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1.
Biomed Pharmacother ; 147: 112644, 2022 Mar.
Article in English | MEDLINE | ID: mdl-35051865

ABSTRACT

BACKGROUND: Psoriasis is an autoimmune disease mediated by T cells. However, treatment remains a clinical challenge because of its frequent recurrence. Celastrol (Cel), the main active ingredient in Tripterygium wilfordii Hook F, is efficacious in treating autoimmune diseases such as psoriasis. OBJECTIVE: To investigate the effect and mechanism of topical Cel in imiquimod (IMQ)-induced psoriasis-like inflammation in mice. METHODS: Female C57BL/6 and Langerin-diphtheria toxin receptor (DTR) mice were divided into Vehicle group and Cel gel groups. IMQ was used induce psoriasis-like inflammation to establish the mouse model of psoriasis. Hematoxylin and eosin staining was used to observe changes in local inflammatory cells in the skin lesions. Enzyme-linked immunosorbent assay was used to detect protein expression levels. Flow cytometry was used to detect the cell number and cytokine expression. Polymerase chain reaction was used to detect cytokine gene expression. RESULTS: Cel gel targeted the Langerhans cells. In IMQ-induced psoriatic dermatitis, Cel gel reduced the secretion of interleukin (IL)- 23 by Langerhans cells, suppressed the interaction between Langerhans cells and γδT cells, and decreased the number of activated γδT cells and related IL-17 secretion, alleviating psoriasis-like inflammation. Furthermore, Cel gel demonstrated a glucocorticoid-like effect that could impede the recurrence of psoriasis. CONCLUSION: Cel gel reduces the secretion of IL-23 from LCs and inhibits the interaction between LCs and γδT cells to alleviate psoriasis. It also shows an effect similar to that of glucocorticoids, which can prevent psoriasis recurrence. These findings provide a new direction for the clinical treatment of psoriasis and contribute to the development of novel drugs.


Subject(s)
Langerhans Cells/drug effects , Pentacyclic Triterpenes/pharmacology , Psoriasis/pathology , Animals , Disease Models, Animal , Female , Gels , Imiquimod/pharmacology , Interleukin-23/metabolism , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Tripterygium
2.
Proc Natl Acad Sci U S A ; 119(3)2022 01 18.
Article in English | MEDLINE | ID: mdl-35012988

ABSTRACT

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.


Subject(s)
Antineoplastic Agents/metabolism , Carcinogens/toxicity , Langerhans Cells/metabolism , 4-Nitroquinoline-1-oxide/toxicity , Cell Line, Tumor , Dendritic Cells/drug effects , Dendritic Cells/pathology , Epithelial Cells/metabolism , Epithelium/drug effects , Epithelium/pathology , Gene Expression Regulation, Neoplastic/drug effects , Head and Neck Neoplasms/genetics , Head and Neck Neoplasms/immunology , Head and Neck Neoplasms/pathology , Histones/metabolism , Humans , Immunity/drug effects , Langerhans Cells/drug effects , Phagocytes/drug effects , Phagocytes/metabolism , Phagocytes/pathology , Quinolones/toxicity , Squamous Cell Carcinoma of Head and Neck/genetics , Squamous Cell Carcinoma of Head and Neck/immunology , Squamous Cell Carcinoma of Head and Neck/pathology , T-Lymphocytes, Regulatory/drug effects , T-Lymphocytes, Regulatory/immunology , Tongue/pathology , Transcriptome/genetics
3.
Biochem Biophys Res Commun ; 586: 100-106, 2022 01 01.
Article in English | MEDLINE | ID: mdl-34837833

ABSTRACT

Lipopolysaccharide (LPS) is the principal component of the outer membrane of gram-negative bacteria. The prior oral administration of LPS attenuates inflammatory responses, such as intestinal injury and atopic dermatitis, in mouse models; however, the underlying mechanism remains unclear. Here, we examined the effect of topical LPS application on allergic contact dermatitis and its mechanism of action using a murine contact hypersensitivity (CHS) model. Prolonged LPS application to the skin significantly suppressed 2,4-dinitrofluorobenzene (DNFB)-induced CHS. LPS application to the skin also reduced the phagocytosis of fluorescein isothiocyanate (FITC)-dextran by Langerhans and dendritic cells. Cutaneous cell migration into the skin-draining lymph nodes (LNs) induced by FITC painting was reduced by LPS application. During the CHS response, DNFB application induced T-cell proliferation and inflammatory cytokine production in skin-draining LNs, whereas prolonged LPS application inhibited DNFB-induced T-cell growth and interferon gamma production, indicating suppression of DNFB-induced sensitization. These results suggest that prolonged LPS application suppressed DNFB-induced sensitization and subsequently CHS response. Our findings imply that topical application of LPS may prevent allergic dermatitis such as CHS.


Subject(s)
Dermatitis, Contact/drug therapy , Immunologic Factors/pharmacology , Lipopolysaccharides/pharmacology , Lymphocytes/drug effects , Skin/drug effects , Administration, Cutaneous , Animals , Cell Line , Cell Movement/drug effects , Dendritic Cells/cytology , Dendritic Cells/drug effects , Dendritic Cells/immunology , Dermatitis, Contact/etiology , Dermatitis, Contact/immunology , Dermatitis, Contact/pathology , Dextrans/metabolism , Dinitrofluorobenzene/administration & dosage , Ear , Fluorescein-5-isothiocyanate/analogs & derivatives , Fluorescein-5-isothiocyanate/metabolism , Keratinocytes/cytology , Keratinocytes/drug effects , Keratinocytes/immunology , Langerhans Cells/cytology , Langerhans Cells/drug effects , Langerhans Cells/immunology , Lymph Nodes/cytology , Lymph Nodes/drug effects , Lymph Nodes/immunology , Lymphocyte Activation/drug effects , Lymphocytes/cytology , Lymphocytes/immunology , Mice , Mice, Inbred C57BL , Phagocytosis/drug effects , Primary Cell Culture , Skin/immunology , Skin/pathology
4.
Nat Immunol ; 22(12): 1538-1550, 2021 12.
Article in English | MEDLINE | ID: mdl-34795444

ABSTRACT

The signals driving the adaptation of type 2 dendritic cells (DC2s) to diverse peripheral environments remain mostly undefined. We show that differentiation of CD11blo migratory DC2s-a DC2 population unique to the dermis-required IL-13 signaling dependent on the transcription factors STAT6 and KLF4, whereas DC2s in lung and small intestine were STAT6-independent. Similarly, human DC2s in skin expressed an IL-4 and IL-13 gene signature that was not found in blood, spleen and lung DCs. In mice, IL-13 was secreted homeostatically by dermal innate lymphoid cells and was independent of microbiota, TSLP or IL-33. In the absence of IL-13 signaling, dermal DC2s were stable in number but remained CD11bhi and showed defective activation in response to allergens, with diminished ability to support the development of IL-4+GATA3+ helper T cells (TH), whereas antifungal IL-17+RORγt+ TH cells were increased. Therefore, homeostatic IL-13 fosters a noninflammatory skin environment that supports allergic sensitization.


Subject(s)
Cell Communication , Cell Differentiation , Interleukin-13/metabolism , Langerhans Cells/metabolism , Skin/metabolism , Th17 Cells/metabolism , Th2 Cells/metabolism , Allergens/pharmacology , Animals , CD11b Antigen/genetics , CD11b Antigen/metabolism , Cells, Cultured , Databases, Genetic , Humans , Interleukin-13/genetics , Langerhans Cells/drug effects , Langerhans Cells/immunology , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Knockout , Phenotype , STAT6 Transcription Factor/genetics , STAT6 Transcription Factor/metabolism , Signal Transduction , Skin/cytology , Skin/drug effects , Skin/immunology , Th17 Cells/drug effects , Th17 Cells/immunology , Th2 Cells/drug effects , Th2 Cells/immunology , Transcriptome
5.
Int J Mol Sci ; 22(13)2021 Jun 25.
Article in English | MEDLINE | ID: mdl-34201952

ABSTRACT

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.


Subject(s)
Cellular Senescence/drug effects , Flavonoids/pharmacology , Skin Aging/drug effects , Skin/drug effects , Aging/drug effects , Aging/genetics , Aging/metabolism , Animals , Flavonoids/chemistry , Flavonoids/therapeutic use , Humans , Keratinocytes/drug effects , Keratinocytes/metabolism , Langerhans Cells/drug effects , Langerhans Cells/metabolism , Skin/metabolism , Skin Aging/genetics
6.
Carbohydr Polym ; 268: 118211, 2021 Sep 15.
Article in English | MEDLINE | ID: mdl-34127215

ABSTRACT

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.


Subject(s)
Langerhans Cells/drug effects , Needles , Panax notoginseng/chemistry , Polysaccharides/chemistry , Administration, Cutaneous , Animals , B7-1 Antigen/metabolism , B7-2 Antigen/metabolism , CD40 Antigens/metabolism , Compressive Strength , Doxorubicin/administration & dosage , Fluorescein/administration & dosage , Fluorouracil/administration & dosage , Langerhans Cells/metabolism , Male , Mice , Myosin Heavy Chains/metabolism , Rats, Sprague-Dawley , Skin/cytology , Skin/drug effects , Skin/metabolism , Solubility
7.
Cell ; 184(8): 2151-2166.e16, 2021 04 15.
Article in English | MEDLINE | ID: mdl-33765440

ABSTRACT

Cutaneous mast cells mediate numerous skin inflammatory processes and have anatomical and functional associations with sensory afferent neurons. We reveal that epidermal nerve endings from a subset of sensory nonpeptidergic neurons expressing MrgprD are reduced by the absence of Langerhans cells. Loss of epidermal innervation or ablation of MrgprD-expressing neurons increased expression of a mast cell gene module, including the activating receptor, Mrgprb2, resulting in increased mast cell degranulation and cutaneous inflammation in multiple disease models. Agonism of MrgprD-expressing neurons reduced expression of module genes and suppressed mast cell responses. MrgprD-expressing neurons released glutamate which was increased by MrgprD agonism. Inhibiting glutamate release or glutamate receptor binding yielded hyperresponsive mast cells with a genomic state similar to that in mice lacking MrgprD-expressing neurons. These data demonstrate that MrgprD-expressing neurons suppress mast cell hyperresponsiveness and skin inflammation via glutamate release, thereby revealing an unexpected neuroimmune mechanism maintaining cutaneous immune homeostasis.


Subject(s)
Glutamic Acid/metabolism , Mast Cells/metabolism , Neurons/metabolism , Skin/metabolism , Animals , Cells, Cultured , Dermatitis/metabolism , Dermatitis/pathology , Diphtheria Toxin/pharmacology , Disease Models, Animal , Female , Integrin beta Chains/genetics , Integrin beta Chains/metabolism , Langerhans Cells/cytology , Langerhans Cells/drug effects , Langerhans Cells/metabolism , Mast Cells/cytology , Mast Cells/drug effects , Mice , Mice, Inbred C57BL , Mice, Knockout , Neurons/cytology , Receptors, G-Protein-Coupled/agonists , Receptors, G-Protein-Coupled/deficiency , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/metabolism , Skin/pathology , beta-Alanine/chemistry , beta-Alanine/metabolism , beta-Alanine/pharmacology
8.
Contact Dermatitis ; 83(5): 337-346, 2020 Nov.
Article in English | MEDLINE | ID: mdl-32677096

ABSTRACT

BACKGROUND: The nature of clinically related adverse reactions to titanium is still unknown. OBJECTIVE: To determine whether titanium salts have irritant or sensitizing potential in a reconstructed human skin (RHS) model with integrated Langerhans cells (LCs). METHODS: RHS-LCs (ie, reconstructed epidermis) containing primary differentiated keratinocytes and CFSE+ CD1a+ -LCs generated from the MUTZ-3 cell line on a primary fibroblast-populated collagen hydrogel (dermis) were topically exposed to titanium(IV) bis(ammonium lactato)dihydroxide (TiALH). LC migration and plasticity were determined. RESULTS: TiALH resulted in CFSE+ CD1a+ -LC migration out of the epidermis. Neutralizing antibodies to CCL5 and CXCL12 showed that LC migration was CCL5 and not CXCL12 mediated. LCs accumulating within the dermis after TiALH exposure were CFSE+ Lang+ CD68+ which is characteristic of a phenotypic switch of MUTZ-LC to a macrophage-like cell. Furthermore, TiALH did not result in increased interleukin (IL)-1ß or CCR7 messenger RNA (mRNA) in the dermis, but did result in increased IL-10 mRNA. In addition, monocultures of MUTZ-LCs failed to increase LC maturation biomarkers CD83, CD86, and CXCL-8 when exposed to noncytotoxic concentrations of four different titanium salts. CONCLUSION: These results classify titanium salts as irritants rather than sensitizers and indicate that titanium implant-related complaints could be due to localized irritant-mediated inflammation arising from leachable agents rather than a titanium metal allergy.


Subject(s)
Dermatitis, Allergic Contact/metabolism , Irritants/pharmacology , Langerhans Cells/drug effects , Titanium/pharmacology , Cell Differentiation/drug effects , Cell Line/drug effects , Dermis/metabolism , Epidermis/metabolism , Humans
9.
J Pharm Pharm Sci ; 23(1): 86-99, 2020.
Article in English | MEDLINE | ID: mdl-32338591

ABSTRACT

BACKGROUND: Langerhans cells (LCs) polarize the immune milieu towards a T helper type (Th) 1 or Th2 immune response. We investigated the effects of selected tetracyclines on Th cells development mediated by LCs, and their implications for the treatment of atopic dermatitis (AD). METHODS: Mice were primed with ovalbumin (OVA) peptide-pulsed LCs, which had been treated with each antibiotic, via the hind footpad. After 5 days, the Th1/Th2 cytokine response in the popliteal lymph nodes was investigated by enzyme-linked immunosorbent assay. The expression of cell surface molecules on LCs was investigated using reverse transcriptase polymerase chain reaction. The therapeutic effects of a selected antibiotic on AD-like skin lesions of NC/Nga mice were assessed in terms of the skin severity score, histological changes in the lesioned skin, the serum level of total IgE, and expression of Th1/Th2 cytokines in lymph nodes and skin lesions. RESULTS: Antibiotic-treated, OVA peptide-pulsed LCs inhibited development of Th2 cells but not Th1 cells. This was accompanied by suppression of T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression in LCs. Doxycycline had the greatest activity against Staphylococcus aureus strains isolated from skin lesions of patients with AD, and a strong inhibitory effect on Th2 cell development. Doxycycline suppressed the increase in the skin severity score during the acute phase in NC/Nga mice similar to betamethasone. This suppressive effect was associated with a decrease in the serum IgE level and production of Th2 cytokines in auricular lymph node cells and skin lesions. CONCLUSION: Topical application of doxycycline to AD lesions would act on both superficial S. aureus colonization and epidermal LCs, thus possibly inhibiting the development of Th2 cells in vivo, with benefits for control of acute inflammation in AD.


Subject(s)
Anti-Allergic Agents/therapeutic use , Anti-Bacterial Agents/therapeutic use , Dermatitis, Atopic/drug therapy , Langerhans Cells/drug effects , Tetracyclines/therapeutic use , Th2 Cells/drug effects , Administration, Topical , Allergens , Animals , Anti-Allergic Agents/pharmacology , Anti-Bacterial Agents/pharmacology , Dermatitis, Atopic/chemically induced , Dermatitis, Atopic/immunology , Dermatitis, Atopic/pathology , Immunoglobulin E/blood , Langerhans Cells/immunology , Lymph Nodes/drug effects , Mice, Inbred BALB C , Mice, Transgenic , Ovalbumin , Picryl Chloride , Skin/drug effects , Skin/pathology , Staphylococcus aureus/drug effects , Tetracyclines/pharmacology , Th1 Cells/drug effects , Th1 Cells/immunology , Th2 Cells/immunology
11.
Autophagy ; 16(8): 1380-1395, 2020 08.
Article in English | MEDLINE | ID: mdl-31668121

ABSTRACT

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.


Subject(s)
Adrenergic beta-Antagonists/pharmacology , Interleukin-23/biosynthesis , Langerhans Cells/metabolism , Lysosomes/metabolism , Oxidative Stress/drug effects , Autophagy/drug effects , Cell Differentiation/drug effects , Chloroquine/pharmacology , Dendritic Cells/drug effects , Dendritic Cells/metabolism , Endosomes/drug effects , Endosomes/metabolism , Gene Expression Regulation/drug effects , Humans , Inflammasomes/metabolism , Inflammation Mediators/metabolism , Interleukin-12/genetics , Interleukin-12/metabolism , Langerhans Cells/drug effects , Lysosomal-Associated Membrane Protein 1/metabolism , Lysosomes/drug effects , Mitochondria/drug effects , Mitochondria/metabolism , Mitogen-Activated Protein Kinase 14/metabolism , NF-kappa B/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Propranolol/pharmacology , Reactive Oxygen Species/metabolism , Receptors, Adrenergic, beta/metabolism , Signal Transduction , Th17 Cells/cytology , Th17 Cells/drug effects , Ubiquitinated Proteins/metabolism
12.
Eur. j. anat ; 23(5): 383-388, sept. 2019. ilus, graf, tab
Article in English | IBECS | ID: ibc-183869

ABSTRACT

Langerhans cells (LCs) are the most effective antigen presenting cells against foreign bodies and carcinogens. Since the oral cavity is a portal of entry for these antigens, the aim of this study was to morphologically classify CD1a+ LCs, quantify them in the normal and malignant buccal mucosa, and evaluate their relation to the age of patients. Healthy buccal mucosal samples collected from 16 patients undergoing reconstructive operation, and malignant samples obtained from 15 patients undergoing radical oncological resection. were processed for immunohistochemistry four- to fivemicron thick sections were stained with CD1a antibody (CD1a). At 40X magnification, CD1a+ LCs were morphologically classified and quantified manually for a 25mm length of basement membrane using Cellsens image analysing software and the data was analysed. Two categories of CD1a+ LCs were identified in the normal and malignant buccal mucosa a) typical dendritic LCs and b) non-dendritic LCs (a new entity). Non-dendritic LCs were of significantly higher number compared to the typical dendritic LCs in the normal tissues (p -0.001). In the malignant group, the non-dendritic CD1a+ LCs were significantly fewer in number (p0.004), when compared to the normal group. Nondendritic LCs were also significantly fewer (p0.026) in patients over 60 years of age. This is the first report of non-dendritic Langerhans cells in normal buccal mucosa and malignant buccal mucosa using the CD1a marker. The significantly higher number of these cells in normal tissues and younger individuals supports their role as accessory antigen presenting cells


No disponible


Subject(s)
Humans , Male , Female , Adult , Middle Aged , Langerhans Cells/pathology , Mouth Mucosa/anatomy & histology , Mouth Mucosa/pathology , Langerhans Cells/drug effects , Mouth Mucosa/cytology , Antigens, CD1
13.
Toxicol Lett ; 314: 172-180, 2019 Oct 10.
Article in English | MEDLINE | ID: mdl-31404593

ABSTRACT

Vesicants cause a multitude of cutaneous reactions like erythema, blisters and ulcerations. After exposure to sulfur mustard (SM) and related compounds, patients present dermal symptoms typically known for chemicals categorized as skin sensitizer (e.g. hypersensitivity and flare-up phenomena). However, although some case reports led to the assumption that SM and other alkylating compounds represent sensitizers, a comprehensive investigation of SM-triggered immunological responses has not been conducted so far. Based on a well-structured system of in chemico and in vitro test methods, the Organization for Economic Co-operation and Development (OECD) established procedures to categorize agents on their skin sensitizing abilities. In this study, the skin sensitizing potential of SM and three related alkylating agents (AAs) was assessed following the OECD test guidelines. Besides SM, investigated AAs were chlorambucil (CHL), nitrogen mustard (HN3) and 2-chloroethyl ethyl sulfide (CEES). The methods are described in detail in the EURL ECVAM DataBase service on ALternative Methods to animal experimentation (DB-ALM). In accordance to OECD recommendations, skin sensitization is a pathophysiological process starting with a molecular initiating step and ending with the in vivo outcome of an allergic contact dermatitis. This concept is called adverse outcome pathway (AOP). An AOP links an adverse outcome to various key events which can be assayed by established in chemico and in vitro test methods. Positive outcome in two out of three key events indicates that the chemical can be categorized as a skin sensitizer. In this study, key event 1 "haptenation" (covalent modification of epidermal proteins), key event 2 "activation of epidermal keratinocytes" and key event 3 "activation of dendritic cells" were investigated. Covalent modification of epidermal proteins measured by using the DPRA-assay provided distinct positive results for all tested substances. Same outcome was seen in the KeratinoSens assay, investigating the activation of epidermal keratinocytes. The h-CLAT assay performed to determine the activation of dendritic cells provided positive results for SM and CEES but not for CHL and HN3. Altogether, following OECD requirements, our results suggest the classification of all investigated substances as skin sensitizers. Finally, a tentative AOP for SM-induced skin sensitization is suggested.


Subject(s)
Chemical Warfare Agents/toxicity , Irritants/toxicity , Mustard Gas/toxicity , Skin Irritancy Tests/standards , Skin/drug effects , Biomarkers/metabolism , Chemical Warfare Agents/classification , Chlorambucil/classification , Chlorambucil/toxicity , Guidelines as Topic , Humans , Irritants/classification , Keratinocytes/drug effects , Keratinocytes/immunology , Keratinocytes/metabolism , Langerhans Cells/drug effects , Langerhans Cells/immunology , Langerhans Cells/metabolism , Mechlorethamine/classification , Mechlorethamine/toxicity , Mustard Gas/analogs & derivatives , Mustard Gas/classification , Risk Assessment , Skin/immunology , Skin/metabolism
14.
J Invest Dermatol ; 139(10): 2164-2173.e1, 2019 10.
Article in English | MEDLINE | ID: mdl-30978356

ABSTRACT

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.


Subject(s)
Dermatitis, Allergic Contact/drug therapy , Dermatitis, Allergic Contact/immunology , Haptens/pharmacology , Langerhans Cells/drug effects , Programmed Cell Death 1 Ligand 2 Protein/genetics , RNA, Small Interfering/pharmacology , Administration, Topical , Animals , Biopsy, Needle , Disease Models, Animal , Female , Flow Cytometry/methods , Gene Silencing/drug effects , Immunohistochemistry , Japan , Langerhans Cells/immunology , Mice , Mice, Inbred BALB C , Mice, Knockout , Random Allocation , Reference Values , Risk Factors , Statistics, Nonparametric , Treatment Outcome
15.
J Pharm Pharm Sci ; 22(1): 122-130, 2019 04 11.
Article in English | MEDLINE | ID: mdl-30974054

ABSTRACT

BACKGROUND: It is widely acknowledged that Langerhans cells (LCs) play a primary role in the polarization of T helper type 1 (Th1) or T helper type 2 (Th2) immune responses. Our aim was to find fluoroquinolone ("new quinolone") antibiotics that would inhibit LC-mediated Th2 cell development. METHODS: Expression of LC surface molecules was investigated using the reverse transcriptase polymerase chain reaction. The effects of fluoroquinolone antibiotics on T-cell immunoglobulin and mucin domain-containing protein (TIM)-4 expression in LCs were examined to predict whether they would inhibit Th2 cell development. Mice were primed via the hind footpad with ovalbumin (OVA) peptide-pulsed LCs that had been treated with a selected fluoroquinolone antibiotic, then 5 days later the cytokine response in popliteal lymph nodes was examined by enzyme-linked immunosorbent assay. RESULTS: Norfloxacin was selected as a candidate inhibitor of Th2 cell development. As expected, OVA peptide-pulsed LCs that had been treated with norfloxacin and injected into the hind footpads of mice inhibited Th2 cell development, as represented by down-regulation of interleukin (IL)-4 production, as well as Th1 cell development, as represented by down-regulation of interferon (IFN)- g production. This additional inhibition of Th1 cell development was accompanied by suppression of CD40 expression in LCs. In addition, Staphylococcus aureus strains isolated from skin lesions of patients with atopic dermatitis (AD) were more susceptible to norfloxacin than to gentamicin. Topical treatment with norfloxacin significantly suppressed the increase in the skin severity score in NC/Nga mice with AD-like skin lesions. This suppressive effect was associated with a decrease in the production of IFN-g and IL-4 in auricular lymph node cells. CONCLUSIONS: The present results show that topical application of norfloxacin inhibits the development of AD-like skin lesions in NC/Nga mice. This suggests that topical application of norfloxacin to AD lesions colonized with S. aureus would act on both superficial S. aureus and epidermal LCs, thus possibly inhibiting the development of Th1 and Th2 cells in vivo, and controlling the severity of AD.


Subject(s)
Anti-Bacterial Agents/pharmacology , Fluoroquinolones/pharmacology , Langerhans Cells/drug effects , Norfloxacin/pharmacology , Th1 Cells/drug effects , Th2 Cells/drug effects , Animals , Cytokines/immunology , Down-Regulation/drug effects , Down-Regulation/immunology , Langerhans Cells/immunology , Mice , Mice, Inbred BALB C , Th1 Cells/immunology , Th2 Cells/immunology
16.
J Immunol Res ; 2019: 5143635, 2019.
Article in English | MEDLINE | ID: mdl-30944833

ABSTRACT

Ideal agents for the topical treatment of skin wounds should have antimicrobial efficacy without negative influence on wound healing. Octenidine (OCT) has become a widely used antiseptic in professional wound care, but its influence on several components of the wound healing process remains unclear. In the present study, we have used a superficial wound model using tape stripping on human full-thickness skin ex vivo to investigate the influence of OCT on epidermal Langerhans cells (LCs) and cytokine secretion pattern of skin cells during wound healing in a model without disruption of the normal skin structure. Histological and immunofluorescence studies showed that OCT neither altered human skin architecture nor the viability of skin cells upon 48 hours of culture in unwounded or wounded skin. The epidermis of explants and LCs remained morphologically intact throughout the whole culture period upon OCT treatment. OCT inhibited the upregulation of the maturation marker CD83 on LCs and prevented their emigration in wounded skin. Furthermore, OCT reduced both pro- and anti-inflammatory mediators (IL-8, IL-33, and IL-10), while angiogenesis and growth factor mediators (VEGF and TGF-ß1) remained unchanged in skin explant cultures. Our data provide novel insights into the host response to OCT in the biologically relevant environment of viable human (wounded) skin.


Subject(s)
Anti-Infective Agents/pharmacology , Cytokines/genetics , Epidermis/drug effects , Langerhans Cells/drug effects , Langerhans Cells/immunology , Pyridines/pharmacology , Wound Healing/drug effects , Adult , Cytokines/immunology , Epidermal Cells/drug effects , Epidermal Cells/immunology , Humans , Imines , Middle Aged , Models, Biological , Skin/drug effects , Skin/immunology , Skin/pathology , Surgical Tape , Wound Healing/immunology , Wounds and Injuries/drug therapy , Wounds and Injuries/immunology , Young Adult
17.
Immunol Cell Biol ; 97(8): 700-713, 2019 09.
Article in English | MEDLINE | ID: mdl-30989674

ABSTRACT

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.


Subject(s)
Antigen Presentation/immunology , Immunosuppressive Agents/therapeutic use , Langerhans Cells/immunology , Skin Diseases/immunology , Skin/immunology , Animals , Antigens, Bacterial/immunology , Antigens, Fungal/immunology , Antigens, Neoplasm/immunology , Antigens, Viral/immunology , Disease Models, Animal , Eosinophils/immunology , Humans , Immunosuppressive Agents/pharmacology , Langerhans Cells/drug effects , Skin/microbiology , Skin/pathology , Skin Diseases/drug therapy , Skin Diseases/microbiology , Skin Diseases/pathology , Wound Healing/immunology
18.
J Cutan Med Surg ; 23(4): 370-379, 2019.
Article in English | MEDLINE | ID: mdl-31010295

ABSTRACT

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.


Subject(s)
Aminolevulinic Acid/pharmacology , Condylomata Acuminata/drug therapy , Epidermis/immunology , Langerhans Cells/immunology , Photochemotherapy , Photosensitizing Agents/pharmacology , Adult , Aminolevulinic Acid/therapeutic use , Antigens, CD1/metabolism , CD3 Complex/metabolism , CD4 Lymphocyte Count , CD8-Positive T-Lymphocytes/immunology , Cytokines/genetics , Epidermis/metabolism , Female , Gene Expression/drug effects , Humans , Interferon Regulatory Factor-7/genetics , Interferon-alpha/genetics , Interferon-beta/genetics , Interferon-gamma/genetics , Interleukin-3 Receptor alpha Subunit/metabolism , Langerhans Cells/drug effects , Langerhans Cells/metabolism , Male , Middle Aged , Myxovirus Resistance Proteins/genetics , Photosensitizing Agents/therapeutic use , RNA, Messenger/metabolism , Toll-Like Receptor 9/genetics , Ubiquitins/genetics , Young Adult
19.
Int J Mol Sci ; 20(6)2019 Mar 19.
Article in English | MEDLINE | ID: mdl-30893757

ABSTRACT

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.


Subject(s)
Langerhans Cells/cytology , Lipopolysaccharides/pharmacology , Lymphocyte Activation/immunology , Monocytes/cytology , Peptidoglycan/pharmacology , Th17 Cells/immunology , Antigens, CD/metabolism , CD40 Ligand/metabolism , Cytokines/metabolism , DNA/metabolism , Humans , Interleukins/metabolism , Langerhans Cells/drug effects , Langerhans Cells/metabolism , Lymphocyte Activation/drug effects , Monocytes/drug effects , Monocytes/metabolism , Poly I-C/pharmacology , Poly U/pharmacology , Toll-Like Receptors/agonists , Toll-Like Receptors/metabolism , Up-Regulation/drug effects
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