Skin-resident Dendritic Cells Research Articles

Evaluation of cutaneous immune response in a controlled human in vivo model of mosquito bites

Mosquito-borne viruses are a growing global threat. Initial viral inoculation occurs in the skin via the mosquito ‘bite’, eliciting immune responses that shape the establishment of infection and pathogenesis. Here we assess the cutaneous innate and adaptive immune responses to controlled Aedes aegypti feedings in humans living in Aedes-endemic areas. In this single-arm, cross-sectional interventional study (trial registration #NCT04350905), we enroll 30 healthy adult participants aged 18 to 45 years of age from Cambodia between October 2020 and January 2021. We perform 3-mm skin biopsies at baseline as well as 30 min, 4 h, and 48 h after a controlled feeding by uninfected Aedes aegypti mosquitos. The primary endpoints are measurement of changes in early and late innate responses in bitten vs unbitten skin by gene expression profiling, immunophenotyping, and cytokine profiling. The results reveal induction of neutrophil degranulation and recruitment of skin-resident dendritic cells and M2 macrophages. As the immune reaction progresses T cell priming and regulatory pathways are upregulated along with a shift to Th2-driven responses and CD8+ T cell activation. Stimulation of participants’ bitten skin cells with Aedes aegypti salivary gland extract results in reduced pro-inflammatory cytokine production. These results identify key immune genes, cell types, and pathways in the human response to mosquito bites and can be leveraged to inform and develop novel therapeutics and vector-targeted vaccine candidates to interfere with vector-mediated disease.

Clonal Hematopoiesis Evolves into Dendritic Cell Neoplasm through the Acquisition of UV Mutations in the Skin

Background Expansion of blood cells derived from a single mutated blood stem cell - clonal hematopoiesis - often precedes leukemic transformation. However, the circumstances that promote or prevent transformation are poorly understood. Here, we investigate the evolution of clonal hematopoiesis into blastic plasmacytoid dendritic cell neoplasm (BPDCN), an aggressive blood cancer that often presents with malignant cells isolated to the skin (referred to as skin-only disease). We gain a better understanding of the influence of site-specific selective pressures and the role of TET2 mutations, which may inform interventions to prevent disease progression. Methods We analyzed bone marrow and skin from 13 BPDCN patients using targeted, exome, and whole-genome sequencing, with germline controls. We performed multimodal single-cell sequencing (10x 3' v3.1 single-cell RNA-seq with genotyping of expressed variants) on bone marrow from 6 healthy donors (20,411 cells), 5 skin-only BPDCN patients with malignant cells in the skin but not conventionally detectable in marrow (36,018 cells), and 6 BPDCN patients with overt bone marrow involvement (30,582 cells). For functional studies, we targeted Tet2 using CRISPR/Cas9 in mouse blood progenitor cells immortalized by an estrogen receptor hormone binding domain fused to HoxB8, which undergo synchronized differentiation to myeloid and dendritic cells upon estrogen withdrawal. We used CellRanger, Seurat, RandomForest machine learning, and the R tidyverse package collection for bioinformatics. Results Genetics allowed us to reconstruct the evolution of BPDCN across bone marrow and skin (representative patient, panel a). Invariably, we detected founder mutations in marrow, including TET2 inactivation in 9/13 patients (two mutations, likely bi-allelic, in 5/9). Single-cell sequencing of marrow from 5 skin-only patients showed that pre-malignant mutant stem cells contributed to multiple lineages, and that TET2 mutated clones were myeloerythroid-biased. We generated a gene expression signature to identify rare malignant BPDCN cells in marrow of skin-only patients. In 3/5 patients, we identified rare cells (0.03-0.19%) with the transcriptional profile of fully transformed cells, and concurrent single-cell genetic analysis suggested that these cells originated from the skin tumor (panel b). To assess whether disseminated skin tumor cells establish bone marrow disease, we quantified mutational signatures: skin tumors and malignant BPDCN marrow cells in 5/5 patients harbored UV-induced mutations, likely acquired in a single skin-resident plasmacytoid dendritic cell (pDC) during malignant transformation. Finally, we edited Tet2 in mouse myelo-dendritic progenitors and tested the response to UV irradiation in vitro. In addition to observing that Tet2 inactivation increases dendritic cell commitment, the more striking result was a survival advantage of Tet2-mutated cells in the presence of UV (n = 6, fold change 1.803, P = 0.0012). These results highlight an unexpected function of TET2 that may predispose mutated pDCs to survive UV-induced DNA damage in the skin, leading to disease progression. Conclusion The complex evolution of BPDCN traverses multiple anatomic sites prior to full transformation. This allows for accurate sequencing of oncogenic events. Disease evolution frequently starts with acquisition of a leukemia-associated mutation in a multipotent blood stem cell, leading to clonal expansion. In the most frequently observed scenario, bi-allelic TET2 inactivation promotes myeloerythroid bias, increases dendritic cell commitment, and confers resistance to UV-mediated cell death. Subsequent evolution of skin-resident mutant pDCs leads to a high burden of UV-induced mutations and ultimate transformation. Dissemination of malignant cells, which can be detected by integrating single-cell gene expression and mutation profiles, initiates "retrograde" bone marrow disease. Our discovery that multiple anatomic sites - and local selective pressures - play a role in disease progression provides a template to study multi-site evolution of clonal hematopoiesis. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

A Novel C-Type Lectin Receptor-Targeted α-Synuclein-Based Parkinson Vaccine Induces Potent Immune Responses and Therapeutic Efficacy in Mice.

The progressive accumulation of misfolded α-synuclein (α-syn) in the brain is widely considered to be causal for the debilitating clinical manifestations of synucleinopathies including, most notably, Parkinson’s disease (PD). Immunotherapies, both active and passive, against α-syn have been developed and are promising novel treatment strategies for such disorders. To increase the potency and specificity of PD vaccination, we created the ‘Win the Skin Immune System Trick’ (WISIT) vaccine platform designed to target skin-resident dendritic cells, inducing superior B and T cell responses. Of the six tested WISIT candidates, all elicited higher immune responses compared to conventional, aluminum adjuvanted peptide-carrier conjugate PD vaccines, in BALB/c mice. WISIT-induced antibodies displayed higher selectivity for α-syn aggregates than those induced by conventional vaccines. Additionally, antibodies induced by two selected candidates were shown to inhibit α-syn aggregation in a dose-dependent manner in vitro. To determine if α-syn fibril formation could also be inhibited in vivo, WISIT candidate type 1 (CW-type 1) was tested in an established synucleinopathy seeding model and demonstrated reduced propagation of synucleinopathy in vivo. Our studies provide proof-of-concept for the efficacy of the WISIT vaccine technology platform and support further preclinical and clinical development of this vaccine candidate.

Science, Medicine, and the Anesthesiologist

Science, Medicine, and the Anesthesiologist

Usutu Virus escapes langerin-induced restriction to productively infect human Langerhans cells, unlike West Nile virus

ABSTRACT Usutu virus (USUV) and West Nile virus (WNV) are phylogenetically close emerging arboviruses and constitute a global public health threat. Since USUV and WNV are transmitted by mosquitoes, the first immune cells they encounter are skin-resident dendritic cells, the most peripheral outpost of immune defense. This unique network is composed of Langerhans cells (LCs) and dermal DCs, which reside in the epidermis and the dermis, respectively. Using human skin explants, we show that while both viruses can replicate in keratinocytes, they can also infect resident DCs with distinct tropism: WNV preferentially infects DCs in the dermis, whereas USUV has a greater propensity to infect LCs. Using both purified human epidermal LCs (eLCs) and monocyte derived LCs (MoLCs), we confirm that LCs sustain a faster and more efficient replication of USUV than WNV and that this correlates with a more intense innate immune response to USUV compared with WNV. Next, we show that ectopic expression of the LC-specific C-type lectin receptor (CLR), langerin, in HEK293T cells allows WNV and USUV to bind and enter, but supports the subsequent replication of USUV only. Conversely, blocking or silencing langerin in MoLCs or eLCs made them resistant to USUV infection, thus demonstrating that USUV uses langerin to enter and replicate in LCs. Altogether, our results demonstrate that LCs constitute privileged target cells for USUV in human skin, because langerin favours its entry and replication. Intriguingly, this suggests that USUV efficiently escapes the antiviral functions of langerin, which normally safeguards LCs from most viral infections.

Skin-resident dendritic cells mediate postoperative pain via CCR4 on sensory neurons

Inflammatory pain, such as hypersensitivity resulting from surgical tissue injury, occurs as a result of interactions between the immune and nervous systems with the orchestrated recruitment and activation of tissue-resident and circulating immune cells to the site of injury. Our previous studies identified a central role for Ly6Clow myeloid cells in the pathogenesis of postoperative pain. We now show that the chemokines CCL17 and CCL22, with their cognate receptor CCR4, are key mediators of this response. Both chemokines are up-regulated early after tissue injury by skin-resident dendritic and Langerhans cells to act on peripheral sensory neurons that express CCR4. CCL22, and to a lesser extent CCL17, elicit acute mechanical and thermal hypersensitivity when administered subcutaneously; this response abrogated by pharmacological blockade or genetic silencing of CCR4. Electrophysiological assessment of dissociated sensory neurons from naïve and postoperative mice showed that CCL22 was able to directly activate neurons and enhance their excitability after injury. These responses were blocked using C 021 and small interfering RNA (siRNA)-targeting CCR4. Finally, our data show that acute postoperative pain is significantly reduced in mice lacking CCR4, wild-type animals treated with CCR4 antagonist/siRNA, as well as transgenic mice depleted of dendritic cells. Together, these results suggest an essential role for the peripheral CCL17/22:CCR4 axis in the genesis of inflammatory pain via direct communication between skin-resident dendritic cells and sensory neurons, opening therapeutic avenues for its control.

Influence of vaccine adjuvant physical properties on antigen processing and presentation in the lymph node

Abstract Despite the efficacy of vaccines, there is a growing need for novel vaccine adjuvants that can drive type-1 immunity, a response governed by antigen presenting cells (APCs). Current methods used to evaluate efficacy of novel adjuvants fail to consider the spatiotemporal events of antigen presentation that dictate the development of cellular immunity. Understanding how adjuvants affect the initial development of type-1 immunity would result in the design of more effective adjuvants. We hypothesized that adjuvants induce unique spatiotemporal patterns within lymph nodes that differ based on the extent to which an adjuvant drains to the LN. These patterns are then responsible for shaping the adaptive immune response. We tested this hypothesis by converting alum, a depot-forming adjuvant that remained at the site of inoculation, into nanoparticles (np) that enter the lymphatics and drained to the LN after vaccination. The size and charge of the np alum was measured through dynamic light scattering and zeta potential. The immunostimulatory effects of both alum and np alum were measured through inflammasome activation of bone marrow-derived macrophages. The two adjuvant formulations were then used to examine the influence of depot-forming alum versus lymph-node (LN) draining alum on skin-resident dendritic cell migration to the LN. The number of migratory DCs in the LN after vaccination was measured by flow cytometry. Next, we made use of live ex vivo lymph node slices to measure chemokine secretion in the LN by ELISA and to observe patterns of DC migration through widefield microscopy. This study illustrates the impact of the physical properties of an adjuvant in shaping the early stages of the adaptive immune response.

ALK3 Is Not Required for the Embryonic Development, Homeostasis, and Repopulation of Epidermal Langerhans Cells in Steady and Inflammatory States

Epidermal Langerhans cells (LCs) are skin-resident dendritic cells, although recently LCs were also classified as specialized macrophages on the basis of their embryonic development shared with tissue-resident macrophages (Doebel et al., 2017; Hoeffel et al., 2012). LCs maintain skin immune surveillance and are involved in skin disorders, including psoriasis (Eidsmo and Martini, 2018; Kaplan, 2017). Adult LCs homeostasis is maintained throughout lifetime self-renewal at steady state, but the impaired LCs could arise by bone marrow–derived precursors at inflammatory state (Ginhoux et al., 2006; Seré et al., 2012).

Chromatin accessibility landscapes of skin cells in systemic sclerosis nominate dendritic cells in disease pathogenesis

Systemic sclerosis (SSc) is a disease at the intersection of autoimmunity and fibrosis. However, the epigenetic regulation and the contributions of diverse cell types to SSc remain unclear. Here we survey, using ATAC-seq, the active DNA regulatory elements of eight types of primary cells in normal skin from healthy controls, as well as clinically affected and unaffected skin from SSc patients. We find that accessible DNA elements in skin-resident dendritic cells (DCs) exhibit the highest enrichment of SSc-associated single-nucleotide polymorphisms (SNPs) and predict the degrees of skin fibrosis in patients. DCs also have the greatest disease-associated changes in chromatin accessibility and the strongest alteration of cell–cell interactions in SSc lesions. Lastly, data from an independent cohort of patients with SSc confirm a significant increase of DCs in lesioned skin. Thus, the DCs epigenome links inherited susceptibility and clinically apparent fibrosis in SSc skin, and can be an important driver of SSc pathogenesis.

Smad2/4 Signaling Pathway Is Critical for Epidermal Langerhans Cell Repopulation Under Inflammatory Condition but Not Required for Their Homeostasis at Steady State.

Epidermal Langerhans cells (LCs) are skin-resident dendritic cells that are essential for the induction of skin immunity and tolerance. Transforming growth factor-β 1 (TGFβ1) is a crucial factor for LC maintenance and function. However, the underlying TGFβ1 signaling pathways remain unclear. Our previous research has shown that the TGFβ1/Smad3 signaling pathway does not impact LC homeostasis and maturation. In this study, we generated mice with conditional deletions of either individual Smad2, Smad4, or both Smad2 and Smad4 in the LC lineage or myeloid lineage, to further explore the impact of TGFβ1/Smad signaling pathways on LCs. We found that interruption of Smad2 or Smad4 individually or simultaneously in the LC lineage did not significantly impact the maintenance, maturation, antigen uptake, and migration of LCs in vivo or in vitro during steady state. However, the interruption of both Smad2 and Smad4 pathways in the myeloid lineage led to a dramatic inhibition of bone marrow-derived LCs in the inflammatory state. Overall, our data suggest that canonical TGFβ1/Smad2/4 signaling pathways are dispensable for epidermal LC homeostasis and maturation at steady state, but are critical for the long-term LC repopulation directly originating from the bone marrow in the inflammatory state.

Amblyomma sculptum Salivary PGE2 Modulates the Dendritic Cell-Rickettsia rickettsii Interactions in vitro and in vivo

Amblyomma sculptum is an important vector of Rickettsia rickettsii, causative agent of Rocky Mountain spotted fever and the most lethal tick-borne pathogen affecting humans. To feed on the vertebrate host's blood, A. sculptum secretes a salivary mixture, which may interact with skin resident dendritic cells (DCs) and modulate their function. The present work was aimed at depicting the A. sculptum saliva-host DC network and the biochemical nature of the immunomodulatory component(s) involved in this interface. A. sculptum saliva inhibits the production of inflammatory cytokines by murine DCs stimulated with LPS. The fractionation of the low molecular weight salivary content by reversed-phase chromatography revealed active fractions eluting from 49 to 55% of the acetonitrile gradient. Previous studies suggested that this pattern of elution matches with that observed for prostaglandin E2 (PGE2) and the molecular identity of this lipid mediator was unambiguously confirmed by a new high-resolution mass spectrometry methodology. A productive infection of murine DCs by R. rickettsii was demonstrated for the first time leading to proinflammatory cytokine production that was inhibited by both A. sculptum saliva and PGE2, a result also achieved with human DCs. The adoptive transfer of murine DCs incubated with R. rickettsii followed by treatment with A. sculptum saliva or PGE2 did not change the cytokine profile associated to cellular recall responses while IgG2a-specific antibodies were decreased in the serum of these mice. Together, these findings emphasize the role of PGE2 as a universal immunomodulator of tick saliva. In addition, it contributes to new approaches to explore R. rickettsii-DC interactions both in vitro and in vivo.

Abstract A102: Rescue of lost skin dendritic cells in melanoma is key for the resuscitation of antitumor T-cell responses

Abstract One major characteristic of skin-related cancers is the loss of skin-resident dendritic cell populations. In the tg(Grm1)EPv mouse model, ectopic expression of the metabotropic glutamate receptor-1 in melanocytes leads to a highly proliferative and antiapoptotic phenotype, resulting in melanoma formation within the dermis. The slow progression of these tumors allows for the in depth analysis of the immune infiltrate in growing tumors. We here show that total DCs (CD11c+ cells) are gradually lost as the tumor progresses. Mainly the dermal CD11b+ DCs are affected, whereas epidermal Langerhans cells remain unchanged. We hypothesized that extrinsic cell death of the DCs may be induced within the growing tumor. Indeed, the tumor tissue upregulated the expression of Fas ligand (FasL) that can induce extrinsic apoptosis in immature DCs that express Fas (CD95). Fas-mediated apoptosis depends on the activation of caspase-8 and mice in which the autoproteolytic cleavage site D387 is mutated to alanine (casp8D387A/casp8D387A) show strong resistance to Fas-mediated death. We compared the apoptosis induction in the different DC subsets in casp8WT/casp8WT and in casp8D387A/casp8D387A upon intradermal administration of an agonistic anti-CD95 antibody and we found that, indeed, CD11b+ DCs in WT mice are susceptible to apoptosis via CD95. In order to retrieve the intratumoral DCs, we treated tg(Grm1)EPv mice carrying tumor lesions with Flt3L and the percentages of the CD11b+ subset could be restored back to the levels of tumor-free mice. At the same time, T-cells from both the tumor and the tumor-draining lymph node were able to produce more cytotoxic cytokines. Our data thus indicate that the loss of DCs in melanoma depends on induction of apoptosis within the tumor; rescue of skin DCs may be a promising strategy in order to enhance the efficacy of existing immunotherapeutic strategies against skin-related cancers. Citation Format: Anastasia Prokopi, Christoph H. Tripp, Bart Tummers, Kerstin Komenda, Katharina Hutter, Giuseppe Cappellano, Lydia Bellmann, Mirjana Efremova, Zlatko Trajanoski, Suzie Chen, Bjorn E. Clausen, Douglas R. Green, Patrizia Stoitzner. Rescue of lost skin dendritic cells in melanoma is key for the resuscitation of antitumor T-cell responses [abstract]. In: Proceedings of the Fourth CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; Sept 30-Oct 3, 2018; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2019;7(2 Suppl):Abstract nr A102.

Infectivity and growth kinetics of Herpes Simplex Virus type-2 in MOLT4 CCR5+ and CEM CCR5+ T cell lines

Herpes simplex virus type-2 (HSV-2) is an important sexually transmitted pathogen that infects the genital mucosal epithelial cells causing ulcerative lesions at the site of entry, facilitating HIV infection. The infection of epithelial cells and skin resident dendritic cells with HSV-2 causes a release of chemokine and retinoic acid which attracts CD4+ T-cells to the genital mucosa. In this study, we investigated whether HSV-2 (ATCC VR734) could infect and replicate in two T-cell lines (CEM CCR5+ and MOLT4 CCR5+). The growth of HSV-2 was assessed by plaque assay while the intracellular HSV-2 was identified using infectious center and indirect immunofluorescence assays. The replication of HSV-2 in T-cell lines was compared to a cell line (Vero) which is routinely used for growing HSV-2. Analysis indicated that a low level of infection was detected in the two T-cells lines and was dependent on the infectious dose as well as the time of adsorption. Indirect immunofluorescence showed presence of HSV-2 antigens in the CEM CCR5+ and Vero cell lines but not in MOLT4 CCR5+. The data suggests that T-cells can support growth of HSV-2 which might contribute to changes in gene expression of T-cells. This is an important aspect that needs to be further investigated in relation of HIV-1/HSV-2 viral synergy.

Isolation of Murine Skin Resident and Migratory Dendritic Cells via Enzymatic Digestion.

Dendritic cells (DCs) are a highly specialized subset of professional antigen-presenting cells (APCs) that reside in peripheral and lymphoid tissues. DCs capture antigen in the periphery and migrate to the lymph node where they prime naïve T cells. In addition, DCs have been recently appreciated to have function in innate immunity within tissues. In the skin, heterogeneous populations of DCs reside within the epidermis and the dermis. Analysis of the cutaneous DC subsets is complicated by requirements of distinct enzymatic digestion protocols for isolation of APCs from distinct anatomical compartments of the skin. Here, specific approaches for isolation of DCs from the epidermis, dermis, and the skin-draining lymph nodes of mice are described. © 2018 by John Wiley & Sons, Inc.

A psoriasis target in Langerhans cells

Immunology Psoriasis is an autoimmune skin condition that is linked to the proinflammatory cytokine interleukin-17 (IL-17), which is produced by T cells in the skin. Zheng et al. found that p38α signaling in skin-resident dendritic cells (Langerhans cells) was important for the pathogenesis of

DC-SIGN promotes Japanese encephalitis virus transmission from dendritic cells to T cells via virological synapses

Skin-resident dendritic cells (DCs) likely encounter incoming viruses in the first place, and their migration to lymph nodes following virus capture may promote viral replication. However, the molecular mechanisms underlying these processes remain unclear. In the present study, we found that compared to cell-free viruses, DC-bound viruses showed enhanced capture of JEV by T cells. Additionally, JEV infection was increased by co-culturing DCs and T cells. Blocking the C-type lectin receptor DC-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) with neutralizing antibodies or antagonists blocked JEV transmission to T cells. Live-cell imaging revealed that DCs captured and transferred JEV viral particles to T cells via virological synapses formed at DC-T cell junctions. These findings indicate that DC-SIGN plays an important role in JEV transmission from DCs to T cells and provide insight into how JEV exploits the migratory and antigen-presenting capabilities of DCs to gain access to lymph nodes for dissemination and persistence in the host.

IL-32 induces indoleamine 2,3-dioxygenase+CD1c+ dendritic cells and indoleamine 2,3-dioxygenase+CD163+ macrophages: Relevance to mycosis fungoides progression

ABSTRACTMycosis fungoides (MF) progresses from patch to tumor stage by expansion of malignant T-cells that fail to be controlled by protective immune mechanisms. In this study, we focused on IL-32, a cytokine, highly expressed in MF lesions. Depending on the other cytokines (IL-4, GM-CSF) present during in vitro culture of healthy volunteers' monocytes, IL-32 increased the maturation of CD11c+ myeloid dendritic cells (mDC) and/or CD163+ macrophages, but IL-32 alone showed a clear ability to promote dendritic cell (DC) differentiation from monocytes. DCs matured by IL-32 had the phenotype of skin-resident DCs (CD1c+), but more importantly, also had high expression of indoleamine 2,3-dioxygenase. The presence of DCs with these markers was demonstrated in MF skin lesions. At a molecular level, indoleamine 2,3-dioxygenase messenger RNA (mRNA) levels in MF lesions were higher than those in healthy volunteers, and there was a high correlation between indoleamine 2,3-dioxygenase and IL-32 expression. In contrast, Foxp3 mRNA levels decreased from patch to tumor stage. Increasing expression of IL-10 across MF lesions was highly correlated with IL-32 and indoleamine 2,3-dioxygenase, but not with Foxp3 expression. Thus, IL-32 could contribute to progressive immune dysregulation in MF by directly fostering development of immunosuppressive mDC or macrophages, possibly in association with IL-10.

Uptake of synthetic naked RNA by skin-resident dendritic cells via macropinocytosis allows antigen expression and induction of T-cell responses in mice.

Intradermal administration of antigen-encoding RNA has entered clinical testing for cancer vaccination. However, insight into the underlying mechanism of RNA uptake, translation and antigen presentation is still limited. Utilizing pharmacologically optimized naked RNA, the dose-response kinetics revealed a rise in reporter signal with increasing RNA amounts and a prolonged RNA translation of reporter protein up to 30days after intradermal injection. Dendritic cells (DCs) in the dermis were shown to engulf RNA, and the signal arising from the reporter RNA was significantly diminished after DC depletion. Macropinocytosis was relevant for intradermal RNA uptake and translation in vitro and in vivo. By combining intradermal RNA vaccination and inhibition of macropinocytosis, we show that effective priming of antigen-specific CD8(+) T-cells also relies on this uptake mechanism. This report demonstrates that direct antigen translation by dermal DCs after intradermal naked RNA vaccination is relevant for efficient priming of antigen-specific T-cells.

Commensal-dendritic-cell interaction specifies a unique protective skin immune signature.

Abstract The skin represents the body’s primary interface with the environment, acting as the first line of physical and immunological defense. This organ is also home to trillions of microbes (bacteria, fungi and viruses) that play an important role in tissue homeostasis and local immunity. Skin microbial communities are highly diverse and can be remodeled over time or in response to environmental challenges. How in the context of this complexity, individual commensals may differentially modulate skin immunity and what the consequences of these responses for tissue physiology are remain unclear. Here, we demonstrate that defined commensals dominantly impact skin immunity and identify the cellular mediators involved in this specification. In particular, colonization with Staphylococcus epidermidis induces IL-17A+CD8+T cells that home to the epidermis, enhance innate barrier immunity and limit pathogen invasion. These commensal-specific T cell responses result from the coordinated action of skin resident dendritic cell subsets and are not associated with inflammation, revealing that tissue resident cells are poised to sense and respond to alterations in microbial communities. This dialogue may represent an evolutionary means by which the skin immune system uses fluctuating commensal clues to calibrate barrier immunity and provide heterologous protection against invasive pathogens. Altogether these findings reveal that the skin immune landscape is a highly dynamic environment that can be rapidly and specifically remodeled by encounters with defined commensals, findings that have profound implications for our understanding of tissue specific immunity and pathologies.

DC-SIGN as an attachment factor mediates Japanese encephalitis virus infection of human dendritic cells via interaction with a single high-mannose residue of viral E glycoprotein

The skin-resident dendritic cells (DCs) are thought to be the first defender to encounter incoming viruses and likely play a role in Japanese encephalitis virus (JEV) early infection. In the current study, following the demonstration of JEV productive infection in DCs, we revealed that the interaction between JEV envelope glycoprotein (E glycoprotein) and DC-SIGN was important for such infection as evidenced by antibody neutralization and siRNA knockdown experiments. Moreover, the high-mannose N-linked glycan at N154 of E glycoprotein was shown to be crucial for JEV binding to DC-SIGN and subsequent internalization, while mutation of DC-SIGN internalization motif did not affect JEV uptake and internalization. These data together suggest that DC-SIGN functions as an attachment factor rather than an entry receptor for JEV. Our findings highlight the potential significance of DC-SIGN in JEV early infection, providing a basis for further understanding how JEV exploits DC-SIGN to gain access to dendritic cells.