Cutaneous Permeability Barrier Research Articles

The role of ceramides in the disruption of the cutaneous permeability barrier, a common manifestation of skin disorders

The role of ceramides in the disruption of the cutaneous permeability barrier, a common manifestation of skin disorders

Stress and Common Dermatological Disorders: The Psychophysiological Dermatoses

Abstract Primary dermatological disorders which are frequently exacerbated by stress and whose course is affected by the psychological state of the patient are called psychophysiologic or psychosomatic diseases. Psychosocial stress plays a major role in the onset and/or aggravation of these skin diseases. A close relationship between the skin and the mind mediated via hormones and neurotransmitters and altered cutaneous permeability barrier homeostasis by psychological stress constitute the basic psychopathology. The common skin conditions precipitated/aggravated by stress include psoriasis, vitiligo, atopic dermatitis, acne vulgaris, alopecia areata, urticaria, lichen planus, prurigo, seborrheic dermatitis, and hyperhidrosis. These cutaneous diseases often have a chronic, unpredictable course with multiple remissions and relapses leading to significant psychiatric morbidity, and disease-related stress further acts as an aggravating factor for the cutaneous disease leading to a vicious cycle. Increased awareness about these psychophysiologic diseases among dermatologists has helped to incorporate psychotherapeutic treatment in the form of psychotropic drugs and nonpharmacological interventions in the management of these patients. A multidisciplinary approach consisting of dermatologists along with psychiatrists and psychologists in a liaison framework has proven useful for these patients.

Stress test of the skin: The cutaneous permeability barrier treadmill.

Studying skin barrier function is central to our understanding of many skin disorders. The past decade has seen a surge of skin barrier related investigative work. Genetic, biochemical and cell biology experiments have added much evidence to the importance of the barrier in disease pathogenesis of a variety of disorders including ichthyosis, atopic dermatitis and psoriasis. However, functional assays prove ever more important to demonstrate relevance of any of these findings. A paper published by Monash and Blank 60years ago describes a stress test of the skin barrier, measuring skin barrier recovery, a functional test of tremendous implications. This seminal paper has not been cited for almost 15years, time to acknowledge its critical importance and to review the relevance of this method today.

Cutaneous permeability barrier function in signal transducer and activator of transcription 6-deficient mice is superior to that in wild-type mice

BackgroundTh2 cytokines exhibit a variety of inhibitory effects on permeability barrier function via signal transducer and activator of transcription 6 (STAT6). However, the role of STAT6 signaling on the construction and/or homeostasis of permeability barrier function in the physiological state has not been fully assessed. ObjectiveWe compared permeability barrier function between Stat6-deficient and wild-type C57BL/6 mice at steady state. Methods and resultsMeasurement of transepidermal water loss and quantitative penetration assay revealed that permeability barrier function was superior in Stat6-deficient mice. Accordingly, expressions of loricrin, acidic sphingomyelinase (aSMase) and β-glucocerebrosidase (β-GlcCer’ase) in epidermis and ceramide levels in stratum corneum were elevated in STAT6-deficient mice. On the other hands, up-regulations of loricrin, aSMase and β-GlcCer’ase were not observed in 3-dimensionally cultured human keratinocytes transfected with siRNA for STAT6. Meanwhile, number of mast cells in the dermis was decreased in Stat6-deficient mice. ConclusionsThese results suggest that STAT6 signaling negatively affects permeability barrier function in vivo, even in the physiological state. However, the superior permeability barrier function in Stat6-deficient mice may be a secondary effect exerted via cells other than keratinocytes, such as mast cells, since mast cells are known to influence permeability barrier function in vivo. Blockade of STAT6 signaling might be a strategy to augment the permeability barrier function.

Role of lamellar bodies in the formation of the cutaneous permeability barrier

Role of lamellar bodies in the formation of the cutaneous permeability barrier

TLR3: A Receptor that Recognizes Cell Injury Is Essential for Permeability Barrier Homeostasis Following UV Irradiation

The cutaneous permeability barrier is essential for life and perturbations in this barrier are repaired rapidly. After minimal injury to the stratum corneum alterations in the calcium concentration in the outer epidermis are the primary signal inducing this repair response. In this issue, studies demonstrate that Toll-like receptor 3 has an important role in signaling permeability barrier repair following injury induced by UVB irradiation.

TNF-α and Th2 Cytokines Induce Atopic Dermatitis–Like Features on Epidermal Differentiation Proteins and Stratum Corneum Lipids in Human Skin Equivalents

Atopic dermatitis (AD) is a chronic inflammatory skin disease in which the skin barrier function is disrupted. In this inflammatory AD environment, cytokines are upregulated, but the cytokine effect on the AD skin barrier is not fully understood. We aimed to investigate the influence of Th2 (IL-4, IL-13, IL-31) and pro-inflammatory (tumor necrosis factor alpha (TNF-α)) cytokines on epidermal morphogenesis, proliferation, differentiation, and stratum corneum lipid properties. For this purpose, we used the Leiden epidermal model (LEM) in which the medium was supplemented with these cytokines. Our results show that IL-4, IL-13, IL-31, and TNF-α induce spongiosis, augment TSLP secretion by keratinocytes, and alter early and terminal differentiation-protein expression in LEMs. TNF-α alone or in combination with Th2 cytokines decreases the level of long chain free fatty acids (FFAs) and ester linked ω-hydroxy (EO) ceramides, consequently affecting the lipid organization. IL-31 increases long chain FFAs in LEMs but decreases relative abundance of EO ceramides. These findings clearly show that supplementation with TNF-α and Th2 cytokines influence epidermal morphogenesis and barrier function. As a result, these LEMs show similar characteristics as found in AD skin and can be used as an excellent tool for screening formulations and drugs for the treatment of AD.

Role of Ceramide in the Barrier Function of the Stratum Corneum, Implications for the Pathogenesis of Atopic Dermatitis

Atopic dermatitis is a recurrent dermatitis which is characterized clinically by atopic dry skin and functionally by cutaneous barrier disruption even in the non-lesional skin. These abnormalities have been thought to be mainly attributable to significantly decreased levels of ceramides even in the non-lesional stratum corneum. Recently, in association with the barrier disruption in atopic dermatitis skin, prevalent and rare loss-of-function mutations in the gene encoding filaggrin have been reported to be an important pre-disposing factor for the development of atopic dermatitis. However, a mechanistic connection between filaggrin loss-of-function mutations and barrier disruption has not been resolved and remains controversial. This review article explores the physiological and biochemical basis for the atopic dermatitis phenotype in terms of the barrier abnormality and associated factors such as ceramides and its metabolites in the following sequence: (1) Clinical characteristics as a recurrent dermatitis (2) Abnormality in cutaneous permeability barrier function (3) Is barrier disruption a cause or a result of dermatitis? (4) Is the barrier abnormality inherent or not? (5) Role of ceramides (6) Transglutaminase or filaggrin mutations and barrier disruption (7) Contribution of the barrier abnormality to the Th1/Th2 balance (8) Th1/Th2 balance and ceramide deficiency (9) S. aureus colonization and ceramide deficiency (10) Is the ceramide deficiency inherent or not? (11) Inflammation as a causative factor that downregulates ceramide generation, (12) Biological mechanisms underlying the ceramide deficiency (13) Clinical efficacy of ceramide as a major role for the pathogenesis of atopic dermatitis

Treating atopic dermatitis at the source: corrective barrier repair therapy based upon new pathogenic insights

Until recently, atopic dermatitis (AD) has been linked to Th1/Th2 cell dysregulation. But now, the opinion that inflammation in AD results from a convergence of inherited and acquired insults to the cutaneous permeability barrier, with variable contributions from inherited abnormalities in innate/adaptive immunity, is becoming increasingly accepted. Current therapy is however, still largely directed towards ameliorating immunologically triggered inflammation, rather than correcting the barrier abnormality. In this article, the authors provide an overview of epidermal barrier function; a review of recent molecular genetic studies pointing to a primary barrier abnormality in AD; a detailed description of new pathogenic insights into AD; and they compare the efficacy of several putative ‘barrier repair’ products currently utilized as adjunctive or primary therapy for AD. The authors also explore the potential of ‘next-generation’ barrier repair approaches that attack specific pathogenic mechanisms in AD (hig...

Nicotinic acetylcholine receptor stimulation impairs epidermal permeability barrier function and recovery and modulates cornified envelope proteins

AimTo characterize how nicotinic acetylcholine receptors (nAChRs) influence epidermal barrier function and recovery following prolonged stress or direct nAChR activation or antagonism. Main methodsMice were subjected to psychological stress or treated topically with nAChR agonist or antagonist for 3days. We assessed barrier permeability and recovery by measuring transepidermal water loss (TEWL) before and after barrier disruption. In parallel, we analyzed the production and localization of several epidermal cornified envelope proteins in mouse skin and in human EpiDerm™ organotypic constructs stimulated with a nAChR agonist (nicotine) and/or a nAChR selective antagonist (α-bungarotoxin). Key findingsWe determined that psychological stress in mice impairs barrier permeability function and recovery, an effect that is reversed by application of the α7 selective nAChR antagonist, α-bungarotoxin (Bung). In the absence of stress, both topical nicotine or Bung treatment alone impaired barrier permeability. We further observed that stress, topical nicotine, or topical Bung treatment in mice influenced the abundance and/or localization of filaggrin, loricrin, and involucrin. Similar alterations in these three major cornified envelope proteins were observed in human EpiDerm™ cultures. SignificancePerceived psychological stress and nicotine usage can both initiate or exacerbate several dermatoses by altering the cutaneous permeability barrier. Modulation of nAChRs by topical agonists or antagonists may be used to improve epidermal barrier function in skin diseases associated with defects in epidermal barrier permeability.

TH2 cytokines increase kallikrein 7 expression and function in patients with atopic dermatitis

Here we report that Th2 cytokines increase kallikrein 7 expression and function in keratinocytes. Our finding describes a link between the Th2 environment and epidermal barrier dysfunction in atopic dermatitis.

Efficacy of Combined Peroxisome Proliferator-Activated Receptor-α Ligand and Glucocorticoid Therapy in a Murine Model of Atopic Dermatitis

Although topical glucocorticoids (GCs) display potent anti-inflammatory activity in inflamed skin, they also can exert numerous harmful effects on epidermal structure and function. In contrast, topical applications of ligands of peroxisome proliferator-activated receptor-α (PPARα) not only reduce inflammation, and also improve cutaneous barrier homeostasis. Therefore, we examined whether sequential topical GCs followed by topical Wy14643 (a ligand of PPARα) might be more effective than either alone for atopic dermatitis (AD) in a hapten (oxazolone)-induced, murine model with multiple features of AD (Ox-AD). Despite expected anti-inflammatory benefits, topical GC alone induced: i) epidermal thinning; ii) reduced expression of involucrin, loricrin and filaggrin; and iii) allowed outside-to-inside penetration of an epicutaneous tracer. While Wy14643 alone yielded significant therapeutic benefits in mice with mild or moderate Ox-AD, it was less effective in severe Ox-AD. Yet, topical applications of Wy14643 after GC was not only significantly effective comparable to GC alone, but it also prevented GC-induced structural and functional abnormalities in permeability barrier homeostasis. Moreover, rebound flares were largely absent after sequential treatment with GC and Wy14643. Together, these results show that GC and PPARα ligand therapy together is not only effective but also prevents development of GC-induced side effects, including rebound flares, in murine AD.

Expression and regulation of GPAT isoforms in cultured human keratinocytes and rodent epidermis

Phospholipids are required for epidermal lamellar body formation. Glycerol 3-phosphate acyltransferases (GPATs) catalyze the initial step in the biosynthesis of glycerolipids. Little is known about the expression and regulation of GPATs in epidermis/keratinocytes. Here, we demonstrate that GPAT 1, 3, and 4 are expressed in epidermis/keratinocytes, whereas GPAT2 is not detected. In mouse epidermis, GPAT 3 and 4 are mainly localized to the upper layers whereas GPAT1 is found in both the upper and lower layers. GPAT1 and 3 mRNA increase during fetal rat epidermal development. No change in GPAT expression was observed in adult mice following acute permeability barrier disruption. Calcium-induced human keratinocyte differentiation increased GPAT3 mRNA whereas both GPAT1 and 4 mRNA levels decreased. In parallel, total GPAT activity increased 2-fold in differentiated keratinocytes attributable to an increase in N-ethylmaleimide (NEM) sensitive GPAT activity localized to microsomes with little change in NEM resistant activity, consistent with an increase in GPAT3. Furthermore, PPARγ or PPARδ activators increased GPAT3 mRNA, microsomal GPAT activity, and glycerol lipid synthesis without affecting the expression of GPAT1 or 4. Finally, both PPARγ and PPARδ activators increased GPAT3 mRNA via increasing its transcription. Thus, multiple isoforms of GPAT are expressed and differentially regulated in epidermis/keratinocytes.

Activators of PPARs and LXR decrease the adverse effects of exogenous glucocorticoids on the epidermis

While glucocorticoids (GC) exert beneficial effects (anti-inflammatory), they also have adverse effects on the epidermis including decreased epidermal differentiation, decreased keratinocyte proliferation, and decreased cutaneous permeability barrier homeostasis. Thus, the purpose of this study was to develop strategies to prevent these GC toxicities using simultaneous topical treatments in clobetasol-treated mice. While a triple-lipid mixture of stratum corneum lipids (ceramide, free fatty acid and cholesterol) was previously shown to reverse the GC-induced abnormality in cutaneous barrier function [J Invest Dermatol, 120 (2003) 456], this lipid mixture did not prevent the GC-induced abnormalities in either keratinocyte proliferation or differentiation. As activators of PPARalpha, beta/delta, gamma and LXR, regulate keratinocyte proliferation and differentiation and improve permeability barrier homeostasis, we next assessed the effects of these activators during concurrent GC treatment. Co-application of either ciglitazone (PPARgamma activator), clofibrate (PPARalpha activator) or 22R (OH) cholesterol (LXR activator) with clobetasol prevented the decrease in involucrin, filaggrin and loricrin expression. By contrast, a PPARbeta/delta activator (GW501516) normalized only the expression of involucrin and filaggrin but not loricrin. Moreover, topical application of PPARalpha, beta/delta or LXR activators partially prevented the decrease in keratinocyte proliferation in GC-treated murine skin, as measured using PCNA, while no effect was seen after co-treatment with PPARgamma activators. Finally, PPARgamma and PPARbeta/delta activators but not PPARalpha and LXR activators improved permeability barrier homeostasis in GC-treated mice. Together, these studies demonstrate that PPAR and LXR activators can prevent several of the adverse effects of topical GC on the epidermis.

Deficiency of PPARβ/δ in the Epidermis Results in Defective Cutaneous Permeability Barrier Homeostasis and Increased Inflammation

In cultured human keratinocytes or murine epidermis, peroxisome proliferator-activated receptor beta/delta (PPARbeta/delta) (NR1C2) activators (1) stimulate keratinocyte differentiation; (2) decrease keratinocyte proliferation; (3) accelerate permeability barrier repair; (4) increase epidermal lipid synthesis; and (5) reduce cutaneous inflammation. Since these results suggest that PPARbeta/delta could play an important role in cutaneous homeostasis, we assessed here the skin phenotype of mice deficient in PPARbeta/delta. Gross cutaneous abnormalities were not evident, and both stratum corneum (SC) skin hydration and surface pH were normal. However, the epidermis was thickened and proliferating cell nuclear antigen (PCNA) staining was increased, indicating increased cell proliferation. No change in apoptosis was observed but the expression of differentiation markers, such as filaggrin, involucrin, and loricrin, was slightly increased in PPARbeta/delta(-/-) mice. Although basal permeability barrier function was normal, PPARbeta/delta knockout (KO) mice show a significant delay in barrier recovery rates following acute barrier disruption by either acetone treatment or tape-stripping. Delayed barrier recovery correlated with decreased production and secretion of lamellar bodies (LBs), and with reduced numbers of extracellular lamellar membranes in the SC. Finally, PPARbeta/delta KO mice displayed increased inflammation in response to 12-O-tetradecanoylphorbol-13-acetate (TPA) treatment. Together, these results further demonstrate that PPARbeta/delta in the epidermis: (1) is required for permeability barrier homeostasis; (2) regulates keratinocyte proliferation; and (3) modulates cutaneous inflammation.

Thematic review series: Skin Lipids. The role of epidermal lipids in cutaneous permeability barrier homeostasis

The permeability barrier is required for terrestrial life and is localized to the stratum corneum, where extracellular lipid membranes inhibit water movement. The lipids that constitute the extracellular matrix have a unique composition and are 50% ceramides, 25% cholesterol, and 15% free fatty acids. Essential fatty acid deficiency results in abnormalities in stratum corneum structure function. The lipids are delivered to the extracellular space by the secretion of lamellar bodies, which contain phospholipids, glucosylceramides, sphingomyelin, cholesterol, and enzymes. In the extracellular space, the lamellar body lipids are metabolized by enzymes to the lipids that form the lamellar membranes. The lipids contained in the lamellar bodies are derived from both epidermal lipid synthesis and extracutaneous sources. Inhibition of cholesterol, fatty acid, ceramide, or glucosylceramide synthesis adversely affects lamellar body formation, thereby impairing barrier homeostasis. Studies have further shown that the elongation and desaturation of fatty acids is also required for barrier homeostasis. The mechanisms that mediate the uptake of extracutaneous lipids by the epidermis are unknown, but keratinocytes express LDL and scavenger receptor class B type 1, fatty acid transport proteins, and CD36. Topical application of physiologic lipids can improve permeability barrier homeostasis and has been useful in the treatment of cutaneous disorders.

Regulation of ABCA1 expression in human keratinocytes and murine epidermis

Keratinocytes require abundant cholesterol for cutaneous permeability barrier function; hence, the regulation of cholesterol homeostasis is of great importance. ABCA1 is a membrane transporter responsible for cholesterol efflux and plays a pivotal role in regulating cellular cholesterol levels. We demonstrate that ABCA1 is expressed in cultured human keratinocytes (CHKs) and murine epidermis. Liver X receptor (LXR) activation markedly stimulates ABCA1 mRNA and protein levels in CHKs and mouse epidermis. In addition to LXR, activators of peroxisome proliferator-activated receptor (PPAR)alpha, PPARbeta/delta, and retinoid X receptor (RXR), but neither PPARgamma nor retinoic acid receptor, also increase ABCA1 expression in CHKs. Increases in cholesterol supply induced by LDL or mevalonate stimulate ABCA1 expression, whereas inhibiting cholesterol synthesis with statins or cholesterol sulfate decreases ABCA1 expression in CHKs. After acute permeability barrier disruption by either tape-stripping or acetone treatment, ABCA1 expression declines, and this attenuates cellular cholesterol efflux, making more cholesterol available for regeneration of the barrier. In addition, during fetal epidermal development, ABCA1 expression decreases at days 18-22 of gestation (term = 22 days), leaving more cholesterol available during the critical period of barrier formation. Together, our results show that ABCA1 is expressed in keratinocytes, where it is negatively regulated by a decrease in cellular cholesterol levels or altered permeability barrier requirements and positively regulated by activators of LXR, PPARs, and RXR or increases in cellular cholesterol levels.

Heavy Water Labeling of Keratin as a Non-Invasive Biomarker of Skin Turnover In Vivo in Rodents and Humans

Measurement of skin turnover has been problematic in humans. Heavy water (2H2O) labeling has recently been developed as a safe, simple method to study in vivo kinetics of many biosynthetic processes, including DNA and protein synthesis. Here, we apply this approach to the measurement of 2H incorporation into skin keratin and show close agreement between keratin and keratinocyte turnover data in the epidermis of rodents. Elevated turnover rates of both keratin and keratinocytes were observed in the epidermis of the flaky skin mouse, although topical treatments effective in human psoriasis had no effect on either turnover rate in these mice. In humans, keratin turnover was monitored non-invasively by serial tape stripping during and after 2H2O labeling. Kinetic data were consistent with previous estimates of epidermal turnover, with a lag time of 18 days before label appeared at the skin surface and a transit time of 4-5 weeks. Variability in skin keratin turnover rates was present among healthy individuals. In summary, 2H2O labeling of skin keratin represents a non-invasive approach for assessing skin turnover dynamics in pre-clinical models and in human subjects.

Expression and regulation of 1-acyl-sn-glycerol- 3-phosphate acyltransferases in the epidermis

Phospholipids are a major class of lipids in epidermis, where they serve as a source of free fatty acids that are important for the maintenance of epidermal permeability barrier function. The phospholipid biosynthetic enzyme, 1-acyl-sn-glycerol-3-phosphate acyltransferase (AGPAT), catalyzes the acylation of lysophosphatidic acid to form phosphatidic acid, the major precursor of all glycerolipids. We identified an expression pattern of AGPAT isoforms that is unique to epidermis, with relatively high constitutive expression of mouse AGPAT (mAGPAT) 3, 4, and 5 but low constitutive expression of mAGPAT 1 and 2. Localization studies indicate that all five isoforms of AGPAT were expressed in all nucleated layers of epidermis. Furthermore, rat AGPAT 2 and 5 mRNAs increased in parallel with both an increase in enzyme activity and permeability barrier formation late in rat epidermal development. Moreover, after two methods of acute permeability barrier disruption, mAGPAT 1, 2, and 3 mRNA levels increased rapidly and were sustained for at least 24 h. In parallel with the increase in mRNA levels, an increase in AGPAT activity also occurred. Because upregulation of mAGPAT mRNAs after tape-stripping could be partially reversed by artificial barrier restoration by occlusion, these studies suggest that an increase in the expression of AGPATs is linked to barrier requirements.

Interleukin-4 Suppresses the Enhancement of Ceramide Synthesis and Cutaneous Permeability Barrier Functions Induced by Tumor Necrosis Factor-α and interferon-γ in Human Epidermis

Ceramide is an integral part of the extracellular lipid bilayer of the stratum corneum (SC) that forms the permeability barrier of the skin. The production of SC ceramides is catalyzed by sphingomyelinase (SMase) and glucocerebrosidase (GCase). Acid-ceramidase (acid-CDase) catalyzes the hydrolysis of ceramide in the SC. We examined the effects of T helper (Th)1 and Th2 cytokines on levels of transcripts of genes for acid-CDase, acid-SMase, and GCase, on levels of ceramide, and on the extent of transepidermal water loss (TEWL) in the human epidermis in an effort to determine whether these cytokines affect the permeability barrier functions. Levels of transcripts for acid-SMase and GCase and the amount of ceramide in human epidermal sheets were enhanced by tumor necrosis factor (TNF)-alpha and interferon (IFN)-gamma and these effects were inhibited in the presence of interleukin (IL)-4. In epidermal keratinocytes cultured under submerged conditions, however, no similar inhibitory effects of IL-4 were observed. Consistent with these results, TEWL was suppressed by TNF-alpha and IFN-gamma, and these effects were also inhibited by IL-4. The balance between Th1 and Th2 might affect the construction and/or the repair of the epidermal permeability barrier via regulation of the production of ceramide.