdt C.; Bergstrom,Tomas; Karre,Klas; Carbone,Ennio; Berg,Louise
The Journal of Immunology 2007;178:3536-3543.
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Abstract
NK and T cells are important for combating CMV infection. Some NK and T cells express leukocyte Ig-like receptor-1 (LIR-1), an inhibitory receptor recognizing MHC class I and the CMV-encoded homolog UL18. We previously demonstrated an early increase in LIR-1-expressing blood lymphocytes in lung-transplanted patients later developing CMV disease. We now show that NK and T cells account for the observed LIR-1 augmentation. Coincubation of PBMC from CMV-seropositive donors with virus-infected lung fibroblasts led to a T cell-dependent secretion of IFN-{gamma}, produced mainly by LIR-1+ T cells and by NK cells. Cytokine production during coculture with fibroblasts infected with virus containing the UL18 gene was augmented compared with the UL18 deletion virus, suggesting a stimulatory role for UL18. However, purified UL18Fc proteins inhibited IFN-{gamma} production of LIR-1+ T cells. We propose that cytokine production in the transplant induces NK and T cells to express LIR-1, which may predispose to CMV disease by MHC/LIR-1-mediated suppression. Although the UL18/LIR-1 interaction could inhibit T cell responses, this unlikely plays a role in response to infected cells. Instead, our data point to an activating role for viral UL18 during infection, where indirect intracellular effects cannot be excluded
874
Cutting Edge: Antibody-Mediated TLR7-Dependent Recognition of Viral RNA
Wang,Jennifer P.; Asher,Damon R.; Chan,Melvin; Kurt-Jones,Evelyn A.; Finberg,Robert W.
The Journal of Immunology 2007;178:3363-3367.
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Abstract
TLR7 recognizes the genome of ssRNA viruses such as Coxsackievirus B. Because TLR7 is expressed in intracellular compartments, viral RNA must be internalized before its recognition by TLR7. In this study, we define plasmacytoid dendritic cells (pDC) as peripheral blood mononuclear immune cells that respond to Coxsackievirus. pDC activation by Coxsackievirus B requires the presence of specific antiviral Abs. We show that Fc receptors mediate the recognition of virus-Ab complexes and that TLR7 is required for human and murine pDC production of cytokines. These data define a pathway by which intracellular TLR7 senses viral RNA and indicate a role for TLRs in association with Abs in sustaining virus-specific responses
877
Flavivirus Activation of Plasmacytoid Dendritic Cells Delineates Key Elements of TLR7 Signaling beyond Endosomal Recognition
Wang,Jennifer P.; Liu,Ping; Latz,Eicke; Golenbock,Douglas T.; Finberg,Robert W.; Libraty,Daniel H.
The Journal of Immunology 2006;177:7114-7121.
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Abstract
TLR7 senses RNA in endosomal compartments. TLR7 expression and signaling have been demonstrated in plasmacytoid and myeloid dendritic cells, B cells, and T cells. The regulation of TLR7 signaling can play a crucial role in shaping the immune response to RNA viruses with different cellular tropisms, and in developing adjuvants capable of promoting balanced humoral and cell-mediated immunity. We used unique characteristics of two ssRNA viruses, dengue virus and influenza virus, to delineate factors that regulate viral RNA-human TLR7 signaling beyond recognition in endosomal compartments. Our data show that TLR7 recognition of enveloped RNA virus genomes is linked to virus fusion or uncoating from the endosome. The signaling threshold required to activate TLR7-type I IFN production is greater than that required to activate TLR7-NF-{kappa}B-IL-8 production. The higher order structure of viral RNA appears to be an important determinant of TLR7-signaling potency. A greater understanding of viral RNA-TLR7 activity relationships will promote rational approaches to interventional and vaccine strategies for important human viral pathogens
764
Implications of previous subclinical dengue infection but not virus load in dengue hemorrhagic fever
Yeh,Wen Ting; Chen,Rong Fu; Wang,Lin; Liu,Jien Wei; Shaio,Men Fang; Yang,Kuender D.
FEMS Immunology & Medical Microbiology 2006;48:84-90.
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Abstract
Abstract In a study comparing the virus load and immune reaction between patients with primary and secondary dengue-2 (DEN-2) infections in a hospital-based analysis, we found that 40.7% (55/135) of the 135 patients had secondary DEN-2 infection following a DEN-2 outbreak in southern Taiwan. Most of the secondary infections had subclinical primary dengue infections (78.2%; 43/55). Patients with secondary DEN-2 infections had lower platelet counts, and blood interferon-alpha and virus load, but significantly higher interleukin-10 (P=0.030) and anti-DEN-1 neutralization titers (P=0.013) than those with primary infection. Patients with secondary DEN-2 infection also had a higher rate of dengue hemorrhagic fever (DHF) (61.7% vs. 36.3%). A previous subclinical dengue infection is involved in the secondary DEN-2 infection associated with altered immune reaction and higher DHF rate, but lower blood virus load
1453
