Transcriptional patterns associated with BDCA3 expression on BDCA1+ myeloid dendritic cells.

Myeloid dendritic cells, including BDCA3hi DCs and BDCA1+ DCs (hereafter dubbed DC1 and DC2 for clarity), play a pivotal role in the induction and regulation of immune responses. Interestingly, a fraction of DC2 also express low to intermediate levels of BDCA3. It is unknown whether BDCA3+ DC2 also share other traits with DC1 that are absent in BDCA3- DC2 and/or whether BDCA3 expression renders DC2 functionally distinct from their BDCA3-lacking counterparts. Here, we used expression analysis on a predefined set of immunology-related genes to determine divergence between BDCA3-positive and BDCA3-negative DC2 and their relation to bona fide BDCA3hi DC1. Results showed that mRNA fingerprints of BDCA3+ DC2 and BDCA3- DC2 are very similar, and clearly distinct from that of DC1. Differences in mRNA expression, however, were observed between BDCA3+ DC2 and BDCA3- DC2 that pointed toward a more activated status of BDCA3+ DC2. In line with this, higher steady state maturation marker expression and TLR-induced maturation marker expression and inflammatory cytokine production by BDCA3+ DC2 were observed. This dataset provides insight into the relationship between myeloid DC populations and contributes to further understanding of DC immunobiology.

The Effect of Chronic Hepatitis B Virus Infection on BDCA3+ Dendritic Cell Frequency and Function.

Chronic hepatitis B virus (HBV) infection results from inadequate HBV-specific immunity. BDCA3+ dendritic cells (DCs) are professional antigen presenting cells considered to be important for antiviral responses because of specific characteristics, including high interferon-λ production. BDCA3+ DCs may thus also have a role in the immune response against HBV, and immunotherapeutic strategies aiming to activate DCs, including BDCA3+ DCs, in patient livers may represent an interesting treatment option for chronic HBV. However, neither the effect of chronic hepatitis B (CHB) infection on the frequency and function of BDCA3+ DCs in liver and blood, nor the effect of the viral surface protein (HBsAg) that is abundantly present in blood of infected individuals are known. Here, we provide an overview of BDCA3+ DC frequency and functional capacity in CHB patients. We find that intrahepatic BDCA3+ DC numbers are increased in CHB patients. BDCA3+ DCs from patient blood are not more mature at steady state, but display an impaired capacity to mature and to produce interferon-λ upon polyI:C stimulation. Furthermore, in vitro experiments exposing blood and intrahepatic BDCA3+ DCs to the viral envelope protein HBsAg demonstrate that HBsAg does not directly induce phenotypical maturation of BDCA3+ DCs, but may reduce IFN-λ production via an indirect unknown mechanism. These results suggest that BDCA3+ DCs are available in the blood and on site in HBV infected livers, but measures may need to be taken to revive their function for DC-targeted therapy.

Hepatitis B Virus Surface Antigen Activates Myeloid Dendritic Cells via a Soluble CD14-Dependent Mechanism.

UNLABELLED: Hepatitis B virus (HBV) infection can cause chronic liver disease, which is associated with increased risk of liver cirrhosis, liver failure, and liver cancer. Clearance of HBV infection requires effective HBV-specific immunity; however, the immunological mechanisms that determine the development of effective HBV-specific immunity are poorly understood. Dendritic cells (DC) play a pivotal role in the regulation of antiviral immunity. Here, we investigated the interaction between HBV surface antigen (HBsAg), the main envelope glycoprotein of HBV, and BDCA1(+) myeloid dendritic cells (mDC). Exposure of peripheral blood-derived BDCA1(+) mDC to HBsAg resulted in strong DC maturation, cytokine production, and enhanced capacity to activate antigen-specific cytotoxic T cells (CTLs). By using neutralizing antibodies, crucial roles for CD14 and Toll-like receptor 4 (TLR4) in HBsAg-mediated BDCA1(+) mDC maturation were identified. Concordantly, HBsAg-mediated DC maturation required fetal calf serum (FCS) or human plasma, naturally containing soluble CD14 (sCD14). Intriguingly, HBsAg-induced DC maturation was significantly reduced in umbilical cord blood plasma, which contained less sCD14 than adult plasma, indicating that sCD14 is an important host factor for recognition of HBsAg by DC and subsequent DC activation. A direct interaction between sCD14 and HBsAg was demonstrated by using enzyme-linked immunosorbent assay (ELISA). Moreover, sCD14-HBsAg complexes were detected both in vitro and in sera of HBV-infected patients. The abundance of sCD14-HBsAg complexes varied between chronic HBV disease stages and correlated with activation of BDCA1(+) mDC in vivo We conclude that HBsAg activates BDCA1(+) DC via an sCD14-dependent mechanism. These findings provide important novel insights into the initiation of HBV-specific immunity and facilitate development of effective immunotherapeutic interventions for HBV. IMPORTANCE: Hepatitis B virus (HBV) infection is a significant health problem, as it causes progressive liver injury and liver cancer in patients with chronic HBV infection, which affects approximately 250 million individuals worldwide. Some of the infected adults and the majority of neonates fail to mount an effective immune response and consequently develop chronic infection. The viral and host factors involved in the initiation of effective HBV-specific immune responses remain poorly understood. Here we identified CD14 and TLR4 as receptors for HBsAg, the main HBV envelope antigen. HBsAg induced strong maturation of dendritic cells (DC), which have a central role in regulation of virus-specific immunity. These results provide essential novel insights into the mechanisms underlying the initiation of HBV-specific immunity. Intriguingly, since neonates have naturally low sCD14, the finding that serum-derived sCD14 is a crucial host factor for recognition of HBsAg by DC may have implications for immunity of neonates to HBV infection.

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

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

BDCA3(+)CLEC9A(+) human dendritic cell function and development.

Dendritic cells (DC) are the most potent antigen presenting cells (APC). They comprise a family of different subsets and play an essential role in the induction and regulation of immune responses. Recently, gene expression profiling identified BDCA3(+)CLEC9A(+) DC as a separate human DC subset. This subset was identified in blood, where they represent the smallest population of human DC, as well as in lymphoid and peripheral tissues. This review summarizes the phenotypic, functional and developmental characteristics of BDCA3(+)CLEC9A(+) DC in relation to their mouse equivalents CD8α(+) DC and CD103(+) DC and other human DC subsets. Apart from being potent antigen presenting cells, their specialized functional capacities compared to other human DC subsets, indicate that these BDCA3(+)CLEC9A(+) DC are of major importance in the induction of anti-viral and anti-tumor immunity. Further characterization of their functional properties, developmental pathways and underlying molecular mechanisms may identify target molecules to fully exploit the immune modulatory function of BDCA3(+)CLEC9A(+) DC and potential use of these cells in immunotherapy.

Understanding MHC class I presentation of viral antigens by human dendritic cells as a basis for rational design of therapeutic vaccines.

Effective viral clearance requires the induction of virus-specific CD8(+) cytotoxic T lymphocytes (CTL). Since dendritic cells (DC) have a central role in initiating and shaping virus-specific CTL responses, it is important to understand how DC initiate virus-specific CTL responses. Some viruses can directly infect DC, which theoretically allow direct presentation of viral antigens to CTL, but many viruses target other cells than DC and thus the host depends on the cross-presentation of viral antigens by DC to activate virus-specific CTL. Research in mouse models has highly enhanced our understanding of the mechanisms underlying cross-presentation and the dendritic cells (DC) subsets involved, however, these results cannot be readily translated toward the role of human DC in MHC class I-antigen presentation of human viruses. Here, we summarize the insights gained in the past 20 years on MHC class I presentation of viral antigen by human DC and add to the current debate on the capacities of different human DC subsets herein. Furthermore, possible sources of viral antigens and essential DC characteristics for effective induction of virus-specific CTL are evaluated. We conclude that cross-presentation is not only an efficient mechanism exploited by DC to initiate immunity to viruses that do not infect DC but also to viruses that do infect DC, because cross-presentation has many conceptual advantages and bypasses direct immune modulatory effects of the virus on its infected target cells. Since knowledge on the mechanism of viral antigen presentation and the preferred DC subsets is crucial for rational vaccine design, the obtained insights are very instrumental for the development of effective anti-viral immunotherapy.