Preferential depletion of CD2(low) plasmacytoid dendritic cells in HIV-infected subjects

Plasmacytoid dendritic cells (pDCs) are decreased in number and are functionally impaired in HIV act reasons for pDCs depletion are still unknown. It was recently reported that pDCs can be divided into two functionally distinct populations based on their CD2 expression level. To determine how the CD2(high) and CD2(low) populations are affected by HIV infection, we analyzed their frequencies in the peripheral blood of HIV-infected subjects and healthy controls. We found that the CD2(low) pDC subset was preferentially depleted in infected individuals. The frequency of CD2(low) pDCs correlated with the CD4(+) T-cell count but not with the plasma viral load. This finding furthers our understanding of the causes and consequences of pDC depletion during HIV infection.     

Human dendritic cell 1 and dendritic cell 2 subsets express FcepsilonRI: correlation with serum IgE and allergic asthma

BACKGROUND: Type 1 dendritic cells (DC1) express the high-affinity IgE receptor (FcepsilonRI); however, the regulation of FcepsilonRI expression by DCs is not well understood. Type 2 DC (DC2) expression of FcepsilonRI has not been demonstrated. OBJECTIVE: We hypothesized that DC2 cellsalso express FcepsilonRI and that expression of FcepsilonRI by the DC1 and DC2 subsets correlates with serum IgE and allergic asthma disease status. METHODS: To test these hypotheses, we quantitated FcepsilonRI alpha chain expression by the peripheral blood precursor DC1 (pDC1) and pDC2 subsets by using flow cytometry. RESULTS: FcepsilonRI was expressed by the pDC1 and pDC2 subsets, as well as tissue DCs from tonsils. Relative FcepsilonRI expression by basophil, pDC1, and pDC2 subsets was 12:6.5:1, respectively. In both pDC subsets, FcepsilonRI expression was significantly greater in allergic asthmatic subjects than in nonatopic control subjects. pDC1 and pDC2 expression of FcepsilonRI was highly correlated to serum IgE concentration. The pDC1, pDC2, and basophil subsets demonstrated a similar magnitude of increase in FcepsilonRI expression relative to changes in serum IgE. CONCLUSIONS: FcepsilonRI expression is characteristic of both the DC1 and DC2 subsets. Furthermore, FcepsilonRI expression by these cells is highly correlated to serum IgE and to basophil FcepsilonRI expression and is greater in subjects with allergic asthma. These data support the concept that novel therapeutic approaches directly targeted at FcepsilonRI expression would affect both the sensitization and the effector phases of the allergen-specific immune response.     

Defects in CD54 and CD86 up-regulation by plasmacytoid dendritic cells during pregnancy

Physiological modulation of the immune system is required for foetal tolerance during pregnancy. However, this immune regulation might lead to impaired self-defence against pathogens. Indeed, pregnant women are more susceptible to newly encountered viruses comparing to non-pregnant women, as exemplified by the prevalence of severe complications in pregnant women infected with the pandemic influenza virus in 2009. Plasmacytoid dendritic cells (pDCs) are specialized dendritic cells that recognise viral antigens and initiate both innate and adaptive immune responses. We therefore sought to determine whether the number and/or the functions of peripheral blood pDCs are regulated during pregnancy. pDC maturation and interferon (IFN)-α production were analysed in response to Toll-like receptor (TLR) stimulation of peripheral blood mononuclear cells from pregnant and non-pregnant women. Our results reveal that pDC frequency is slightly decreased, while the IFN-α production in response to TLR stimulation increases during pregnancy. Interestingly, the up-regulation of the co-stimulatory receptors CD54 (ICAM1) and CD86 is significantly decreased in pDCs from pregnant women as compared to controls, suggesting a possible impact on T-cell responses. In conclusion, we propose that the modulation of CD54 and CD86 expression on peripheral blood pDCs during pregnancy might decrease the initiation of adaptive antiviral immune responses.     

Replication-independent activation of human plasmacytoid dendritic cells by the paramyxovirus SV5 Requires TLR7 and autophagy pathways

The paramyxovirus Simian Virus 5 (SV5) is a poor inducer of interferon (IFN) secretion in all cell types tested so far, including primary epithelial cells and primary human myeloid dendritic cells. SV5 is hypothesized to limit induction of antiviral responses through control of viral gene expression and production of the V protein antagonist. Plasmacytoid dendritic cells (pDCs) are known to uniquely express toll-like receptor (TLR)-7 and are a main producer of IFN-alpha among peripheral blood mononuclear cells in response to many viruses. Here, we tested whether SV5 would remain a poor inducer of IFN in primary human pDCs. The efficiency of SV5 infection of pDCs could be increased by an increasing multiplicity of infection. pDCs infected by both live and UV-inactivated SV5 induced large amounts of IFN-alpha secretion and resulted in upregulation of maturation markers CD80 and CD86. However, IL-6 secretion was not induced by SV5 infection. When TLR7 signaling was inhibited, SV5 induced less IFN secretion and CD80 expression, and there was a corresponding increase in number of infected cells. Similar effects were seen with inhibitors of cellular autophagy pathways, suggesting that the SV5 activation of pDC requires access to the cytoplasm and autophagic sampling of cytoplasmic contents. These results have implications for control of SV5 infections in vivo and for development of SV5 as a vaccine vector.     

Expression of the high-affinity IgE receptor on peripheral blood dendritic cells: differential binding of IgE in atopic asthma

BACKGROUND: Dendritic cells can express the high-affinity IgE receptor (FcepsilonRI), which, in the presence of specific IgE, facilitates the uptake of allergen, leading to increased activation of allergen-specific T cells. FcepsilonRI expression by dendritic cells is higher in the airways of atopic asthmatic subjects than in those of healthy, nonatopic control subjects. OBJECTIVE: The aims of this study were to determine whether a similar difference in FcepsilonRI expression occurs between dendritic cells in the peripheral blood of atopic asthmatic subjects and healthy individuals and also whether an altered ability of FcepsilonRI(+) peripheral blood dendritic cells to bind IgE accompanies the atopic asthmatic state. METHODS: Flow cytometry was used to analyze the surface expression of FcepsilonRI and exogenously bound IgE on dendritic cells identified as lineage negative (CD3, CD14, CD16, CD19, and CD56) and HLA-DR bright. RESULTS: The total expression of FcepsilonRI on the surface of dendritic cells from healthy and asthmatic subjects was not significantly different. However, in vivo, dendritic cells from atopic asthmatic subjects had higher levels of receptor occupancy by IgE and bound exogenous IgE in vitro more efficiently than dendritic cells from healthy subjects. CONCLUSION: The similar levels of expression of FcepsilonRI on peripheral blood dendritic cells from healthy and asthmatic subjects suggest that the local environment in the airway is responsible for the upregulation of surface FcepsilonRI on airway dendritic cells in asthma. The results also suggest that the functional ability of FcepsilonRI to bind IgE is differentially controlled in the atopic state.     

Plasmacytoid dendritic cell ablation impacts early interferon responses and antiviral NK and CD8(+) T cell accrual

Plasmacytoid dendritic cells (pDCs) mediate type I interferon (IFN-I) responses to viruses that are recognized through the Toll-like receptor 7 (TLR7) or TLR9 signaling pathway. However, it is unclear how pDCs regulate the antiviral responses via innate and adaptive immune cells. We generated diphtheria toxin receptor transgenic mice to selectively deplete pDCs by administration of diphtheria toxin. pDC-depleted mice were challenged with viruses known to activate pDCs. In murine cytomegalovirus (MCMV) infection, pDC depletion reduced early IFN-I production and augmented viral burden facilitating the expansion of natural killer (NK) cells expressing the MCMV-specific receptor Ly49H. During vesicular stomatitis virus (VSV) infection, pDC depletion enhanced early viral replication and impaired the survival and accumulation of virus-specific cytotoxic T lymphocytes. We conclude that pDCs mediate early antiviral IFN-I responses and influence the accrual of virus-specific NK or CD8(+) T?cells in?a virus-dependent manner.     

Decreased interferon-alpha production in HIV-infected patients correlates with numerical and functional deficiencies in circulating type 2 dendritic cell precursors.

Peripheral blood mononuclear cells from patients with human immunodeficiency virus (HIV) infection exhibit a progressively marked decrease in the production of virus-induced interferon (IFN)-alpha, a finding that correlates with and is highly predictive of disease progression and opportunistic infections. The major IFN-alpha producing population has recently been defined as the precursor to type 2 dendritic cells (pDC2) or plasmacytoid DC (pDC). Using four-color flow cytometry, we have enumerated the pDC2 vs non-IFN-alpha producing myeloid DC1 in peripheral blood from HIV-infected patients and healthy controls and related these values to CD4 cell numbers, viral load, and functional activity. The patients had reductions in the numbers of both pDC2 (lin-/HLA-DR+/CD123(bright)) and DC1 (lin1-/HLA-DR+/CD123(dim)/CD11c+), both at an absolute level and as a percentage of cells. The decreases were most evident in patients with decreased CD4 levels. Viral load correlated with the functional frequency of the IFN producing cells but not with absolute pDC2 levels. Using intracellular flow cytometric analysis for IFN-alpha, the patients were demonstrated to have fewer pDC2, as well as a lower percentage of responding cells among those remaining. We conclude that deficient production of IFN-alpha by pDC2 from HIV-infected patients results from both selective loss of these cells and their qualitative dysfunction. Given the central role of DC, and in particular, DC2, in linking innate and adaptive immune responses, these qualitative and quantitative changes in pDC2 are likely to be key contributors to HIV pathogenesis.     

Common variable immunodeficiency revisited: normal generation of naturally occurring dendritic cells that respond to Toll‐like receptors 7 and 9

Patients with common variable immunodeficiency (CVID) have reduced numbers and frequencies of dendritic cells (DCs) in blood, and there is also evidence for defective activation through Toll‐like receptors (TLRs). Collectively, these observations may point to a primary defect in the generation of functional DCs. Here, we measured frequencies of plasmacytoid DCs (pDCs) and myeloid DCs (mDCs) in peripheral blood of 26 CVID patients and 16 healthy controls. The results show that the patients have reduced absolute counts of both subsets. However, the decreased numbers in peripheral blood were not reflected in reduced frequencies of CD34⁺ pDC progenitors in the bone marrow. Moreover, studies at the single cell level showed that DCs from CVID patients and healthy controls produced similar amounts of interferon‐α or interleukin‐12 and expressed similar levels of activation markers in response to human cytomegalovirus and ligands for TLR‐7 and TLR‐9. The study represents the most thorough functional characterization to date, and the first to assess bone marrow progenitor output, of naturally occurring DCs in CVID. In conclusion, it seems unlikely that CVID is secondary to insufficient production of naturally occurring DCs or a defect in their signalling through TLR‐7 or TLR‐9.     

Disparate effects of acute and chronic infection with SIVmac239 or SHIV-89.6P on macaque plasmacytoid dendritic cells

Blood plasmacytoid dendritic cells (pDCs) contribute to both innate and adaptive immune responses by secreting high levels of IFN-alpha following acute bacterial and viral infections and indirectly by augmenting cell-mediated immunity. Cross-sectional studies have shown that the number of circulating pDCs in HIV patients, compared to that in uninfected individuals, is reduced. However, since the time of infection is usually unknown in HIV-infected patients, pDC-virus interactions that occur immediately after virus exposure are poorly understood. The current study investigated pDC dynamics during acute and chronic infections of macaques with either SIVmac239 or the pathogenic SIV-HIV chimera, SHIV-89.6P, as models for HIV infection. In three rhesus and three pig-tailed macaques infected intravenously with SIVmac239, the percentages of pDCs in blood declined 2- to 6-fold during the first 6 weeks after infection and remained depressed throughout the disease course. Surprisingly, no consistent, comparable decline in peripheral blood pDCs was observed in six macaques infected with SHIV-89.6P. In this latter group, percentages of pDCs did not correlate with CD4(+) T cells, but there was an inverse relationship with viral load. In addition, when compared to na?ve controls, the percentages of pDCs were reduced in spleens and peripheral lymph nodes of SIVmac239- but not SHIV-89.6P-infected animals that had progressed to AIDS. Proviral DNA was detected during the acute phase in pDCs isolated from macaques infected with either virus. These results imply that, even though macaque pDCs can be infected by both SIVmac239 and SHIV-89.6P, the subsequent effects on in vivo pathogenesis differ. The underlying mechanism(s) for these differences is unclear, but the selection of SIV or SHIV as a challenge virus might influence the outcome of some studies, such as those evaluating vaccines or the therapeutic efficacy of drugs.     

CD49d/CD29‐integrin controls the accumulation of plasmacytoid dendritic cells into the CNS during neuroinflammation

Plasmacytoid dendritic cells (pDCs) are found in the CNS during neuroinflammation and have been reported to exert regulatory functions in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). However, the mechanisms of entry of pDCs into the CNS as well as their phenotype and innate functional properties, once recruited into the CNS, have not been thoroughly examined. Herein, we show that pDCs rapidly accumulate into the brain and spinal cord during the acute phase of EAE, and maintain the expression of numerous phenotypic markers typical of peripheral pDCs. Functionally, CNS‐pDCs constitutively expressed IRF7 and were able to rapidly produce type I IFNs and IL‐12p40 upon ex vivo TLR‐9 stimulation. Using adoptive transfer experiments, we provide evidence that CNS‐pDC are recruited from the blood and accumulate into the CNS during the acute phase of EAE. Accumulation of pDCs into the CNS was strongly inhibited in the absence of CD29, but not CD18, suggesting a major role for ß1 but not ß2 integrins. Indeed, blocking the CD49d α4‐integrins during acute EAE drastically diminished CNS‐pDC numbers. Together, our results demonstrate that circulating pDCs are actively recruited into the CNS during acute EAE through a mechanism largely dependent on CD49d/CD29‐integrins.     

Analysis of HLDA9 mAbs on plasmacytoid dendritic cells

Dendritic cells are a heterogeneous population of bone marrow derived leucocytes that play a crucial role in both pathogen recognition and the initiation of primary T cell immune responses. Plasmacytoid dendritic cells (pDCs), also known as natural interferon-producing cells, comprise one of two major human dendritic cell subsets that strongly influences immune balance. pDCs remain a poorly characterized subset. Several studies have suggested the existence of a close phenotypic and functional relationship between B-cells and pDCs. The surface reactivity of a panel of 96 monoclonal antibodies submitted to the ninth Human Leukocyte Differentiation Antigens workshop (HLDA9) B cell section was analyzed using pDCs as target cells. The results showed that eight of the mAbs reacted positively on pDCs: CD86, CD229, CD319, CD305, CD184, CD84, CD85g and Fc?RIa, confirming previously published reports. Interestingly, this study also revealed the expression of eight surface molecules not previously described on pDCs, including CD352(NTBA), CD272(BTLA), CD357(GITR), CD48, CD270(HVEM), Galectin-3, CD148, and CD361. The present report summarizes the expression of these molecules on freshly isolated pDCs. Significantly, we have identified several new molecules expressed by these intriguing cells, ones we believe will open new avenues for the study of pDC functionality and their role in human health and diseases.     

Unraveling the functions of plasmacytoid dendritic cells during viral infections,autoimmunity, and tolerance

Summary: Plasmacytoid dendritic cells (pDCs) are bone marrow-derived cells that secrete large amounts of type I interferon (IFN) in response to viruses. Type I IFNs are pleiotropic cytokines with antiviral activity that also enhance innate and adaptive immune responses. Viruses trigger activation of pDCs and type I IFN responses mainly through the Toll-like receptor pathway. However, a variety of activating and inhibitory pDC receptors fine tune the amplitude of type I IFN responses. Chronic activation and secretion of type I IFN in the absence of infection can promote autoimmune diseases. Furthermore, while activated pDCs promote immunity and autoimmunity, resting or alternatively activated pDCs may be tolerogenic. The various roles of pDCs have been extensively studied in vitro and in vivo with depleting antibodies. However, depleting antibodies cross-react with other cell types that are critical for eliciting protective immunity, potentially yielding ambiguous phenotypes. Here we discuss new approaches to assess pDC functions in vivo and provide preliminary data on their potential roles during viral infections. Such approaches would also prove useful in the more specific evaluation of how pDCs mediate tolerance and autoimmunity. Finally, we discuss the emergent role of pDCs and one of their receptors, tetherin, in human immunodeficiency virus pathogenesis.     

Individual plasmacytoid dendritic cells are major contributors to the production of multiple innate cytokines in an organ-specific manner during viral infection

Plasmacytoid dendritic cells (pDCs) are an important source of IFN-alpha/beta in response to a variety of viruses in vivo, including murine cytomegalovirus (MCMV). However, the respective contributions of various infected organs, and within these of pDCs, conventional dendritic cells and other cells, to the systemic production of IFN-alpha/beta or other innate cytokines during viral infections in vivo is largely unknown. Whether a specialization of pDC subsets in the production of different patterns of innate cytokines exists in vivo in response to a viral infection has not been investigated. Here, by analyzing for the first time directly ex vivo, at the single-cell level, the simultaneous production of up to three cytokines in pDCs isolated from MCMV-infected mice, we show that (i) pDCs are the quasi-exclusive source of IFN-alpha/beta, IL-12 and tumor necrosis factor (TNF)-alpha, early during MCMV infection, in two immunocompetent mouse lines and with two viral strains, (ii) pDC activation for IFN-alpha/beta production is organ specific and (iii) a significant proportion of pDCs simultaneously produce IFN-alpha/beta, TNF-alpha and IL-12, although TNF-alpha and IFN-alpha/beta appear more often co-expressed with one another than each of them with IL-12. Altogether, these results show a broad and non-redundant role of pDCs in early innate detection of, and defense against, viral infection. The data also show differences in the responsiveness of pDCs from different tissues and suggest distinct molecular requirements for pDC production of various cytokines. These observations must be taken into account when designing new antiviral vaccination strategies aimed at harnessing pDC responses.     

Dendritic cells in atopic dermatitis: expression of FcepsilonRI on two distinct inflammation-associated subsets

BACKGROUND: Dendritic cells (DCs) represent a major portion within the infiltrate of atopic dermatitis (AD) lesions. As antigen-presenting cells they have the ability to regulate both the quantity and quality of T-cell responses and, thus, are likely to play a key role in the pathogenesis of T-cell-dominated skin diseases such as AD. Thus we sought to identify the DC repertoire occurring in AD patients. METHODS: For this purpose, we phenotypically analyzed various defined DC subsets of AD patients and healthy controls in skin biopsies and peripheral blood by immunofluorescence staining. RESULTS: In AD lesions, two inflammation-associated DC subsets with varying expression of costimulatory molecules occurred besides epidermal Langerhans cells (LCs) and dermal myeloid DCs (dmDCs) indigenously residing in normal skin: (1) CD1a+/CD1c+/FcepsilonRI+/IgE+/CD207- myeloid DCs (mDCs) in the epidermis and dermis and (2) CD123+/BDCA-2+/CD45RA+/CD68+ plasmacytoid DCs (pDCs) in the dermis. In the peripheral blood of the patients, these cells exhibited an immature phenotype. Interestingly, we found FcepsilonRI and cell-bound IgE to be expressed not only on myeloid, but also on plasmacytoid DCs from both the skin and peripheral blood of AD patients. CONCLUSIONS: It is tempting to speculate that the disease-regulating role of inflammatory DCs in AD is influenced by both FcepsilonRI occupancy and their degree of maturity.     

Alloantigen-presenting plasmacytoid dendritic cells mediate tolerance to vascularized grafts

The induction of alloantigen-specific unresponsiveness remains an elusive goal in organ transplantation. Here we identify plasmacytoid dendritic cells (pDCs) as phagocytic antigen-presenting cells essential for tolerance to vascularized cardiac allografts. Tolerizing pDCs acquired alloantigen in the allograft and then moved through the blood to home to peripheral lymph nodes. In the lymph node, alloantigen-presenting pDCs induced the generation of CCR4+ CD4+ CD25+ Foxp3+ regulatory T cells (Treg cells). Depletion of pDCs or prevention of pDC lymph node homing inhibited peripheral Treg cell development and tolerance induction, whereas adoptive transfer of tolerized pDCs induced Treg cell development and prolonged graft survival. Thus, alloantigen-presenting pDCs home to the lymph nodes in tolerogenic conditions, where they mediate alloantigen-specific Treg cell development and allograft tolerance.     

Plasmacytoid dendritic cells act as the most competent cell type in linking antiviral innate and adaptive immune responses

Appropriate in vivo control of plasmacytoid dendritic cell （pDC） recruitment and activation is a fundamental requirement for defense against viral infection. During this process, a pivotal event that influences the outcome of viral infection is the production of high levels of type I interferon by pDCs. In particular, recent research findings showed that pDCs not only shape the nature of innate resistance, but are also responsible for the successful transition from innate to adaptive immunity for viral resistance. In addition, pDCs can differentiate into antigen presenting cells that may regulate tolerance to a given pathogen. Importantly, in a series of recent clinical studies, pDCs appeared to be defective in number and function in conditions of chronic viral diseases such as infected with HIV-1, HBV or HCV. pDC-associated clinical antiviral therapy is also emerging. This review describes research findings exatnining the functional and antiviral properties of in vivo pDC plasticity. Cellular ＆ Molecular Immunology. 2005;2（6）：411- 417.     

TLR9/TLR7-triggered downregulation of BDCA2 expression on human plasmacytoid dendritic cells from healthy individuals and lupus patients

Plasmacytoid dendritic cells (pDCs) can produce a large amount of interferon-alpha (IFN-alpha) upon exposure to TLR9 or TLR7 agonists. Human pDCs have been shown to play an important role in the pathogenesis of systemic lupus erythematosus (SLE) through increased production of IFN-alpha. So, how to negatively regulate activation of pDCs and how to evaluate the activation of pDC in SLE patients attract much attention. BDCA2 is selectively expressed on human pDCs, acting as a hallmark of human pDCs. In this study, we showed that BDCA2 expression on pDCs decreased along maturation of pDCs, and TLR7 or TLR9 agonists could further significantly downregulate pDCs to express BDCA2, suggesting that the activated pDCs exhibit decreased expression of BDCA2. Functionally, BDCA2 ligation significantly inhibited upregulation of CD40, CD86 and CCR7 expression, IFN-alpha, IFN-beta and IL-6 production by pDCs stimulated with CpG ODN. Moreover, BDCA2 ligation suppressed CpG ODN-activated pDCs to mediate Th1 response, including T cell proliferation, IFN-gamma production, and CD4(+)CCR5(+)Th1 development, confirming that BDCA2 is a negative regulator of TLR9-dependent activation of human pDCs. BDCA2 expression on pDCs from SLE patients decreased significantly but IFN-alpha production of these patients increased markedly as compared to that from healthy donors. Therefore, these results suggest that downregulation of BDCA2 expression on pDCs may reflect the activation of pDCs accumulated in SLE patients, and may be one marker for indication of the disease activity of SLE patients.     

Comparative analysis of nasal and oral mucosa dendritic cells

BACKGROUND: Mucosal dendritic cells (DC) play a crucial role in tolerance induction as seen in mucosal immunotherapy of atopic diseases. Nevertheless little is known about the phenotypical differences of oral and nasal mucosal DC (nmDC). Recently, we could show that oral mucosal myeloid CD1a(+) DC (omDC) differ from their skin counterparts especially by the expression of high affinity receptor for immunoglobulin E (IgE; FcepsilonRI). However, expression pattern of FcepsilonRI and phenotypical characteristics of CD1a(+) nmDC have not been elucidated in detailed yet. METHODS: We performed detailed phenotypical comparison of nmDC and omDC of atopic and nonatopic individuals. RESULTS: As reported for omDC, FcepsilonRI on nmDC of atopic donors was elevated and mostly occupied by IgE while FcepsilonRI was present only in low amounts on nmDC of nonatopic donors. Nevertheless, the highest FcepsilonRI expression has been observed on omDC. Furthermore, significant amounts of costimulatory molecules CD40, CD80 and CD86 could be detected on nmDC that expressed more CD80 compared with omDC. Moreover, nmDC displayed less major histocompatability complex (MHC) class I and II molecules than omDC. In addition, nmDC expressed more C-type lectins CD205, CD206 as well as myeloid marker CD11b while omDC displayed increased expression of CD207 and lipopolysaccharide (LPS) receptor CD14. CONCLUSION: Together these data imply that nmDC phenotypical differ from omDC which might result in diverse functional properties and might be of relevance for selecting routes for immunotherapy of atopic diseases. Moreover these data provide a basis for further studies investigating immunological mechanisms underlying mucosal immunotherapy.