Vehicle-treated Skin Research Articles

LB986 A novel three-dimensional imaging and analysis of imiquimod-induced mouse skin

Here we report a novel three-dimensional (3D) assessment technique for TLR7/8 agonist-induced mouse skin samples using the TissueCyte Serial Two-Photon Plus (STP2) imaging platform. Wild-type (WT) mouse skin was treated with either TLR7/8 agonist imiquimod (IMQ) cream, a commonly used method for modeling psoriasis in mice, or vehicle. Explants were sectioned serially and imaged to visualize tissue autofluorescence and collagen fibril signatures using second harmonic generation (SHG) imaging. The hypodermis, dermis, and epidermis skin layers across each section were segmented and the layer volume (millimeter3) was computed. Results indicate an increase in volume of all assessed tissue layers of the IMQ-treated versus vehicle-treated skin, by 1.6-fold in the hypodermis, 1.4-fold in the dermis, and 3.0-fold in the epidermis. This study demonstrates that IMQ cream produces inflammation throughout the hypodermis, dermis, and epidermis of mouse skin. The ability to process TLR7/8 agonized mouse skin samples using the STP2 imaging platform demonstrates a new method to produce a 3D model and analyze total gross morphology with collagen fibril signatures. The techniques developed in this study hold promise for assessing inflammation type and disease severity of psoriasis, with future application to thoroughly evaluate pharmacodynamics and pharmacokinetics of drug treatment.

Tirbanibulin: A New Topical Therapy for Actinic Keratoses With a Novel Mechanism of Action and Improved Ease of Use.

Tirbanibulin: A New Topical Therapy for Actinic Keratoses With a Novel Mechanism of Action and Improved Ease of Use.

Anti-allergic effect of the Src family kinase inhibitor saracatinib.

The aim of this study was to evaluate the anti-anaphylactic and anti-allergic potentials of saracatinib, a Src family kinase inhibitor that was already shown to be safe in clinical trials when it was used as an anti-cancer drug. Using in vitro mast cell models, we found that saracatinib inhibited the degranulation response and cytokine production in RBL2H3 cells that were stimulated with IgE and antigen without affecting cell viability. Phosphorylation of Lyn, Akt, a PI3K substrate, and MAPKs including ERK, JNK, and p38, as well as the intracellular Ca2+ increase induced by this stimulation were also suppressed by saracatinib. This drug also inhibited symptoms in our established anaphylaxis mouse model, anaphylaxis-dependent spotted distribution of immune complex in skin (ASDIS). The intravenous injection of the mixture of IgE and antigen induced acute spotted distribution of immune complex in skin in hairless HR-1 mice, and its inhibition by intradermal injection of saracatinib was observed. Moreover, toluidine blue-stained skin sections indicated that the degranulation ratio of dermal mast cells was reduced in saracatinib-treated skin compared with vehicle-treated skin. Because only a few signaling inhibitors are used as anti-anaphylaxis and anti-allergic drugs, these results indicated the valuable suggestion that saracatinib and the Src family kinase inhibitors are good candidates for anti-anaphylaxis and anti-allergic drugs.

Long-term Topical Oestrogen Treatment of Sun-exposed Facial Skin in Post-menopausal Women Does Not Improve Facial Wrinkles or Skin Elasticity, But Induces Matrix Metalloproteinase-1 Expression

It is controversial whether treatment with oestrogen stimulates collagen production or accumulation in sun-exposed skin. The aim of this study was to determine the effect of long-term treatment with topical oestrogen on photoaged facial skin, with regard to wrinkle severity, and expression of procollagen and matrix metalloproteinase-1 enzyme. Two groups of 40 post-menopausal women applied either 1 g of 1% oestrone or vehicle cream once daily to the face for 24 weeks. Visiometer R1-R5 values (skin wrinkles) and Cutometer values (skin elasticity) were not significantly improved in the oestrone group after 24 weeks of treatment. Type I procollagen immunostaining did not increase in the oestrone group compared with the control group. However, levels of matrix metalloproteinase-1 mRNA increased robustly (10.3 times) in oestrone-treated skin compared with vehicle-treated skin. Thus, treatment with topical oestrogen may be deleterious in ultraviolet-induced skin ageing, at least in part, through induction of matrix metalloproteinase-1 (MMP-1) expression in human skin.

In Vivo Imaging of Human and Mouse Skin with a Handheld Dual-Axis Confocal Fluorescence Microscope

Advancing molecular therapies for the treatment of skin diseases will require the development of new tools that can reveal spatiotemporal changes in the microanatomy of the skin and associate these changes with the presence of the therapeutic agent. For this purpose, we evaluated a handheld dual-axis confocal (DAC) microscope that is capable of in vivo fluorescence imaging of skin, using both mouse models and human skin. Individual keratinocytes in the epidermis were observed in three-dimensional image stacks after topical administration of near-infrared (NIR) dyes as contrast agents. This suggested that the DAC microscope may have utility in assessing the clinical effects of a small interfering RNA (siRNA)-based therapeutic (TD101) that targets the causative mutation in pachyonychia congenita (PC) patients. The data indicated that (1) formulated indocyanine green (ICG) readily penetrated hyperkeratotic PC skin and normal callused regions compared with nonaffected areas, and (2) TD101-treated PC skin revealed changes in tissue morphology, consistent with reversion to nonaffected skin compared with vehicle-treated skin. In addition, siRNA was conjugated to NIR dye and shown to penetrate through the stratum corneum barrier when topically applied to mouse skin. These results suggest that in vivo confocal microscopy may provide an informative clinical end point to evaluate the efficacy of experimental molecular therapeutics.

Retinoids suppress cysteine‐rich protein 61 (CCN1), a negative regulator of collagen homeostasis, in skin equivalent cultures and aged human skin in vivo

Alterations in connective tissue collagen are prominent features of both chronologically aged and photoaged (ageing because of sun exposure) human skin. These age-related abnormalities are mediated in part by cysteine-rich protein 61 (CCN1). CCN1 is elevated in the dermis of both chronologically aged and photoaged human skin in vivo and promotes aberrant collagen homeostasis by down-regulating type I collagen, the major structural protein in skin, and promoting collagen degradation. Vitamin A and its metabolites have been shown to improve chronologically aged and photoaged skin by promoting deposition of new collagen and preventing its degradation. Here, we investigated regulation of CCN1 expression by retinoids in skin equivalent cultures and chronologically aged and photoaged human skin in vivo. In skin equivalent cultures, all-trans retinoic acid (RA), the major bioactive form of vitamin A in skin, significantly increased type I procollagen and reduced collagenase (matrix metalloproteinases-1, MMP-1). Addition of recombinant human CCN1 to skin equivalent cultures significantly reduced type I procollagen and increased MMP-1. Importantly, RA significantly reduced CCN1 expression in skin equivalent cultures. Topical treatment with retinol (vitamin A, 0.4%) for 7days significantly reduced CCN1 mRNA and protein expression in both chronologically aged (80+years) and photoaged human skin in vivo, compared to vehicle-treated skin. These data indicate that the mechanism by which retinoids improve aged skin, through increased collagen production, involves down-regulation of CCN1.

Clinical and histological evaluation of 1% nadifloxacin cream in the treatment of acne vulgaris in Korean patients

Nadifloxacin is a fluoroquinolone with broad-spectrum antibacterial activity. Although it is used as an acne treatment in some European countries, it has not been used to treat Korean acne patients. We aimed to evaluate the clinical efficacy and safety of 1% nadifloxacin cream and the histological changes it incurs when used to treat mild to moderate facial acne in Korean patients. An eight-week, randomized, prospective, split-face, double-blind, vehicle-controlled trial was performed. All participants were treated with 1% nadifloxacin cream on one-half of the face and vehicle cream on the other, twice per day for eight weeks. At final visits, inflammatory acne lesions were reduced by 70% on nadifloxacin-treated skin and increased by 13.5% on vehicle-treated skin; non-inflammatory acne lesions showed reductions of 48.1 and 10.1%, respectively. A significant difference was observed between the two treatments at four weeks. Histopathological examinations of the acne lesions showed decreased inflammation and interleukin-8 expression but no change in transforming growth factor-β expression in nadifloxacin-treated skin compared with vehicle-treated skin after eight weeks of treatment. Nadifloxacin 1% cream is an effective, safe, and well-tolerated topical treatment for Korean patients with mild to moderate acne vulgaris. Histopathological changes after nadifloxacin treatment were well correlated with clinical outcomes. Therefore, nadifloxacin can be used as an effective and safe treatment option in the management of mild to moderate acne in Asian subjects.

Effects of Serine Palmitoyltransferase Inhibitor ISP-I on the Stratum Corneum of Intact Mouse Skin

Serine palmitoyltransferase (SPT) is involved in the ceramide synthesis pathway. We investigated the effects of ISP-I, a potent inhibitor of SPT, on the stratum corneum (SC) of hairless mouse skin. Application of ISP-I for one week resulted in a significant decrease in the amount of ceramide, which was associated with a decrease in SC hydration. However, there was an increase in the number of SC layers and less transepidermal water loss than control. Transmission Electron Microscopy observation revealed that the number of desmosome-like structures in the layers immediately above the stratum granulosum (SG) was significantly increased in ISP-I-treated skin compared to vehicle-treated skin. The activity of serine protease-an enzyme associated with the process of desquamation-was lower in the SC of ISP-I-treated skin than control. Furthermore, immunoelectronmicroscopy revealed that glucosylceramide and corneodesmosin tended to remain in corneocytes and were not secreted into the intercellular spaces of the SC in the ISP-I-treated skin. These results indicate that the application of ISP-I decreases ceramide and skin hydration, while at the same time increases the number of SC layers. The accumulation of corneocyte layers may originate from an aberrant desquamation process related to the decrease in the serine protease activity as well as an alteration in the transport of desquamation-related proteases by lamellar bodies.

The Effects of Serine Palmitoyltransferase Inhibitor, ISP-I, on UV-Induced Barrier Disruption in the Stratum Corneum

We examined the effects of ISP-I (myriocin, thermozymocidin) - a potent inhibitor of serine palmitoyltransferase (SPT) which is involved in the ceramide synthetic pathway-on skin barrier function in post-UVB-irradiated hairless mouse skin. Disruption of the skin barrier function after UVB irradiation as represented by the increase in transepidermal water loss (TEWL) was significantly suppressed with ISP-I treatment. In the ISP-I-treated skin, the peak of cell proliferation was observed 24 h earlier than in vehicle-treated skin. In addition, the number of apoptotic cells in ISP-I-treated skin showed a sharp decrease at 48 and 72 h post-irradiation. The number of stratum corneum cell layers was increased in ISP-I-treated skin at 72 h after UVB irradiation; at this time, TEWL in ISP-I-treated skin was lower than that in the vehicle-treated skin. We suggest ISP-I treatment altered cell proliferation and apoptosis after UVB exposure by modulating ceramide synthesis in epidermal cells, resulting in an increase of stratum corneum layers which lessened the effects of irradiation-induced barrier disruption.

Erbb2 Suppresses DNA Damage-Induced Checkpoint Activation and UV-Induced Mouse Skin Tumorigenesis

The Erbb2 receptor is activated by UV irradiation, the primary cause of non-melanoma skin cancer. We hypothesized that Erbb2 activation contributes to UV-induced skin tumorigenesis by suppressing cell cycle arrest. Consistent with this hypothesis, inhibition of Erbb2 in v-ras(Ha) transgenic mice before UV exposure resulted in both 56% fewer skin tumors and tumors that were 70% smaller. Inhibition of the UV-induced activation of Erbb2 also resulted in milder epidermal hyperplasia, S-phase accumulation, and decreased levels of the cell cycle regulator Cdc25a, suggesting altered cell cycle regulation on inhibition of Erbb2. Further investigation using inhibition or genetic deletion of Erbb2 in vitro revealed reduced Cdc25a levels and increased S-phase arrest in UV-irradiated cells lacking Erbb2 activity. Ectopic expression of Cdc25a prevented UV-induced S-phase arrest in keratinocytes lacking Erbb2 activity, demonstrating that maintenance of Cdc25a by Erbb2 suppresses cell cycle arrest. Examination of checkpoint pathway activation upstream of Cdc25a revealed Erbb2 activation did not alter Ataxia Telangiectasia and Rad3-related/Ataxia Telangiectasia Mutated activity but increased inhibitory phosphorylation of Chk1-Ser(280). Since Akt phosphorylates Chk1-Ser(280), the effect of Erbb2 on phosphatidyl inositol-3-kinase (PI3K)/Akt signaling during UV-induced cell cycle arrest was determined. Erbb2 ablation reduced the UV-induced activation of PI3K while inhibition of PI3K/Akt increased UV-induced S-phase arrest. Thus, UV-induced Erbb2 activation increases skin tumorigenesis through inhibitory phosphorylation of Chk1, Cdc25a maintenance, and suppression of S-phase arrest via a PI3K/Akt-dependent mechanism.

Topical treatment with CYP26 inhibitor talarozole (R115866) dose dependently alters the expression of retinoid-regulated genes in normal human epidermis

An alternative approach to retinoid therapy is to inhibit the cytochrome P450 (CYP)-mediated catabolism of endogenous all-trans retinoic acid in the skin by applying retinoic acid metabolism blocking agents such as talarozole (R115866). To study the effects of topical talarozole on retinoid biomarkers in normal skin in a randomized phase I trial. Gels containing talarozole (0.35% or 0.07%) and vehicle were applied once daily for 9 days on either buttock of 16 healthy volunteers. Epidermal shave biopsies (for mRNA analysis) and punch biopsies (for histology and immunofluorescence analysis) were collected from the treatment areas. Genes encoding the following were studied by quantitative real-time polymerase chain reaction: cellular retinoic acid binding protein 2 (CRABP2), cytokeratins (KRT2 and KRT4), CYP26A1, CYP26B1, CYP26C1 and CYP2S1, two enzymes in the retinol metabolism (retinal dehydrogenase-2 and retinol acyltransferase) and two proinflammatory cytokines [interleukin (IL)-1alpha and tumour necrosis factor-alpha]. Talarozole treatment increased the mRNA expression of CRABP2, KRT4, CYP26A1 and CYP26B1 dose dependently, and decreased the expression of KRT2 and IL-1alpha compared with vehicle-treated skin. No mRNA change in retinol-metabolizing enzymes was obtained. There was no induction of epidermal thickness or overt skin inflammation in talarozole-treated skin. Immunofluorescence analysis confirmed an upregulation of KRT4 protein, but no upregulation of CYP26A1 and CYP26B1 expression was detected. Talarozole influences the biomarker pattern consistently with increased retinoic acid stimulation. The low irritancy of talarozole at the two examined dosages is a possible advantage over topical retinoids.

Evaluation of efficacy and safety of rucinol serum in patients with melasma: a randomized controlled trial

Melasma is a hyperpigmentation disorder predominantly affecting sun-exposed areas in women, which is often refractory to treatment. Most commercially available treatments incorporate inhibitors of tyrosinase, a key enzyme in melanin production within the melanocyte. In general, however, the efficacy of these therapies is somewhat limited. Recent studies have identified other enzymes that play an important role in melanogenesis, including tyrosinase-related protein-1 (TRP-1), which catalyses the oxidation of the melanogenetic intermediate 5,6-dihydroxyindole-2-carbolylic acid. Rucinol (4-n-butylresorcinol) has been shown to inhibit the activity of both tyrosinase and TRP-1. To assess the efficacy of rucinol serum 0.3% vs. the corresponding vehicle as a treatment for melasma. Secondary objectives were to evaluate local and general tolerability and to assess the skin acceptability of rucinol serum in the target population. In this prospective, single-centre, double-blind, randomized, vehicle-controlled, bilateral (split-face) comparative trial, 32 women with melasma were provided with two identical tubes containing rucinol serum 0.3% or vehicle. The products were each applied to one-half of the face, according to the randomization scheme, twice daily for 12 weeks (phase 1). A broad-spectrum sunscreen (sun protection factor 60) was also applied daily. Assessments at baseline, 4, 8 and 12 weeks included clinical evaluations by a dermatologist, chromametry, ultraviolet and standard photography, and assessments of skin acceptability and tolerability. After 12 weeks, patients were given the option of an additional 3-month treatment period of open full-face rucinol treatment, with reviews at 16, 20 and 24 weeks (phase 2). Twenty-eight patients completed phase 1 and 26 patients completed phase 2. After 12 weeks, the clinical pigmentation score for rucinol-treated skin was significantly lower than for vehicle-treated skin (P = 0.027). During phase 2, rucinol induced a significant reduction in mean pigmentation score on the half of the face previously treated with vehicle. There was also a further, significant improvement on the rucinol-treated side of the face. Chromametry measurements showed that skin was significantly lighter and less yellow, with a strong trend towards reduced redness, following rucinol therapy compared with vehicle. Rucinol serum showed good tolerability and acceptability and was considered to have good or fair efficacy by 78% of the patient population. Rucinol serum was shown to have significant efficacy compared with vehicle alone in improving melasma after 3 months of treatment, according to clinical and objective assessments of skin colour.

Ferulic Acid Stabilizes a Solution of Vitamins C and E and Doubles its Photoprotection of Skin

Ferulic acid is a potent ubiquitous plant antioxidant. Its incorporation into a topical solution of 15%l-ascorbic acid and 1%alpha-tocopherol improved chemical stability of the vitamins (C+E) and doubled photoprotection to solar-simulated irradiation of skin from 4-fold to approximately 8-fold as measured by both erythema and sunburn cell formation. Inhibition of apoptosis was associated with reduced induction of caspase-3 and caspase-7. This antioxidant formulation efficiently reduced thymine dimer formation. This combination of pure natural low molecular weight antioxidants provides meaningful synergistic protection against oxidative stress in skin and should be useful for protection against photoaging and skin cancer.

Topical Pretreatment of Diabetic Rats With All-trans Retinoic Acid Improves Healing of Subsequently Induced Abrasion Wounds

In the current study, rats were made diabetic with streptozotocin (STZ) and maintained for 8 weeks, during which time they were treated topically on alternative days with a solution of 0.1% all-trans retinoic acid in a vehicle of 70:30% ethanol/propylene glycol. STZ-induced diabetic rats treated with vehicle served as controls. Additional nondiabetic rats were treated with all-trans retinoic acid or vehicle in parallel. At the end of the 8-week period, rats from all four treatment groups were subjected to abrasion wound formation. Wounds healed more rapidly in vehicle-treated nondiabetic skin than in vehicle-treated diabetic skin (96% of the wound surface area closed in nondiabetic rats within 6 days vs. 41% closed in diabetic rats). Wounds in all-trans retinoic acid-treated diabetic skin healed more rapidly than wounds in vehicle-treated diabetic skin (85% of the wound surface area closed in all-trans retinoic acid-treated diabetic rats vs. 41% closed in vehicle-treated diabetic rats). At the histological level, recently healed skin from vehicle-treated diabetic rats was shown to contain a thin, wispy provisional matrix in which many of the embedded cells were rounded and some were pycnotic. In contrast, a much denser provisional matrix with large numbers of embedded spindle-shaped cells was observed in healed wounds from diabetic skin that had been pretreated with all-trans retinoic acid. The all-trans retinoic acid-treated diabetic skin was histologically similar to vehicle-treated (or all-trans retinoic acid-treated) skin from nondiabetic animals. In light of these findings, we suggest that prophylactic use of retinoid-containing preparations might be useful in preventing the development of nonhealing skin ulcers resultant from minor traumas in at-risk skin.

Topical retinoic acid alters the expression of cellular retinoic acid-binding protein-I and cellular retinoic acid-binding protein-II in non-lesional but not lesional psoriatic skin.

Therapeutic retinoids have profound effects on psoriatic skin pathology but their interactions with various retinoid-binding proteins in lesional vs non-lesional skin have not been investigated. Using quantitative real-time PCR the mRNA expression of cellular retinol-binding protein I (CRBPI) and retinoic acid-binding protein I/II (CRABPI/CRABPII) was studied in psoriatic and healthy control (=normal) skin after 4 days of occlusive RA/vehicle treatment (n=6). Untreated psoriatic lesions showed a markedly elevated CRABPII/CRABPI ratio, while the CRBPI level was reduced in lesional and non-lesional skin as compared to normal skin. In RA-treated normal and non-lesional skin, the mRNA expression of CRBPI was unaltered while that of CRABPI and CRABPII was reduced by approximately 80% and increased approximately 5-fold, respectively, as compared to vehicle-treated skin. In contrast, lesional skin exposed to RA showed an almost 90% increase in CRBPI transcripts but unaltered expression of CRABPI and CRABPII, yet, the mRNA expression of several inflammatory mediators, e.g. inducible nitric oxide synthase, interferon-gamma and interleukin-1beta, was clearly reduced. Immunohistochemistry localized CRABPII to suprabasal keratinocytes in normal skin and revealed markedly elevated levels in lesional skin. RA treatment induced CRABPII protein expression in normal and non-lesional skin, to similar levels as in untreated lesions. The results indicate that the effects of RA differ in normal/non-lesional psoriatic skin and lesional skin. Whether the high expression of CRABPII in psoriatic skin lesions is due to increased amounts of endogenous retinoids in lesional skin or reflects an abnormal regulation of the CRABPII gene in psoriasis remains to be studied.

The role of elastases secreted by fibroblasts in wrinkle formation: implication through selective inhibition of elastase activity.

We have previously demonstrated that decreases in skin elasticity, accompanied by increases in the tortuosity of elastic fibers, are important early events in wrinkle formation. In order to study the role of elastases in the degeneration of elastic fibers during wrinkle formation we examined the effects of an inhibitor of skin fibroblast elastase, N-phenethylphosphonyl-L-leucyl-L-tryptophane (NPLT), on wrinkle formation in hairless mice skin following UV irradiation. Dorsal skins of hairless mice were exposed daily to UV light for 18 weeks at doses of 65-95 mJ/cm2 and treated topically with 100 microL of 1 mM NPLT immediately after each UV irradiation. Wrinkles on dorsal skins were evaluated from week 6 through week 18. The daily exposure of mouse skin to UV light with less than 1 minimal erythemal dose significantly enhanced the activity of elastase in the exposed skin by week 4, and the elevated levels of elastase activity were significantly reduced by the in vitro incubation with NPLT in a dose-dependent manner to a level similar to that in unexposed mice skin, indicating that NPLT can efficiently inhibit the UV-inducible elastase activity. Topical application of NPLT significantly suppressed wrinkle formation when compared with vehicle controls by week 15 of treatment (P < 0.05). Histochemistry of elastic fibers with Orcein staining demonstrated that there were no obvious decreases of the fine elastic fibers in UV-exposed NPLT-treated skin in contrast to their marked decreases in the UV-exposed vehicle-treated skin. These findings suggest that skin fibroblast elastase plays a decisive role in wrinkle formation through the degeneration of elastic fiber.

CD 2394, a novel synthetic retinoid, initiates an embryonic type of differentiation in hyperproliferative skin

In human skin, there are 2 types of epidermal differentiation: normal differentiation, characterized by keratin 10 expression, and alternative differentiation. Alternative differentiation may be regeneration-associated differentiation (keratin 6 and 16) or re-induction of embryonic differentiation (expression of keratin 13, 15 and 19). The purpose of this study was to investigate the effect of the novel synthetic retinoid CD 2394 on hyperproliferative human skin, with respect to embryonic differentiation in particular. The effects of CD 2394 were compared with untreated and vehicle-treated skin 48 h after tape-stripping. In a multiparameter flow cytometric assay, parameters of proliferation, normal differentiation, embryonic differentiation and inflammation were assessed. With respect to proliferation, treatment with CD 2394 resulted in a decreased number of cells in the G2M-phase. Normal differentiation was decreased in CD 2394 treated skin. Furthermore, most of the CD 2394 treated samples showed expression of keratin 13, which was not seen in the otherwise treated skin. A correlation between keratin 10 and keratin 13 expression could not be demonstrated. This study showed that CD 2394 is capable of inducing an embryonic pathway of differentiation, which is distinct from normal differentiation or regeneration-associated differentiation.

Omega-Hydroxyceramides are Required for Corneocyte Lipid Envelope (CLE) Formation and Normal Epidermal Permeability Barrier Function

Omega-hydroxyceramides (omega-OHCer) are the predominant lipid species of the corneocyte lipid envelope in the epidermis. Moreover, their omega-esterified-derivatives (acylCer) are major components of the stratum corneum extracellular lamellae, which regulate cutaneous permeability barrier function. Because epidermal omega-OHCer appear to be generated by a cytochrome P450-dependent process, we determined the effects of a mechanism-based inhibitor of omega-hydroxylation, aminobenzotriazole (ABT), on epidermal omega-OH Cer formation and barrier function. We first ascertained that ABT, but not hydroxybenzotriazole (OHBT), a chemical relative with no P450 inhibitory activity, inhibited the incorporation of [14C]-acetate into the omega-OH-containing Cer species in cultured human keratinocytes (68.1% +/- 6.9% inhibition versus vehicle-treated controls; p < 0.001), without altering the synthesis of other Cer and fatty acid species. In addition, ABT significantly inhibited the omega-hydroxylation of very long-chain fatty acids in cultured human keratinocytes. Topical application of ABT, but not OHBT, when applied to the skin of hairless mice following acute barrier disruption by tape-stripping, resulted in a significant delay in barrier recovery (e.g., 38.3% delay at 6 h versus vehicle-treated animals), assessed as increased transepidermal water loss. The ABT-induced barrier abnormality was associated with: (i) a significant decrease in the quantities of omega-OHCer in both the unbound and the covalently bound Cer pools; (ii) marked alterations of lamellar body structure and contents; and (iii) abnormal stratum corneum extracellular lamellar membrane structures, with no signs of cellular toxicity. Furthermore, pyridine-extraction of ABT- versus vehicle-treated skin, which removes all of the extracellular lamellae, leaving the covalently attached lipids, showed numerous foci with absent corneocyte lipid envelope in ABT- versus vehicle-treated stratum corneum. These results provide the first direct evidence for the importance of omega-OHCer for epidermal permeability function, and suggest further that acylCer and/or corneocyte lipid envelope are required elements in permeability barrier homeostasis.

Atopy patch testing--a diagnostic tool?

Atopy patch testing--a diagnostic tool?

Down-regulation of protein kinase C isoforms in irritant contact dermatitis.

Protein kinase C (PKC) isoforms are important in cell signal transduction associated with regulation of cell proliferation and differentiation. In this study, alterations of PKC isoform levels in irritant patch test reactions were detected by Western immunoblots. 4 chemically and structurally different irritants, 4% and 8% hydrochloric acid (HCl); 1% and 2% sodium hydroxide (NaOH); 20% and 40% nonanoic acid (NON) in 1-propanol; and 5% and 10% sodium dodecyl sulfate (SDS) were applied on the backs of mice in Finn Chambers and fixed with surgical dressings. The patches were kept on the skin for 24 h and removed. 24 h after removal, mild erythema with thickening of skin without vesicles was observed in all irritated skin, except that 4% HCl and 1% NaOH treated skin showed unremarkable skin reaction. No visible skin reaction was detected in vehicle-treated skin. PKC isoform alpha, beta, gamma, and delta levels in irritated skin revealed a 10% to 65% decrease compared to vehicle treated skin. These results indicate that in HCl, NaOH, NON and SDS-induced irritation, activation of the PKC related cell signal transduction cascade may be involved, and that PKC mediated events may be a common phenomenon in irritant contact dermatitis.