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1.
Lasers Surg Med ; 56(1): 100-106, 2024 Jan.
Article in English | MEDLINE | ID: mdl-37855626

ABSTRACT

PURPOSE: In postoperative wound healing after surgical operations or ablative laser treatments, recent studies suggest the timely use of non-ablative fractional laser treatments with the aim to improve wound healing and prevent pathological scar formation. However, the underlying molecular mechanisms are poorly understood. The aim of this study was to investigate the effects of laser-assisted scar healing (LASH) at the molecular level and to combine it with already established wound healing-promoting local treatments. METHODS: We irradiated full-thickness 3D skin models with a fractional ablative Er:YAG laser to set standardized lesions to the epidermal and upper dermal layer. Subsequently, LASH was induced by irradiating the models with either a fractional non-ablative 1540 nm Er:Glass or 1550 nm diode laser. In addition, we tested the combination of non-ablative fractional laser treatment and topical aftercare with a dexpanthenol-containing ointment (DCO). RESULTS: Histological analysis revealed that models irradiated with the 1540 nm Er:Glass or 1550 nm diode laser exhibited accelerated but not complete wound closure after 16 h. In contrast, additional topical posttreatment with DCO resulted in complete wound closure. At gene expression level, both non-ablative laser systems showed similar effects on epidermal differentiation and mild anti-inflammatory properties. The additional posttreatment with DCO enhanced the wound-healing effects of LASH, especially the upregulation of epidermal differentiation markers and anti-inflammatory cytokines at the gene expression level. CONCLUSION: This in vitro study deciphers the biological effects of LASH with a fractional non-ablative 1540 nm Er:Glass or a 1550 nm diode laser in 3D skin models. These data help to better understand the biological properties of the LASH technique and is important to optimize its application.


Subject(s)
Laser Therapy , Lasers, Solid-State , Humans , Cicatrix/metabolism , Lasers, Semiconductor/therapeutic use , Skin/metabolism , Wound Healing , Lasers, Solid-State/therapeutic use , Anti-Inflammatory Agents/metabolism , Anti-Inflammatory Agents/pharmacology , Laser Therapy/methods
2.
Proc Natl Acad Sci U S A ; 114(13): E2748-E2757, 2017 03 28.
Article in English | MEDLINE | ID: mdl-28289191

ABSTRACT

The proinflammatory cytokine IL-36γ is highly expressed in epithelial cells and is a pivotal mediator of epithelial inflammation. In particular, IL-36γ is strongly associated with the inflammatory skin disease psoriasis. As with other IL-1 cytokines, IL-36γ is expressed as an inactive precursor and must be processed by specific proteases to become bioactive. Our aim therefore was to identify protease/s capable of IL-36γ activation and explore the importance of this activation in psoriasis. Using a keratinocyte-based activity assay in conjunction with small-molecule inhibitors and siRNA gene silencing, cathepsin S was identified as the major IL-36γ-activating protease expressed by epithelial cells. Interestingly, cathepsin S activity was strongly up-regulated in samples extracted from psoriasis patients relative to healthy controls. In addition, IL-36γ-Ser18, identified as the main product of cathepsin S-dependent IL-36γ cleavage, induced psoriasiform changes in human skin-equivalent models. Together, these data provide important mechanistic insights into the activation of IL-36γ and highlight that cathepsin S-mediated activation of IL-36γ may be important in the development of numerous IL-36γ-driven pathologies, in addition to psoriasis.


Subject(s)
Cathepsins/metabolism , Interleukin-1/genetics , Psoriasis/genetics , Amino Acid Motifs , Cathepsins/genetics , Cell Line , Epithelial Cells/enzymology , Epithelial Cells/metabolism , Humans , Inflammation Mediators/metabolism , Interleukin-1/metabolism , Psoriasis/enzymology , Psoriasis/immunology
3.
Exp Dermatol ; 27(9): 1009-1014, 2018 09.
Article in English | MEDLINE | ID: mdl-29851147

ABSTRACT

Atopic dermatitis (AD) is a chronically relapsing, pruritic inflammation of the skin with dryness and disturbed skin barrier function. Recently, we established that IL-31 treatment of human 3D skin models resulted in a disrupted skin barrier phenotype resembling AD. In this model, we found that IL-31 interferes with the differentiation of keratinocytes and inhibits the expression of terminal differentiation markers. In the present study, we investigated the effects of a ceramide-containing water-in-oil skin care ointment on the physical skin barrier structure and function in disrupted skin barrier models, generated either by using primary normal human epidermal keratinocytes (NHEK) or HaCaT cells. We observed that the physical skin barrier of the models recovered after daily topical treatment with the ceramide-containing ointment. Topical application of the ointment prevented downregulation of filaggrin and disorganization of other differentiation markers, such as keratin 10 and ß4-integrin, as demonstrated by immunohistological analysis. The expression of Ki67 was also upregulated in response to the ointment. Furthermore, functional studies revealed that local application of the ointment diminished the increased uptake of fluorescently labelled recombinant allergens of timothy grass (phl p1) in our model. In conclusion, our data revealed that topical application of a ceramide-containing skin care ointment reduced IL-31 induced impairments of the physical skin barrier and skin barrier function in an in vitro model of the disrupted skin barrier. This standardized model can be utilized in the future to monitor ex vivo effects of various topical therapies on skin morphology, physiology, and gene expression.


Subject(s)
Ceramides/pharmacology , Dermatologic Agents/pharmacology , Interleukins/pharmacology , Skin Physiological Phenomena/drug effects , Water Loss, Insensible/drug effects , Bioartificial Organs , Cell Differentiation/drug effects , Cell Line , Fibroblasts/metabolism , Filaggrin Proteins , Humans , Keratinocytes/metabolism , Ointment Bases , Ointments , Recombinant Proteins/pharmacology , Water/metabolism
4.
FASEB J ; 31(2): 526-543, 2017 02.
Article in English | MEDLINE | ID: mdl-27825106

ABSTRACT

The response of the skin to harmful environmental agents is shaped decisively by the status of the immune system. Keratinocytes constitutively express and secrete the chemokine-like mediator, macrophage migration inhibitory factor (MIF), more strongly than dermal fibroblasts, thereby creating a MIF gradient in skin. By using global and epidermis-restricted Mif-knockout (Mif-/- and K14-Cre+/tg; Miffl/fl) mice, we found that MIF both recruits and maintains antigen-presenting cells in the dermis/epidermis. The reduced presence of antigen-presenting cells in the absence of MIF was associated with accelerated and increased formation of nonmelanoma skin tumors during chemical carcinogenesis. Our results demonstrate that MIF is essential for maintaining innate immunity in skin. Loss of keratinocyte-derived MIF leads to a loss of control of epithelial skin tumor formation in chemical skin carcinogenesis, which highlights an unexpected tumor-suppressive activity of MIF in murine skin.-Brocks, T., Fedorchenko, O., Schliermann, N., Stein, A., Moll, U. M., Seegobin, S., Dewor, M., Hallek, M., Marquardt, Y., Fietkau, K., Heise, R., Huth, S., Pfister, H., Bernhagen, J., Bucala, R., Baron, J. M., Fingerle-Rowson, G. Macrophage migration inhibitory factor protects from nonmelanoma epidermal tumors by regulating the number of antigen-presenting cells in skin.


Subject(s)
Macrophage Migration-Inhibitory Factors/metabolism , Skin Neoplasms/chemically induced , Skin/cytology , Skin/immunology , Animals , Anthracenes/toxicity , Antigens, CD/genetics , Antigens, CD/metabolism , Carcinogenesis , Gene Expression Regulation/physiology , Inflammation/metabolism , Keratinocytes/metabolism , Macrophage Migration-Inhibitory Factors/genetics , Mice , Mice, Inbred BALB C , Mice, Knockout , Mice, Transgenic , Piperidines/toxicity , Pyridines/toxicity , Receptors, CXCR/genetics , Receptors, CXCR/metabolism
5.
J Immunol ; 196(8): 3233-44, 2016 Apr 15.
Article in English | MEDLINE | ID: mdl-26944931

ABSTRACT

Atopic dermatitis, a chronic inflammatory skin disease with increasing prevalence, is closely associated with skin barrier defects. A cytokine related to disease severity and inhibition of keratinocyte differentiation is IL-31. To identify its molecular targets, IL-31-dependent gene expression was determined in three-dimensional organotypic skin models. IL-31-regulated genes are involved in the formation of an intact physical skin barrier. Many of these genes were poorly induced during differentiation as a consequence of IL-31 treatment, resulting in increased penetrability to allergens and irritants. Furthermore, studies employing cell-sorted skin equivalents in SCID/NOD mice demonstrated enhanced transepidermal water loss following s.c. administration of IL-31. We identified the IL-1 cytokine network as a downstream effector of IL-31 signaling. Anakinra, an IL-1R antagonist, blocked the IL-31 effects on skin differentiation. In addition to the effects on the physical barrier, IL-31 stimulated the expression of antimicrobial peptides, thereby inhibiting bacterial growth on the three-dimensional organotypic skin models. This was evident already at low doses of IL-31, insufficient to interfere with the physical barrier. Together, these findings demonstrate that IL-31 affects keratinocyte differentiation in multiple ways and that the IL-1 cytokine network is a major downstream effector of IL-31 signaling in deregulating the physical skin barrier. Moreover, by interfering with IL-31, a currently evaluated drug target, we will have to consider that low doses of IL-31 promote the antimicrobial barrier, and thus a complete inhibition of IL-31 signaling may be undesirable.


Subject(s)
Dermatitis, Atopic/pathology , Interleukin-1alpha/metabolism , Interleukin-1beta/metabolism , Interleukins/metabolism , Tight Junctions/metabolism , Animals , Antimicrobial Cationic Peptides/biosynthesis , Cell Differentiation/drug effects , Cell Differentiation/physiology , Cell Line , Filaggrin Proteins , Humans , Interleukin 1 Receptor Antagonist Protein/pharmacology , Interleukins/pharmacology , Intermediate Filament Proteins/metabolism , Keratinocytes/cytology , Mice , Mice, Inbred NOD , Mice, SCID , Receptors, Interleukin-1/antagonists & inhibitors , Signal Transduction/physiology , Skin/cytology , Skin/growth & development , Tight Junctions/drug effects
6.
Environ Res ; 164: 221-228, 2018 07.
Article in English | MEDLINE | ID: mdl-29501832

ABSTRACT

Polychlorinated biphenyls (PCB) are well known persistent and toxic environmental pollutants. Our aim was to identify effects of moderate-high exposure to dioxin-like (dl) and non-dioxin-like (ndl)-PCBs on the skin in order to provide more insight in the pathophysiological effects of these compounds. We performed a dermatological examination on 92 former workers from a transformer recycling company with known elevated serum PCB and/or dioxin (polychlorinated dibenzo-p-dioxin/polychlorinated dibenzo-p-furan (PCDD/F)) levels. In addition, we performed a skin cancer screening over a period of seven years (2010-2016) on resp. 268, 271, 210, 149, 92, 129 and 79 participants. We found a higher incidence of acne and malignancies of the skin (malignant melanoma, basal cell carcinoma and mycosis fungoides) in the workers compared to normal population. The probability of having hyperpigmentation on the skin was statistically significantly higher in workers with higher sumPCBs- (OR:1.09(1.12-2.17)), dioxin-like (dl)-PCBs- (OR:1.56(1.12-2.17)) and dioxin (PCDD/Fs) (OR:1.09(1.02-1.16)) levels. Age was a confounding factor in this model. Formation of hyperpigmentation could be an indicator for (moderate-high) exposure to toxic compounds like PCBs. The higher incidence of cutaneous malignancies found in the workers might be associated with PCB- and dioxin exposure, warranting further investigation on larger cohorts.


Subject(s)
Benzofurans , Dioxins , Environmental Pollutants , Hyperpigmentation , Polychlorinated Biphenyls , Polychlorinated Dibenzodioxins , Skin Neoplasms , Adult , Aged , Benzofurans/toxicity , Dibenzofurans, Polychlorinated/toxicity , Environmental Pollutants/toxicity , Female , Humans , Hyperpigmentation/epidemiology , Incidence , Male , Middle Aged , Polychlorinated Biphenyls/toxicity , Polychlorinated Dibenzodioxins/toxicity , Skin Neoplasms/epidemiology
7.
Dermatol Surg ; 44(12): 1525-1536, 2018 12.
Article in English | MEDLINE | ID: mdl-30045108

ABSTRACT

BACKGROUND: Immunosuppression is often considered as an indication for antibiotic prophylaxis to prevent surgical site infections (SSI) while performing skin surgery. However, the data on the risk of developing SSI after dermatologic surgery in immunosuppressed patients are limited. PATIENTS AND METHODS: All patients of the Department of Dermatology and Allergology at the University Hospital of RWTH Aachen in Aachen, Germany, who underwent hospitalization for a dermatologic surgery between June 2016 and January 2017 (6 months), were followed up after surgery until completion of the wound healing process. The follow-up addressed the occurrence of SSI and the need for systemic antibiotics after the operative procedure. Immunocompromised patients were compared with immunocompetent patients. The investigation was conducted as a retrospective analysis of patient records. RESULTS: The authors performed 284 dermatologic surgeries in 177 patients. Nineteen percent (54/284) of the skin surgery was performed on immunocompromised patients. The most common indications for surgical treatment were nonmelanoma skin cancer and malignant melanomas. Surgical site infections occurred in 6.7% (19/284) of the cases. In 95% (18/19), systemic antibiotic treatment was needed. Twenty-one percent of all SSI (4/19) were seen in immunosuppressed patients. CONCLUSION: According to the authors' data, immunosuppression does not represent a significant risk factor for SSI after dermatologic surgery. However, larger prospective studies are needed to make specific recommendations on the use of antibiotic prophylaxis while performing skin surgery in these patients.


Subject(s)
Dermatologic Surgical Procedures/adverse effects , Immunocompromised Host , Immunosuppression Therapy/adverse effects , Skin Neoplasms/surgery , Surgical Wound Infection/etiology , Adolescent , Adult , Aged , Aged, 80 and over , Anti-Bacterial Agents/therapeutic use , Child , Child, Preschool , Female , Follow-Up Studies , Humans , Keratosis, Actinic/surgery , Length of Stay , Male , Middle Aged , Retrospective Studies , Risk Factors , Surgical Wound Infection/drug therapy , Young Adult
8.
Dermatol Surg ; 43(7): 928-933, 2017 Jul.
Article in English | MEDLINE | ID: mdl-28272086

ABSTRACT

BACKGROUND: Digital mucoid cysts have a tendency for recurrence after operative intervention. Several procedures are in use. OBJECTIVE: Retrospective evaluation for effectiveness, safety and patient satisfaction by using a questionnaire after treatment for digital mucoid cysts with targeted surgical excision and closure by flap-design. MATERIALS AND METHODS: All patients treated with surgical excision for digital mucoid cysts at the Dermatology Department of the Ludwigshafen City Hospital between 2007 and 2011 were evaluated using a specially designed questionnaire. RESULTS: We evaluated 31 patients. The patient group consisted of 65% women, the median age was 61 years. Seventy-eight percent of patients with nail involvement had a marked improvement or complete resolution of this complaint after surgery. A few complications (e.g., redness, pain or hematoma) were observed after treatment, but no patients required oral antibiotics. Patient evaluation of cosmetic outcome revealed high satisfaction with the procedure, nevertheless recurrence of the digital mucoid cysts was observed in 22.5% of all cases. CONCLUSION: Surgical excision in treatment of digital mucoid cysts was shown to be effective and safe. However, possible advantages and disadvantages of this treatment option should be discussed with the patients before a decision on the kind of therapy is reached.


Subject(s)
Synovial Cyst/surgery , Adolescent , Adult , Aged , Dermatologic Surgical Procedures/adverse effects , Dermatologic Surgical Procedures/methods , Female , Fingers , Humans , Male , Middle Aged , Retrospective Studies , Treatment Outcome , Young Adult
9.
Skin Pharmacol Physiol ; 29(6): 281-290, 2016.
Article in English | MEDLINE | ID: mdl-28013301

ABSTRACT

Tasisulam is a promising antitumor agent with complex pharmacology, which is used as an antiproliferative agent in patients with metastatic melanoma and other solid tumors. Phase 2 melanoma studies showed promising results but had to be stopped because of insufficient tasisulam clearance leading to toxic side effects. To reduce the negative effects of tasisulam, we synthesized a novel sulfonimidamide-based analog to evaluate its antiproliferative effects in comparison to the original compound by performing a cell proliferation assay in melanoma cell lines SKMel23 and A375. The results revealed that the analog had inhibitory effects on the proliferation comparable to tasisulam in both investigated cell lines. These results could contribute to a reduced toxicity of tasisulam and lead to further clinical trials in metastatic melanoma.


Subject(s)
Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Benzamides/chemical synthesis , Benzamides/pharmacology , Sulfonamides/chemical synthesis , Sulfonamides/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Humans , Melanoma
10.
Lasers Med Sci ; 31(3): 397-404, 2016 Apr.
Article in English | MEDLINE | ID: mdl-26796701

ABSTRACT

Clinical experiences with non-ablative fractional erbium glass laser therapy have demonstrated promising results for dermal remodelling and for the indications of striae, surgical scars and acne scars. So far, molecular effects on human skin following treatment with these laser systems have not been elucidated. Our aim was to investigate laser-induced effects on skin morphology and to analyse molecular effects on gene regulation. Therefore, human three-dimensional (3D) organotypic skin models were irradiated with non-ablative fractional erbium glass laser systems enabling qRT-PCR, microarray and histological studies at same and different time points. A decreased mRNA expression of matrix metalloproteinases (MMPs) 3 and 9 was observed 3 days after treatment. MMP3 also remained downregulated on protein level, whereas the expression of other MMPs like MMP9 was recovered or even upregulated 5 days after irradiation. Inflammatory gene regulatory responses measured by the expression of chemokine (C-X-C motif) ligands (CXCL1, 2, 5, 6) and interleukin expression (IL8) were predominantly reduced. Epidermal differentiation markers such as loricrin, filaggrin-1 and filaggrin-2 were upregulated by both tested laser optics, indicating a potential epidermal involvement. These effects were also shown on protein level in the immunofluorescence analysis. This novel standardised laser-treated human 3D skin model proves useful for monitoring time-dependent ex vivo effects of various laser systems on gene expression and human skin morphology. Our study reveals erbium glass laser-induced regulations of MMP and interleukin expression. We speculate that these alterations on gene expression level could play a role for dermal remodelling, anti-inflammatory effects and increased epidermal differentiation. Our finding may have implications for further understanding of the molecular mechanism of erbium glass laser-induced effects on human skin.


Subject(s)
Cicatrix/radiotherapy , Lasers, Solid-State/therapeutic use , Skin/radiation effects , Cicatrix/pathology , Filaggrin Proteins , Gene Expression/radiation effects , Gene Expression Regulation/radiation effects , Humans , Low-Level Light Therapy/methods , Models, Biological , Skin/pathology , Tissue Culture Techniques
11.
Exp Dermatol ; 24(9): 663-8, 2015 Sep.
Article in English | MEDLINE | ID: mdl-25809190

ABSTRACT

Inter-α-trypsin inhibitors are protease inhibitors that are thought to be important regulators in various acute-phase processes. They are composed of one light chain (bikunin) and different heavy chains (ITIHs). The only function known so far of ITIHs is the covalent linkage to hyaluronan (HA). As there is virtually no knowledge on the distribution and function of ITIH proteins in skin tissue, we performed a systematic characterization of ITIH expression in healthy and diseased skin. Using GeneChip(®) Human Exon 1.0 ST expression profiling, we found that ITIH5 represents the major ITIH family member expressed in human skin. Moreover, the use of quantitative reverse transcription PCR and a customized ITIH5-specific antibody indicated that ITIH5 is predominantly produced by dermal fibroblasts. Immunohistochemical analysis revealed a clearly detectable ITIH5 protein expression in normal skin. Interestingly, ITIH5 expression was significantly up-regulated in inflammatory skin diseases. Furthermore, 3D skin models employing murine Itih5(-/-) epidermal keratinocytes and dermal fibroblasts as well as skin specimens of Itih5(-/-) mice revealed a significantly altered epidermal structure compared to wild-type controls. Hence, we can strengthen the presumption that ITIH5 may constitute a novel regulatory molecule of the human skin that could play an important role in inflammation via its interaction with HA.


Subject(s)
Epidermis/metabolism , Gene Expression , Proteinase Inhibitory Proteins, Secretory/genetics , Proteinase Inhibitory Proteins, Secretory/metabolism , Skin Diseases/genetics , Skin Diseases/metabolism , Alpha-Globulins/genetics , Animals , Blood Proteins/genetics , Cells, Cultured , Epidermis/chemistry , Epidermis/pathology , Female , Fibroblasts , Gene Expression Profiling , Glycoproteins/genetics , Humans , Hyaluronic Acid/metabolism , Inflammation/genetics , Keratinocytes , Mice , Mice, Knockout , Models, Anatomic , Oligonucleotide Array Sequence Analysis , Proteinase Inhibitory Proteins, Secretory/analysis , RNA, Messenger/metabolism , Up-Regulation
12.
Lasers Surg Med ; 47(3): 257-65, 2015 Mar.
Article in English | MEDLINE | ID: mdl-25771913

ABSTRACT

BACKGROUND AND OBJECTIVE: At present, there is no standardized in vitro human skin model for wound healing. Therefore, our aim was to establish and characterize an in vitro/ex vivo three-dimensional (3D) wound healing model, which we employed to analyze the effects of dexpanthenol on wound healing and gene regulation. MATERIALS AND METHODS: The novel human 3D skin wound healing model using scaffold and collagen 3D organotypic skin equivalents was irradiated with a non-sequential fractional ultrapulsed CO2 laser. These standardized injured full-thickness skin equivalents enable qRT-PCR, microarray, and histological studies analyzing the effect of topically or systemically applied compounds on skin wound healing. RESULTS: These human laser-irradiated skin models were found to be appropriate for in vitro wound healing analysis. Topical treatment of skin wounds with a 5% dexpanthenol water-in-oil emulsion or two different 5% dexpanthenol oil-in-water emulsions clearly enhanced wound closure compared to laser-irradiated untreated control models. To find out whether this positive effect is caused by the active substance dexpanthenol, laser-irradiated skin models were cultured in calciumpantothenate containing medium (20 µg/ml) compared to skin equivalents cultured without calciumpantothenate. 3D models cultured in calciumpantothenate revealed considerably faster wound closure compared to the control models. Quantitative RT-PCR studies showed enhanced mRNA expression of MMP3, IL1α, keratin-associated protein 4-12 (KRTAP4-12), and decreased expression of S100A7 in laser-irradiated skin models cultured in medium containing calciumpantothenate. CONCLUSION: This novel standardized human 3D skin wound healing model proves useful for topical pharmacological studies on wound healing and reveals new insights into molecular mechanisms of dexpanthenol-mediated effects on wound healing. In addition, these novel 3D model systems can be used to monitor ex vivo effects of various laser systems on gene expression and morphology of human skin.


Subject(s)
Lasers, Gas/therapeutic use , Models, Biological , Pantothenic Acid/analogs & derivatives , Wound Healing/drug effects , Cells, Cultured , Dermatologic Surgical Procedures/methods , Humans , Pantothenic Acid/pharmacology , Wound Healing/genetics
13.
Skin Pharmacol Physiol ; 28(4): 205-212, 2015.
Article in English | MEDLINE | ID: mdl-25721651

ABSTRACT

BACKGROUND/AIM: Vitamin A (all- trans -retinol, ATRol) serves as a precursor for all- trans -retinoic acid (ATRA), a ligand for the retinoic acid receptor (RAR), representing a potent regulator for many physiological processes. While murine melanoma cells are highly sensitive to retinoid treatment, human melanoma cells have developed still unidentified mechanisms that mediate cellular retinoid resistance. One of the key retinoid metabolizing enzymes is lecithin retinol acyltransferase (LRAT), which catalyzes the transformation of ATRol into inactive retinyl esters. LRAT is highly expressed in human melanoma cells. The aim of this study was to identify the mechanisms in retinol metabolism that are responsible for cellular retinoid sensitivity in the murine melanoma cell line B16F10. METHODS: mRNA expression analysis, cell viability assessment and determination of intracellular retinoid levels using HPLC analysis of a generated LRAT-overexpressing B16F10 cell line compared to the control B16F10 cell line. RESULTS: We found that the murine retinoid-sensitive B16F10 cell line does not express the enzyme LRAT. LRAT overexpression decreased the antiproliferative effects of retinoid treatment in these melanoma cells. The RAR-regulated enzyme Cyp26a1 showed a significantly lower expression in LRAT-overexpressing B16F10 cells. Cyp26a1 expression was restored after ATRA incubation. HPLC analysis revealed that the level of inactive retinyl ester increased after ATRol treatment, and levels of the substrate ATRol and biologically active ATRA significantly decreased in LRAT-overexpressing murine melanoma. Consistently with this, levels of 4-oxoretinoic acid, an ATRA metabolite and Cyp26a1 product, were also decreased in LRAT-overexpressing cells. CONCLUSION: Our results revealed a direct link between LRAT expression and regulation of ATRA levels indicating that the absence of LRAT-catalyzed retinol esterification is important for mediating retinoid sensitivity in murine melanoma cells. Thus, our data suggest that LRAT overexpression represents a novel mechanism by which tumor cells can escape high supplementary ATRA levels that mediate tumor-suppressive RAR signaling.


Subject(s)
Acyltransferases/metabolism , Melanoma, Experimental/metabolism , Retinaldehyde/pharmacology , Tretinoin/pharmacology , Vitamin A/pharmacology , Acyltransferases/genetics , Animals , Cell Line, Tumor , Cytochrome P-450 Enzyme System/genetics , Mice , Retinaldehyde/analogs & derivatives , Retinoic Acid 4-Hydroxylase
14.
Exp Dermatol ; 23(11): 832-7, 2014 Nov.
Article in English | MEDLINE | ID: mdl-25236354

ABSTRACT

Retinoids such as all-trans retinoic acid (ATRA) influence cell growth, differentiation and apoptosis and may play decisive roles in tumor development and progression. An essential retinoid-metabolizing enzyme known as lecithin retinol acyltransferase (LRAT) is expressed in melanoma cells but not in melanocytes catalysing the esterification of all-trans retinol (ATRol). In this study, we show that a stable LRAT knockdown (KD) in the human melanoma cell line SkMel23 leads to significantly increased levels of the substrate ATRol and biologically active ATRA. LRAT KD restored cellular sensitivity to retinoids analysed in cell culture assays and melanoma 3D skin models. Furthermore, ATRA-induced gene regulatory mechanisms drive depletion of added ATRol in LRAT KD cells. PCR analysis revealed a significant upregulation of retinoid-regulated genes such as CYP26A1 and STRA6 in LRAT KD cells, suggesting their possible involvement in mediating retinoid resistance in melanoma cells. In conclusion, LRAT seems to be important for melanoma progression. We propose that reduction in ATRol levels in melanoma cells by LRAT leads to a disturbance in cellular retinoid level. Balanced LRAT expression and activity may provide protection against melanoma development and progression. Pharmacological inhibition of LRAT activity could be a promising strategy for overcoming retinoid insensitivity in human melanoma cells.


Subject(s)
Acyltransferases/genetics , Melanoma/metabolism , Skin Neoplasms/metabolism , Tretinoin/chemistry , Catalysis , Cell Line, Tumor , Cells, Cultured , Disease Progression , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Neoplastic , Gene Knockdown Techniques , Humans , Keratinocytes/cytology , Melanocytes/metabolism , Vitamin A/chemistry , Melanoma, Cutaneous Malignant
15.
J Allergy Clin Immunol ; 129(2): 426-33, 433.e1-8, 2012 Feb.
Article in English | MEDLINE | ID: mdl-22177328

ABSTRACT

BACKGROUND: Atopic dermatitis (AD) is an inflammatory skin disease affecting 10% to 20% of children and 1% to 3% of adults in industrialized countries. Enhanced expression of IL-31 is detected in skin samples of patients with AD, but its physiological relevance is not known. OBJECTIVE: We sought to determine the role of IL-31 in skin differentiation. METHODS: We used human 3-dimensional organotypic skin models with either primary keratinocytes or HaCaT keratinocytes with inducible IL-31 receptor α to evaluate the effect of IL-31. The consequences were studied by using histology, the expression of markers analyzed by immunofluoresence and quantitative RT-PCR, and gene expression arrays. RESULTS: We observed that IL-31 interferes with keratinocyte differentiation. Gene expression analysis revealed a limited set of genes deregulated in response to IL-31, including IL20 and IL24. In HaCaT keratinocytes with inducible IL-31 receptor α, IL-31 inhibited proliferation upon induction of IL-31 receptor α by inducing cell cycle arrest. As in primary cells, IL-31-treated HaCaT cells elicited a differentiation defect in organotypic skin models, associated with reduced epidermal thickness, disturbed epidermal constitution, altered alignment of the stratum basale, and poor development of the stratum granulosum. The differentiation defect was associated with a profound repression of terminal differentiation markers, including filaggrin, an essential factor for skin barrier formation, and a reduced lipid envelope. The highly induced proinflammatory cytokines IL-20 and IL-24 were responsible for part of the effect on FLG expression and thus for terminal differentiation. CONCLUSION: Our study suggests that IL-31 is an important regulator of keratinocyte differentiation and demonstrates a link between the presence of IL-31 in skin, as found in patients with AD, and filaggrin expression.


Subject(s)
Dermatitis, Atopic/metabolism , Interleukins/metabolism , Intermediate Filament Proteins/metabolism , Keratinocytes/cytology , Cell Differentiation/drug effects , Cell Line , Cells, Cultured , Filaggrin Proteins , Humans , Interleukins/genetics , Keratinocytes/metabolism , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Skin/metabolism
16.
Exp Dermatol ; 20(5): 450-2, 2011 May.
Article in English | MEDLINE | ID: mdl-21366702

ABSTRACT

Several transport proteins are constitutively expressed in skin cells, but the putative role of the ABC transporter P-glycoprotein (P-gp) in human skin is yet unknown. Therefore, we analysed mRNA and protein expression and localization of P-gp in human skin. Using qRT-PCR, we demonstrated a strong MDR1 mRNA expression in whole skin specimens and dermis, whereas the expression of MDR1 in epidermis, epidermal keratinocytes or dermal fibroblasts was only weak. Immunohistochemistry confirmed mRNA data and revealed a marked expression of P-gp within sweat ducts, vessels, nerve sheaths and muscles of human skin and a moderate expression in basal epidermis. Our findings closely correlate with previous studies in murine skin supporting the role of P-gp in the uptake of compounds from the epidermal compartment and their secretion into the bloodstream and sweat ducts. It may also prevent the uptake of xenobiotics into the skin by functioning as a barrier located in the dermal vasculature.


Subject(s)
ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Dermis/metabolism , Gene Expression/genetics , Skin/metabolism , ATP Binding Cassette Transporter, Subfamily B , Blood Vessels/metabolism , Carcinoma, Squamous Cell/metabolism , Cells, Cultured , Epidermis/metabolism , Fibroblasts/metabolism , Humans , Keratinocytes/metabolism , Liver/metabolism , Muscle, Smooth/metabolism , Myelin Sheath/metabolism , Skin/blood supply , Skin Neoplasms/metabolism , Sweat Glands/metabolism
18.
Arch Biochem Biophys ; 508(2): 212-6, 2011 Apr 15.
Article in English | MEDLINE | ID: mdl-21284934

ABSTRACT

The multidrug resistance related proteins (MRPs) function as efflux transporters of a variety of large organic anions or their conjugates. In recent studies we demonstrated that antigen-presenting cells express a specific pattern of MRPs. MRP-mediated efflux activity of human monocyte-derived dendritic cells (moDCs) was analyzed using an in vitro transport assay. The efflux transport of radiolabeled contact allergens was inhibited using the specific MRP inhibitor indomethacin. Treatment with indomethacin increased intracellular concentration of [³H] eugenol and [³H] isoeugenol in moDCs. In addition by using MRP1 expressing inside-out membrane vesicles we revealed that the transport of eugenol is mediated by MRP1. Human DCs were employed to assess the sensitizing potential of contact allergens and alters their cytokine gene expression profile. Hence, to survey the functionality of indomethacin after stimulation with contact allergens IL-8 and TRIM16 regulation was measured by a DC-based in vitro assay. Incubation with isoeugenol after pre-treatment with indomethacin leads to increased IL-8 and TRIM16 gene expression. These results strongly support the functional role of MRPs in the active efflux of contact allergens also in antigen-presenting cells like moDCs, a novel mechanism which could possibly play a role in the pathogenesis of contact allergy.


Subject(s)
Allergens/metabolism , Dendritic Cells/metabolism , Monocytes/cytology , Multidrug Resistance-Associated Proteins/metabolism , Biological Transport, Active/drug effects , Dendritic Cells/cytology , Eugenol/metabolism , Humans , Indomethacin/pharmacology , Multidrug Resistance-Associated Proteins/antagonists & inhibitors
19.
EMBO Rep ; 10(7): 783-9, 2009 Jul.
Article in English | MEDLINE | ID: mdl-19483673

ABSTRACT

Y-box protein (YB)-1 of the cold-shock protein family functions in gene transcription and RNA processing. Extracellular functions have not been reported, but the YB-1 staining pattern in inflammatory glomerular diseases, without adherence to cell boundaries, suggests an extracellular occurrence. Here, we show the secretion of YB-1 by mesangial and monocytic cells after inflammatory challenges. It should be noted that YB-1 was secreted through a non-classical mode resembling that of the macrophage migration inhibitory factor. YB-1 release requires ATP-binding cassette transporters, and microvesicles protect YB-1 from protease degradation. Two lysine residues in the YB-1 carboxy-terminal domain are crucial for its release, probably because of post-translational modifications. The addition of purified recombinant YB-1 protein to different cell types results in increased DNA synthesis, cell proliferation and migration. Thus, the non-classically secreted YB-1 has extracellular functions and exerts mitogenic as well as promigratory effects in inflammation.


Subject(s)
Extracellular Space/metabolism , Mitogens/metabolism , Secretory Pathway , Y-Box-Binding Protein 1/metabolism , Cell Movement/drug effects , Cell Proliferation/drug effects , Extracellular Space/drug effects , Humans , Lipopolysaccharides/pharmacology , Monocytes/cytology , Monocytes/drug effects , Monocytes/metabolism , Secretory Pathway/drug effects , Secretory Vesicles/drug effects , Secretory Vesicles/metabolism
20.
Life (Basel) ; 11(8)2021 Aug 19.
Article in English | MEDLINE | ID: mdl-34440590

ABSTRACT

Psoriasis is a chronic skin disease affecting 2-3% of the global population. The proinflammatory IL-17A is a key cytokine in psoriasis. Accumulating evidence has revealed that IL-36γ plays also a pathogenic role. To understand more precisely the role of the IL-17A-IL-36γ cytokine network in skin pathology, we used an ear injection model. We injected IL-17A or IL-36γ alone and in combination into the ear pinnae of mice. This resulted in a significant increase in ear thickness measured over time. Histological evaluation of IL-17A + IL-36γ-treated skin showed a strong acanthosis, hyperparakeratosis and infiltration of neutrophils. The same histological features were found in mice after injection of IL-36γ alone, but to a lesser extent. IL-17A alone was not able to induce psoriasis-like changes. Genes encoding proteins of the S100 family, antimicrobial peptides and chemo-attractants for neutrophils were upregulated in the IL-17A + IL-36γ group. A much weaker expression was seen after the injection of each cytokine alone. These results strengthen the hypothesis that IL-17A and IL-36γ drive psoriatic inflammation via a synergistic interaction. Our established intradermal ear injection model can be utilized in the future to monitor effects of various inhibitors of this cytokine network.

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