Early antitumor activity of oral Langerhans cells is compromised by a carcinogen.

Early diagnosis of oral squamous cell carcinoma (OSCC) remains an unmet clinical need. Therefore, elucidating the initial events of OSCC preceding tumor development could benefit OSCC prognosis. Here, we define the Langerhans cells (LCs) of the tongue and demonstrate that LCs protect the epithelium from carcinogen-induced OSCC by rapidly priming αßT cells capable of eliminating γH2AX+ epithelial cells, whereas γδT and natural killer cells are dispensable. The carcinogen, however, dysregulates the epithelial resident mononuclear phagocytes, reducing LC frequencies, while dendritic cells (DCs), macrophages, and plasmacytoid DCs (pDCs) populate the epithelium. Single-cell RNA-sequencing analysis indicates that these newly differentiated cells display an immunosuppressive phenotype accompanied by an expansion of T regulatory (Treg) cells. Accumulation of the Treg cells was regulated, in part, by pDCs and precedes the formation of visible tumors. This suggests LCs play an early protective role during OSCC, yet the capacity of the carcinogen to dysregulate the differentiation of mononuclear phagocytes facilitates oral carcinogenesis.

Flavonoids in Skin Senescence Prevention and Treatment.

Skin aging is associated with the accumulation of senescent cells and is related to many pathological changes, including decreased protection against pathogens, increased susceptibility to irritation, delayed wound healing, and increased cancer susceptibility. Senescent cells secrete a specific set of pro-inflammatory mediators, referred to as a senescence-associated secretory phenotype (SASP), which can cause profound changes in tissue structure and function. Thus, drugs that selectively eliminate senescent cells (senolytics) or neutralize SASP (senostatics) represent an attractive therapeutic strategy for age-associated skin deterioration. There is growing evidence that plant-derived compounds (flavonoids) can slow down or even prevent aging-associated deterioration of skin appearance and function by targeting cellular pathways crucial for regulating cellular senescence and SASP. This review summarizes the senostatic and senolytic potential of flavonoids in the context of preventing skin aging.

Dissolvable microneedles based on Panax notoginseng polysaccharide for transdermal drug delivery and skin dendritic cell activation.

This work explored the feasibility of using biological polysaccharide to fabricate dissolvable microneedles (MNs) for the purpose of transdermal drug delivery and skin dendritic cell (DC) activation. Panax notoginseng polysaccharide (PNPS), a naturally derived immunoactive macromolecule, was used to fabricate dissolvable MNs. The prepared PNPS MNs showed a satisfactory mechanical strength and a skin penetration depth. By Franz diffusion cell assay, the PNPS MNs demonstrated a high transdermal delivery amount of model drugs. Furthermore, with the assistance of MNs, PNPS easily penetrated across the stratum corneum and target ear skin DCs, activating the maturation and migration of immunocytes by increasing the expressions of CD40, CD80, CD86, and MHC II of skin DCs. Consequently, the matured DCs migrated to the auricular draining lymph nodes and increased the proportions of CD4+ T and CD8+ T cells. Thus, PNPS might be a promising biomaterial for transdermal drug delivery, with adjuvant potential.

Lysosomotropic beta blockers induce oxidative stress and IL23A production in Langerhans cells.

Oxidative stress and Th17 cytokines are important mediators of inflammation. Treatment with beta-adrenoceptor (ADRB) antagonists (beta-blockers) is associated with induction or aggravation of psoriasis-like skin inflammation, yet the underlying mechanisms are poorly understood. Herein, we identify lysosomotropic beta-blockers as critical inducers of IL23A in human monocyte-derived Langerhans-like cells under sterile-inflammatory conditions. Cytokine release was not mediated by cAMP, suggesting the involvement of ADRB-independent pathways. NFKB/NF-κB and MAPK14/p38 activation was required for propranolol-induced IL23A secretion whereas the NLRP3 inflammasome was dispensable. MAPK14 regulated recruitment of RELB to IL23A promoter regions. Without affecting the ubiquitin-proteasome pathway, propranolol increased lysosomal pH and induced a late-stage block in macroautophagy/autophagy. Propranolol specifically induced reactive oxygen species production, which was critical for IL23A secretion, in Langerhans-like cells. Our findings provide insight into a potentially crucial immunoregulatory mechanism in cutaneous dendritic cells that may explain how lysosomotropic drugs regulate inflammatory responses. ABBREVIATIONS: ATF: activating transcription factor; DC: dendritic cell; ChIP: chromatin immunoprecipitation; gDNA: genomic DNA; IL: interleukin; LAMP1: lysosomal associated membrane protein 1; LC: Langerhans cell; LPS: lipopolysaccharide; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPK: mitogen-activated protein kinase; MoDC: monocyte-derived DC; MoLC: monocyte-derived Langerhans-like cell; mtDNA: mitochondrial DNA; NAC: N-acetyl-L-cysteine; NLRP3: NLR family pyrin domain containing 3; PBMC: peripheral blood mononuclear cell; PI: propidium iodide; PYCARD/ASC: PYD and CARD domain containing; qRT-PCR: quantitative real-time PCR; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1; TLR: Toll-like receptor; TRAF6: TNF receptor associated factor 6; TNF: tumor necrosis factor; Ub: ubiquitin.

Silencing of PD-L2/B7-DC by Topical Application of Small Interfering RNA Inhibits Elicitation of Contact Hypersensitivity.

PD-L2 is a ligand for the immune checkpoint receptor PD-1; however, its regulatory function is unclear. We previously reported that silencing of CD86 in cutaneous dendritic cells by topical application of small interfering RNA (siRNA) inhibits the elicitation of contact hypersensitivity (CHS). Here, we investigated the effects of topical application of PD-L2 siRNA on allergic skin disease. PD-L2 was induced in dendritic cells concurrently with the elevation of major histocompatibility complex class II and CD86 expression. Topical application of PD-L2 siRNA inhibited the elicitation of CHS by suppressing early proinflammatory cytokine expression and migration of hapten-carrying dendritic cells into lymph nodes. Local injection of neutralizing anti-PD-L2 mAb inhibited CHS to the same extent. PD-L2 siRNA treatment inhibited CHS in PD-1/PD-L1 double knockout mice and in the sensitized T-cell-transferred skin. These results suggest that the effects of PD-L2 silencing are independent of PD-1 but dependent on local memory T cells. Most of the inhibitory effects of PD-L2 and CD86 silencing on CHS were comparable, but PD-L2 siRNA treatment did not inhibit atopic disease-like manifestations and T helper type 2 responses in NC/Nga mice. Our results suggest that PD-L2 in cutaneous dendritic cells acts as a costimulator rather than a regulator. Local PD-L2 silencing by topical application of siRNA represents a therapeutic approach for contact allergy.

The role of Langerhans cells in pathologies of the skin.

Langerhans cells (LCs) are epidermal immune cells of myeloid origin. Although these cells were primarily thought to play a defensive role in the skin, evidence now indicates a diverse range of LC-mediated effects including the relay of viral antigens in herpes simplex infection, recruitment of eosinophils in atopic dermatitis and promotion of a Th17 response in Candida infection. LCs may have a protective or suppressive function in pathologies of the skin, with differing functions being driven by the skin milieu. Understanding LC function will help guide the development of interventions that modulate these cells for therapeutic benefit.

Photodynamic Therapy for Genital Warts Causes Activation of Local Immunity.

BACKGROUND: 5-aminolevulinic acid photodynamic therapy (PDT) for genital warts is effective, safe, and can prevent recurrence. It is believed that PDT can induce immune responses, but the mechanism is not completely understood. OBJECTIVES: The objectives of this article are to confirm the effect of PDT for genital warts on local immunity and to investigate the recruitment and significance of immune cells in tissues. METHODS: Local immune changes in T lymphocytes (CD3+, CD4+, CD8+), plasmacytoid dendritic cells (pDCs) (CD123+), and myeloid dendritic cells (CD1a+) after PDT in patients were evaluated by immunohistochemistry staining. Changes in mRNA levels of IFN-γ, IFN-α, IFN-ß, interferon-stimulated gene 15 kDa (ISG-15), Mx2, Toll-like receptor 9 (TLR9), and interferon regulatory factor 7 (IRF7) were analyzed by real-time quantitative polymerase chain reaction. RESULTS: At 4 hours after PDT, CD4+ increased, accompanied by increased levels of mRNA expression of IFN-γ, but CD4+ and mRNA expression levels of IFN-γ were decreased at 24 hours after PDT. CD123+ pDCs showed an increasing trend. CD1a+ LCs in the epidermis gradually decreased, and DCs in the epidermis gradually increased. CD3+ infiltrated and migrated to the superficial dermis, but CD8+ did not change significantly after PDT. The mRNA expression levels of IFN-α, IFN-ß, ISG-15, Mx2, TLR9, and IRF7 showed an increasing trend after PDT. As compared with the patients without significantly increased IFN-α and IFN-ß after PDT sessions, patients with significant increases needed fewer sessions of PDT for remission. CONCLUSIONS: PDT for genital warts can activate T lymphocyte-mediated, DC-related, and pDC-related immunity. The clinical efficacy of PDT for genital warts may be related to the increased levels of IFN-α and IFN-ß after treatment.

In Vitro Induction of T Helper 17 Cells by Synergistic Activation of Human Monocyte-Derived Langerhans Cell-Like Cells with Bacterial Agonists.

In the case of epidermal barrier disruption, pathogens encounter skin-resident Langerhans cells (LCs) and are recognized by pathogen recognition receptors such as Toll-like receptors (TLRs). As the majority of microorganisms exhibit more than one TLR ligand, the mechanisms of subsequent T cell differentiation are complex and far from clear. In this study, we investigated combinatory effects on Th cell polarization by bacterial cell wall compounds peptidoglycan (PGN) and lipopolysaccharide (LPS) and by bacterial nucleic acid (DNA). Expression of maturation markers CD40, CD80, HLA-DR and CCR7 and the release of IL-1ß, IL-6 and IL-23 was strongly enhanced by simultaneous exposure to PGN, LPS and DNA in LCs. As all these factors were potential Th17 driving cytokines, we investigated the potency of combinatory TLR stimuli to induce Th17 cells via LC activation. High amounts of IL-17A and IL-22, key cytokines of Th17 cells, were detected. By intracellular costaining of IL-17⁺T cells, IL-22- (Th17) and IL-22⁺ (immature Th17) cells were identified. Interestingly, one population of LPS stimulated cells skewed into IL-9⁺Th cells, and LPS synergized with PGN while inducing high IL-22. In conclusion, our data indicates that when mediated by a fine-tuned signal integration via LCs, bacterial TLR agonists synergize and induce Th17 differentiation.

In vivo cancer vaccination: Which dendritic cells to target and how?

The field of cancer immunotherapy has been revolutionized with the use of immune checkpoint blockade antibodies such as anti-programmed cell death 1 protein (PD-1) and chimeric antigen receptor T cells. Significant clinical benefits are observed in different cancer types with these treatments. While considerable efforts are made in augmenting tumor-specific T cell responses with these therapies, other immunotherapies that actively stimulate endogenous anti-tumor T cells and generating long-term memory have received less attention. Given the high cost of cancer immunotherapies especially with chimeric antigen receptor T cells, not many patients will have access to such treatments. The next-generation of cancer immunotherapy could entail in vivo cancer vaccination to activate both the innate and adaptive anti-tumor responses. This could potentially be achieved via in vivo targeting of dendritic cells which are an indispensable link between the innate and adaptive immunities. Dendritic cells highly expressed toll-like receptors for recognizing and eliminating pathogens. Synthetic toll-like receptors agonists could be synthesized at a low cost and have shown promise in preclinical and clinical trials. As different subsets of human dendritic cells exist in the immune system, activation with different toll-like receptor agonists could exert profound effects on the quality and magnitude of anti-tumor T cell responses. Here, we reviewed the different subsets of human dendritic cells. Using published preclinical and clinical cancers studies available on PubMed, we discussed the use of clinically approved and emerging toll-like receptor agonists to activate dendritic cells in vivo for cancer immunotherapy. Finally, we searched www.clinicaltrials.gov and summarized the active cancer trials evaluating toll-like receptor agonists as an adjuvant.

Immunological dysfunction in chronic arsenic exposure: From subclinical condition to skin cancer.

Exposure to arsenic is a global health issue. Long-term arsenic exposure may associate with various cancers and many other pathological effects. Over 100 million people worldwide are exposed to arsenic particularly in countries such as Bangladesh, Chile, China, India, Mexico, Taiwan and the USA. Drinking of water contaminated with arsenic is the major route of human exposure. Skin lesions are considered to be the most common adverse effects associated with chronic arsenic exposure. Skin lesions usually develop with the latency period spanning more than 20 years from first exposure. Arsenic-induced Bowen's disease, the most frequently encountered carcinoma in situ resulting from chronic arsenic exposure, is characterized by multiple and recrudescent lesions. Long-term arsenic exposure results in impaired immunity in susceptible individuals. In the prenatal stage, enhanced placental inflammatory responses and reduced placental T cells by arsenic may result in decreased thymic size and functions in newborns. In childhood, arsenic exposure may reduce peripheral CD4+ cells and interleukin-2 secretion which leads to susceptibility to opportunistic infections. There was an impairment of macrophage function and oxidative DNA damage of peripheral polymorphonuclear leukocytes in adults with skin lesions. In arsenic-induced Bowen's disease lesions, a decrease in the number and functions of Langerhans cells and, in parallel, a selective CD4+ cell apoptosis was noticed. These findings provide scientific evidence for understanding the phenomenon of arsenic-induced immune escape in the early stage of carcinogenesis and a reasonable explanation for multiple and recrudescent arsenic cancers in the skin.

Activation profile of THP-1 derived dendritic cells stimulated by allergen Mal f 1 beyond its IgE-binding ability.

BACKGROUND: Mal f 1, the first allergen cloned from Malassezia furfur, has positive IgE reactivity in sera from atopic dermatitis (AD) patients. The mechanism by which Mal f 1 induces the maturation of human dendritic cells (DCs) and maintains the symptoms of AD is not well understood. OBJECTIVE: The present study aims to explore the activation profile of THP-1 derived dendritic cells (TDDCs) stimulated by recombinant Mal f 1, as well as to explore the IgE-binding ability of rMal f 1 and its correlation with IgE-binding activity of complete allergens of M. furfur. METHODS: rMal f 1 was produced by expression in E. coli and purification with affinity chromatography. The ability of rMal f 1 and ImmunoCAP complete allergens of M. furfur to bind to serum specific IgE was assayed in parallel by ELISA and immunoblotting. Immature TDDCs were stimulated with rMal f 1 or an enzyme-digested product of rMal f 1. The expression levels of markers, CD83, CD80, CD86, and HLA-DR, were investigated by flow cytometry. The levels of interleukin (IL)-6, IL-10, IL-12p70 and tumor necrosis factor (TNF)-α in culture supernatants were determined by ELISA. RESULTS: Eighteen patient sera were identified that reacted positively to the complete allergens of M. furfur as determined by ImmunoCAP and also showed positive responses to rMal f 1. Five patient sera were identified that had no reaction to ImmunoCAP complete allergens of M. furfur and also exhibited negative response to rMal f 1. All sera, except for one, had no reaction to the unrelated allergen Bet v 1. rMal f 1 upregulated the maturation surface marker CD83 on TDDCs. In addition, rMal f 1 also induced high levels of CD80 and CD86. Increased expression of HLA-DR, a first signal for T cell activation, was observed. Secretion of IL-6, TNF-α and IL-10 by TDDCs increased significantly (P < 0.0001 for IL-6, P < 0.01 for TNF-α and P < 0.05 for IL-10) after stimulation by rMal f 1, while the IL-12p70 level was unaltered. CONCLUSION: We have shown that rMal f 1 has ideal IgE binding ability and good correlation with binding activity to M. furfur. Moreover, we have revealed a hitherto unknown DC activation profile after rMal f 1 stimulation whereby TNF-α, IL-6, and IL-10 were significantly increased and IL-12 was unaltered, suggesting that rMal f 1 can predispose a DC bias toward the TH22/TH17 pathway beyond the routine IgE-dependent TH2 pathway, thus providing intriguing clues for clinical treatment involving both pathways.

Epicutaneous administration of the pattern recognition receptor agonist polyinosinic-polycytidylic acid activates the MDA5/MAVS pathway in Langerhans cells.

Together with keratinocytes (KCs) and the dense network of Langerhans cells (LCs), the epidermis is an ideal portal for vaccine delivery. Pattern recognition receptor agonists, in particular polyinosinic-polycytidylic acid [p(I:C)], are promising adjuvant candidates for therapeutic vaccination to generate protective T-cell immunity. Here we established an ex vivo skin explant model to study the expression and activation of double-stranded RNA (dsRNA)-sensing pattern recognition receptors in LCs and KCs in human skin. Whereas KCs expressed all known dsRNA sensing receptors at a constitutive and inducible level, LCs exclusively expressed melanoma differentiation-associated protein 5 (MDA5) in untreated skin and freshly isolated cells. Comparative assessments of downstream signaling pathways induced by p(I:C) revealed distinct mitochondrial antiviral-signaling protein, IFN-regulatory factor 3, and NF-κB activation in LCs and KCs. Consequently, p(I:C) treatment of LCs significantly induced IFN-α and IFN-ß mRNA expression, while in KCs an up-regulation of IFN-ß and TNF-α mRNA was detectable. Stimulation of LCs with specific ligands revealed that not the TLR3- but only the MDA5-specific ligand induced IFN-α2, IFN-ß, and TNF-α cytokines, but no IL-6 and -8. In KCs, both ligands induced production of high IL-6 and IL-8 levels, and low IFN-α2 and IFN-ß levels, indicating that different dsRNA-sensing receptors and/or downstream signaling pathways are activated in both cell types. Our data suggest that MDA5 may be an attractive adjuvant target for epicutaneous delivery of therapeutic vaccines with the goal to target LCs.-Tajpara, P., Schuster, C., Schön, E., Kienzl, P., Vierhapper, M., Mildner, M., Elbe-Bürger, A. Epicutaneous administration of the pattern recognition receptor agonist polyinosinic-polycytidylic acid activates the MDA5/MAVS pathway in Langerhans cells.

Microparticles produced by human papillomavirus type 16 E7-expressing cells impair antigen presenting cell function and the cytotoxic T cell response.

High-risk, cancer-causing human papillomaviruses (HPV) cause infections of the epidermis that may progress to cancer, including cervical cancer. Viral persistence, contributed to by viral evasion of the host immune response, is associated with the likelihood of cancer developing. Langerhans cells (LCs) are the only professional antigen presenting cells located in the epidermis, therefore may influence the antiviral immune response. Microparticles, or microvesicles, are small membrane particles shed by cells that can exert effects on other cells at both a local and systemic level. We found increased numbers of microparticles were shed from human or mouse keratinocytes expressing the HPV16 E7 oncoprotein, compared with control keratinocytes. Co-culture of LCs with microparticles from E7-expressing cells suppressed the cytotoxic T cell response. We attributed this, at least in part, to the reduction in surface of CD40 and intracellular pro-inflammatory cytokine IL-12 p40 subunit that we measured in the LCs. The evidence provided here shows that co-culture of E7-microparticles with LCs inhibits antigen-specific cytotoxicity. This is an important finding, suggesting that microparticles from HPV-infected cells could suppress the T cell response by regulating LCs, potentially contributing to persistence of HPV infection and cancer.

Physiological Doses of Red Light Induce IL-4 Release in Cocultures between Human Keratinocytes and Immune Cells.

Phototherapy is routinely used for the treatment of various skin conditions and targeted therapy of superficial cancers. However, the molecular mechanisms behind their biological effects and the need for efficacy enhancing photosensitizers are not well addressed. Particularly, not much is known about the inherent effect of light from the visible spectrum on cytokine release and its downstream effects in keratinocytes and immune cells located in skin and therefore exposed to light. To address this, we delivered calibrated doses of well-defined light qualities (380 to 660 nm) to cocultures of human keratinocytes and macrophage/dendritic cells in the absence or presence of the commonly used photosensitizer 8-methoxypsoralen (8-MOP). The experiments identified IL-4 as a key effector cytokine released by this coculture model with need for 8-MOP in the UVA1 /blue (380 nm) and no requirement for photosensitizer in the red light spectrum (627 nm). 3D organotypic skin cultures treated with IL-4 showed thickening of the epidermal layer and delayed differentiation. However unlike IL-4 and UVA1 /blue light treatment, red light did not reduce the expression of keratinocyte differentiation markers or increase signs of photo-oxidative damage. This supports the application of isolated red light as a possible alternative for photo-immunotherapy without need for additional photosensitizers.

Lower levels of interleukin-1ß gene expression are associated with impaired Langerhans' cell migration in aged human skin.

Langerhans' cells (LC) play pivotal roles in skin immune responses, linking innate and adaptive immunity. In aged skin there are fewer LC and migration is impaired compared with young skin. These changes may contribute to declining skin immunity in the elderly, including increased skin infections and skin cancer. Interleukin-1ß (IL-1ß) and tumour necrosis factor-α (TNF-α) are mandatory signals for LC migration and previous studies suggest that IL-1ß signalling may be dysregulated in aged skin. Therefore, we sought to explore the mechanisms underlying these phenomena. In skin biopsies of photoprotected young (< 30 years) and aged (> 70 years) human skin ex vivo, we assessed the impact of trauma, and mandatory LC mobilizing signals on LC migration and gene expression. Biopsy-related trauma induced LC migration from young epidermis, whereas in aged skin, migration was greatly reduced. Interleukin-1ß treatment restored LC migration in aged epidermis whereas TNF-α was without effect. In uncultured, aged skin IL-1ß gene expression was lower compared with young skin; following culture, IL-1ßmRNA remained lower in aged skin under control and TNF-α conditions but was elevated after culture with IL-1ß. Interleukin-1 receptor type 2 (IL1R2) gene expression was significantly increased in aged, but not young skin, after cytokine treatment. Keratinocyte-derived factors secreted from young and aged primary cells did not restore or inhibit LC migration from aged and young epidermis, respectively. These data suggest that in aged skin, IL-1ß signalling is diminished due to altered expression of IL1B and decoy receptor gene IL1R2.

Transcutaneous immunization with a novel imiquimod nanoemulsion induces superior T cell responses and virus protection.

BACKGROUND: Transcutaneous immunization (TCI) is a novel vaccination strategy utilizing the skin associated lymphatic tissue to induce immune responses. TCI using a cytotoxic T lymphocyte (CTL) epitope and the Toll-like receptor 7 (TLR7) agonist imiquimod mounts strong CTL responses by activation and maturation of skin-derived dendritic cells (DCs) and their migration to lymph nodes. However, TCI based on the commercial formulation Aldara only induces transient CTL responses that needs further improvement for the induction of durable therapeutic immune responses. OBJECTIVE: Therefore we aimed to develop a novel imiquimod solid nanoemulsion (IMI-Sol) for TCI with superior vaccination properties suited to induce high quality T cell responses for enhanced protection against infections. METHODS: TCI was performed by applying a MHC class I or II restricted epitope along with IMI-Sol or Aldara (each containing 5% Imiquimod) on the shaved dorsum of C57BL/6, IL-1R, Myd88, Tlr7 or Ccr7 deficient mice. T cell responses as well as DC migration upon TCI were subsequently analyzed by flow cytometry. To determine in vivo efficacy of TCI induced immune responses, CTL responses and frequency of peptide specific T cells were evaluated on day 8 or 35 post vaccination and protection in a lymphocytic choriomeningitis virus (LCMV) infection model was assessed. RESULTS: TCI with the imiquimod formulation IMI-Sol displayed equal skin penetration of imiquimod compared to Aldara, but elicited superior CD8+ as well as CD4+ T cell responses. The induction of T-cell responses induced by IMI-Sol TCI was dependent on the TLR7/MyD88 pathway and independent of IL-1R. IMI-Sol TCI activated skin-derived DCs in skin-draining lymph nodes more efficiently compared to Aldara leading to enhanced protection in a LCMV infection model. CONCLUSION: Our data demonstrate that IMI-Sol TCI can overcome current limitations of previous imiquimod based TCI approaches opening new perspectives for transcutaneous vaccination strategies and allowing the use of this enhanced cutaneous drug-delivery system to be tailored for the improved prevention and treatment of infectious diseases and cancers.

In vivo quantification of quantum dot systemic transport in C57BL/6 hairless mice following skin application post-ultraviolet radiation.

BACKGROUND: Previous work has demonstrated size, surface charge and skin barrier dependent penetration of nanoparticles into the viable layers of mouse skin. The goal of this work was to characterize the tissue distribution and mechanism of transport of nanoparticles beyond skin, with and without Ultraviolet Radiation (UVR) induced skin barrier disruption. Atomic absorption spectroscopy (AAS), flow cytometry and confocal microscopy were used to examine the effect of UVR dose (180 and 360 mJ/cm2 UVB) on the skin penetration and systemic distribution of quantum dot (QD) nanoparticles topically applied at different time-points post UVR using a hairless C57BL/6 mouse model. RESULTS: Results indicate that QDs can penetrate mouse skin, regardless of UVR exposure, as evidenced by the increased cadmium in the local lymph nodes of all QD treated mice. The average % recovery for all treatment groups was 69.68% with ~66.84% of the applied dose recovered from the skin (both epicutaneous and intracutaneous). An average of 0.024% of the applied dose was recovered from the lymph nodes across various treatment groups. When QDs are applied 4 days post UV irradiation, at the peak of the skin barrier defect and LC migration to the local lymph node, there is an increased cellular presence of QD in the lymph node; however, AAS analysis of local lymph nodes display no difference in cadmium levels due to UVR treatment. CONCLUSIONS: Our data suggests that Langerhans cells (LCs) can engulf QDs in skin, but transport to the lymph node may occur by both cellular (dendritic and macrophage) and non-cellular mechanisms. It is interesting that these specific nanoparticles were retained in skin similarly regardless of UVR barrier disruption, but the observed skin immune cell interaction with nanoparticles suggest a potential for immunomodulation, which we are currently examining in a murine model of skin allergy.

Langerhans cell markers CD1a and CD207 are the most rapidly responding genes in lesional psoriatic skin following adalimumab treatment.

TNFα-, IL-23- and IL-17-targeting drugs are highly effective in the treatment of psoriasis. However, the precise molecular mechanism remains unknown. In psoriatic skin, the presence of Langerhans cells (LCs) is reduced, but the role of LC is poorly understood. The purpose of this study was to investigate the impact of TNFα and IL-23/IL-17 on the presence of LC in the skin during treatment. Therefore, psoriatic skin was investigated before and after 4 days of adalimumab or ustekinumab treatment. Furthermore, TNFα and IL-17A stimulation was investigated in an ex vivo model of epidermis and dermis from healthy normal skin kept in cultures at an air-liquid interphase for 4 days. In a gene array analysis, we found that the two LC markers, CD1a and CD207, were among the most up- or downregulated genes in psoriatic skin after anti-TNFα therapy. Validation showed that both mRNA expression and protein level followed the same pattern and became significantly upregulated after 4 days of treatment. No changes were seen after ustekinumab treatment. In the ex vivo skin model, a decrease in the CD1a level was seen after TNFα stimulation and it was caused by LC migration from epidermis. No response in LC migration was seen after IL-17A stimulation. Taken together, we demonstrated that changes in the LC level in epidermis precede the histological and clinical changes during adalimumab treatment in psoriatic skin. Furthermore, TNFα plays a prominent role in orchestrating LC migration in the skin. This seems not to be the true for the IL-23/IL-17A pathway.

Langerin-mediated internalization of a modified peptide routes antigens to early endosomes and enhances cross-presentation by human Langerhans cells.

The potential of the skin immune system to generate immune responses is well established, and the skin is actively exploited as a vaccination site. Human skin contains several antigen-presenting cell subsets with specialized functions. In particular, the capacity to cross-present exogenous antigens to CD8+ T cells is of interest for the design of effective immunotherapies against viruses or cancer. Here, we show that primary human Langerhans cells (LCs) were able to cross-present a synthetic long peptide (SLP) to CD8+ T cells. In addition, modification of this SLP using antibodies against the receptor langerin, but not dectin-1, further enhanced the cross-presenting capacity of LCs through routing of internalized antigens to less proteolytic early endosome antigen 1+ early endosomes. The potency of LCs to enhance CD8+ T-cell responses could be further increased through activation of LCs with the toll-like receptor 3 ligand polyinosinic:polycytidylic acid (pI:C). Altogether, the data provide evidence that human LCs are able to cross-present antigens after langerin-mediated internalization. Furthermore, the potential for antigen modification to target LCs specifically provides a rationale for generating effective anti-tumor or anti-viral cytotoxic T lymphocyte responses.