Topical hesperidin prevents glucocorticoid‐induced abnormalities in epidermal barrier function in murine skin

Systemic and topical glucocorticoids (GC) can cause significant adverse effects not only on the dermis, but also on epidermal structure and function. In epidermis, a striking GC‐induced alteration in permeability barrier function occurs that can be attributed to an inhibition of epidermal mitogenesis, differentiation and lipid production. As prior studies in normal hairless mice demonstrated that topical applications of a flavonoid ingredient found in citrus, hesperidin, improve epidermal barrier function by stimulating epidermal proliferation and differentiation, we assessed here whether its topical applications could prevent GC‐induced changes in epidermal function in murine skin and the basis for such effects. When hairless mice were co‐treated topically with GC and 2% hesperidin twice‐daily for 9 days, hesperidin co‐applications prevented the expected GC‐induced impairments of epidermal permeability barrier homoeostasis and stratum corneum (SC) acidification. These preventive effects could be attributed to a significant increase in filaggrin expression, enhanced epidermal β‐glucocerebrosidase activity and accelerated lamellar bilayer maturation, the last two likely attributable to a hesperidin‐induced reduction in stratum corneum pH. Furthermore, co‐applications of hesperidin with GC largely prevented the expected GC‐induced inhibition of epidermal proliferation. Finally, topical hesperidin increased epidermal glutathione reductase mRNA expression, which could counteract multiple functional negative effects of GC on epidermis. Together, these results show that topical hesperidin prevents GC‐induced epidermal side effects by divergent mechanisms.     

Topical apigenin improves epidermal permeability barrier homoeostasis in normal murine skin by divergent mechanisms

The beneficial effects of certain herbal medicines on cutaneous function have been appreciated for centuries. Among these agents, chrysanthemum extract, apigenin, has been used for skin care, particularly in China, for millennia. However, the underlying mechanisms by which apigenin benefits the skin are not known. In this study, we first determined whether topical apigenin positively influences permeability barrier homoeostasis, and then the basis thereof. Hairless mice were treated topically with either 0.1% apigenin or vehicle alone twice daily for 9 days. At the end of the treatments, permeability barrier function was assessed with either an electrolytic water analyzer or a Tewameter. Our results show that topical apigenin significantly enhanced permeability barrier homoeostasis after tape stripping, although basal permeability barrier function remained unchanged. Improved barrier function correlated with enhanced filaggrin expression and lamellar body production, which was paralleled by elevated mRNA levels for the epidermal ABCA12. The mRNA levels for key lipid synthetic enzymes also were upregulated by apigenin. Finally, both cathelicidin‐related peptide and mouse beta‐defensin 3 immunostaining were increased by apigenin. We conclude that topical apigenin improves epidermal permeability barrier function by stimulating epidermal differentiation, lipid synthesis and secretion, as well as cutaneous antimicrobial peptide production. Apigenin could be useful for the prevention and treatment of skin disorders characterized by permeability barrier dysfunction, associated with reduced filaggrin levels and impaired antimicrobial defenses, such as atopic dermatitis.     

Mite and cockroach allergens activate protease-activated receptor 2 and delay epidermal permeability barrier recovery

Protease-activated receptor-2 (PAR-2) is known to be involved in epidermal permeability barrier function homeostasis. PAR-2 activation occurs after barrier disruption and further activation of PAR-2 by activating peptide significantly delays barrier recovery rate. Cockroach and house dust mite allergens, both known to be associated with the development of asthma, allergic rhinitis, and atopic dermatitis, have protease activity, which can activate PAR-2. In this study, we investigated the effects of both allergens on the epidermal barrier function as well as on the epidermal calcium gradient. Both allergens, when topically applied on the barrier-disrupted site, increased protease activities in the epidermis and delayed barrier recovery and lamellar body secretion in murine skin. The topical application of PAR-2-specific antagonist or protease inhibitors normalized the barrier recovery. Cockroach allergens induced intracellular calcium oscillations in cultured human keratinocytes through PAR-2-involved pathway, which was confirmed by desensitization protocol. Abnormal calcium ion distribution after barrier disruption was also observed in allergens-applied skin. These results suggest that allergens with protease activity can influence the epidermal permeability barrier homeostasis through PAR-2 activation and consequent modulation of the calcium ions in skin.     

Testosterone perturbs epidermal permeability barrier homeostasis

Although there are no known gender-related differences in permeability barrier function in adults, estrogens accelerate whereas testosterone retards barrier development in fetal skin, and male fetuses demonstrate slower barrier development than female littermates. Moreover, prenatal administration of the androgen receptor antagonist, flutamide, equalizes developmental rates in male and female fetuses. Therefore, we evaluated the effects of changes in testosterone on barrier homeostasis in adult murine and human skin. Hypogonadal mice (whether by castration or by treatment with systemic flutamide) displayed significantly faster barrier recovery at 3, 6, and 12 h than did controls, and testosterone replacement slowed barrier recovery in castrated mice. Moreover, testosterone directly effects the skin, as topical flutamide also accelerated barrier recovery in normal male mice. These findings appear to be of physiologic significance, since prepubertal male mice (age 5 wk) displayed accelerated barrier recovery in comparison with adult postpubertal (11 wk) males. These studies also appear to be relevant for humans, as a hypopituitary human subject demonstrated repeated changes in barrier recovery in parallel with peaks and nadirs in serum testosterone levels during intermittent testosterone replacement. Mechanistic studies showed that differences in epidermal lipid synthesis do not account for the testosterone-induced functional alterations. Instead, epidermal lamellar body (LB) formation and secretion both decrease, resulting in decreased extracellular lamellar bilayers in testosterone-replete animals. These studies demonstrate that fluctuations in testosterone modulate barrier function, and that testosterone repletion can have negative consequences for permeability barrier homeostasis.     

Role of PKC-delta as a signal mediator in epidermal barrier homeostasis

The skin shows an important "epidermal permeability barrier homeostasis" in response to barrier disruption. Calcium ion (Ca(2+)), a major regulator in keratinocyte differentiation and proliferation, plays a crucial role in skin barrier homeostasis. Acute barrier disruption induces an immediate depletion of both extra- and intracellular calcium ions in the epidermis, especially in the upper granular layers, and results in the loss of normal epidermal calcium gradient. Currently, we hypothesize that the change in the intracellular calcium ion concentration triggers the barrier repair responses, such as lamellar body (LB) secretion and increased lipid synthesis in the epidermis. In this article, we suggest that PKC-delta is a signaling mediator for the changes in extracellular and intracellular calcium ion concentration.     

Serine protease signaling of epidermal permeability barrier homeostasis

Evidence is growing that protease-activated receptor-2 (PAR-2) plays a key role in epithelial inflammation. We hypothesized here that PAR-2 plays a central role in epidermal permeability barrier homeostasis by mediating signaling from serine proteases (SP) in the stratum corneum (SC). Since the SC contains tryptic- and chymotryptic-like activity, we assessed the influence of SP activation/inhibition on barrier function. Acute barrier disruption increases SP activity and blockade by topical SP inhibitors (SPI) accelerates barrier recovery after acute abrogation. This improvement in barrier function is due to accelerated lamellar body (LB) secretion. Since tryptic SP signal certain downstream responses through PAR-2, we assessed its potential role in mediating the negative effects of SP on permeability barrier. Firstly, PAR-2 is expressed in the outer nucleated layers of the epidermis and most specifically under basal condition to the lipid raft (LR) domains. Secondly, tape stripping-induced barrier abrogation provokes PAR-2 activation, as shown by receptor internalization (i.e. receptor movement from LR to cytolpasmic domains). Thirdly, topical applications of PAR-2 agonist peptide, SLIGRL, delay permeability barrier recovery and inhibit LB secretion, while, conversely, PAR-2 knockout mice display accelerated barrier recovery kinetics and enhanced LB secretion, paralleled by increased LR formation and caveolin-1 expression. These results demonstrate first, the importance of SP/SPI balance for normal permeability barrier homeostasis, and second, they identify PAR-2 as a novel signaling mechanism of permeability barrier, that is, of response linked to LB secretion.     

Barrier function regulates epidermal lipid and DNA synthesis

The stratum corneum, the permeability barrier between the internal milieu and the environment, is composed of fibrous protein-enriched corneocytes and a lipid-enriched intercellular matrix. The lipids are a mixture of sphingolipids, cholesterol and free fatty acids, which form intercellular membrane bilayers. Lipid synthesis occurs in the keratinocytes in all nucleated layers of the epidermis, and the newly synthesized lipids are delivered by lamellar bodies to the interstices of the stratum corneum during epidermal differentiation. Disruption of barrier function by topical acetone treatment results in an increase in the synthesis of free fatty acids, sphingolipids and cholesterol in the living layers of the epidermis, leading to barrier repair. Cholesterol and sphingolipid synthesis are regulated by the rate-limiting enzymes HMG CoA reductase and serine palmitoyi transferase (SPT). respectively. Acute barrier disruption leads to an increase in both enzymes, but with a different time curve: increase in HMG CoA reductase activity begins at 1.5 h, whereas the increase in SPT activity occurs 6 h after barrier impairment. Topical application of HMG CoA reductase or SPT inhibitors after acetone treatment delays barrier repair, providing further evidence for a role of cholesterol and sphingolipids in epidermal barrier function. Repeated application of lovastatin to untreated skin results in disturbed barrier function accompanied by increased DNA synthesis and epidermal hyperplasia. Therefore, we have examined the specific relationship between barrier function and epidermal DNA synthesis. After acute and chronic disturbances not only lipid, but also DNA synthesis, is stimulated. Thus, stimulation of DNA synthesis leading to epidermal hyperplasia may be a second mechanism by which the epidermis repairs defects in barrier function. The link between barrier function and both lipid and DNA synthesis is supported further by occlusion studies. Artificial barrier repair by latex occlusion prevents an increase in both lipid and DNA synthesis. In addition, increased epidermal lipid and DNA synthesis in essential fatty-acid deficiency can be reversed by topical applications of the n-6 unsaturated fatty acids, linoleic or columbinic acid. These studies may be of relevance in understanding the pathogenesis of hyperproliferative skin diseases, such as ichthyosis, psoriasis, atopic dermatitis, and irritant contact dermatitis.     

Mechanisms by which psychologic stress alters cutaneous permeability barrier homeostasis and stratum corneum integrity

Although many skin disorders, including psoriasis and atopic dermatitis, are adversely affected by psychologic stress (PS), the pathophysiologic link between PS and disease expression remains unclear. Recent studies demonstrated PS-induced alterations in permeability barrier homeostasis, mediated by increased endogenous glucocorticoids. Here, we assessed the mechanisms by which PS alters stratum corneum (SC) function. Insomniac psychologic stress (IPS) altered both barrier homeostasis and SC integrity. IPS decreased epidermal cell proliferation, impaired epidermal differentiation, and decreased the density and size of corneodesmosomes (CD), which was linked to degradation of CD proteins (e.g., desmoglein1). Barrier compromise was linked to decreased production and secretion of lamellar bodies (LB), which in turn could be attributed to a decrease in de novo synthesis of epidermal lipids. Topical physiologic lipids (equimolar cholesterol, ceramides, and free fatty acids) normalized both barrier homeostasis and SC integrity in IPS mice, further evidence that lipid deficiency accounted for these functional abnormalities. Thus, PS inhibition of epidermal lipid synthesis results in decreased LB formation and secretion, as well as decreased CD, compromising both permeability barrier homeostasis and SC integrity. These studies suggest that topical treatment with epidermal physiologic lipids could be beneficial in stress-induced, barrier-associated dermatoses, such as psoriasis and atopic dermatitis.     

Cannabinoid receptors 1 and 2 oppositely regulate epidermal permeability barrier status and differentiation

Cannabinoid receptors (CBR) 1 and 2 have been implicated in keratinocyte differentiation/proliferation. How CB receptors affect epidermal permeability barrier and stratum corneum structure and function remains unclear. Permeability barrier abrogation was induced by sequential tape-stripping of the SC and assessed in both CB1R and CB2R knockout (-/-) mice in comparison with wild-type (+/+) littermates. Absence of CB1R delays permeability barrier recovery, while the latter was found to be accelerated in CB2R -/- mice. While increased lamellar body (LB) secretion is observed in CB2R -/- mice accounting for the enhanced recovery, CB1R -/- animals display strong alterations in lipid bilayer structures. Markers for epidermal differentiation (i.e. filaggrin, loricrin and involucrin) and terminal differentiation (i.e. TUNEL assay and caspase-14 activation) were respectively decreased and increased in CB1R and CB2R -/- mice. Surprisingly, CB1R agonist treatment of human cultured keratinocytes increases mRNA of p21 and cytokeratin 1 and 10 and decreases cyclin D1 but protein levels remained unchanged. Such paradox could partially be explained by the increase in non-phosphorylated-4E-BP1, an inhibitor of mRNA translation, following CB1R agonist treatment. Altogether, these observations put forward the importance and the complexity of cannabinoid signalling for the regulation of permeability barrier and epidermal differentiation.     

Deficiency of PPARbeta/delta 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.     

Skin barrier function, epidermal proliferation and differentiation in eczema

Skin permeability barrier function is impaired in eczema, particularly in contact and atopic dermatitis (AD). In contact dermatitis disruption of the barrier by irritants and allergens is the primary event, followed by sensitization, inflammation, increased epidermal proliferation and changes in differentiation. Genetically impaired skin barrier function is already present in non-lesional and more pronounced in lesional skin in AD. Increased epidermal proliferation and disturbed differentiation, including changes in lipid composition, cause impaired barrier function in AD. Defective permeability barrier function enables the enhanced penetration of environmental allergens into the skin and initiates immunological reactions and inflammation. Barrier dysfunction is therefore crucially involved in the pathogenesis of AD. The atopic syndrome represents a genetically impaired skin barrier function as well as impaired nasal, bronchial, and intestinal mucous membranes leading to AD, allergic rhinitis, bronchial asthma or aggravation of AD. Common treatment strategies for eczema include the application of lipid-based creams and ointments, which aim toward the restoration of the defective permeability barrier, thus helping to normalize proliferation and differentiation.     

Stimulation of PPARalpha promotes epidermal keratinocyte differentiation in vivo

Our recent studies have demonstrated that PPARalpha activators stimulate differentiation and inhibit proliferation in cultured human keratinocytes and accelerate epidermal development and permeability barrier formation in fetal rat skin explants. As the role of PPARalpha activation in adult epidermis is not known, the aim of this study was to determine if topically applied PPARalpha ligands regulate keratinocyte differentiation in murine epidermis. Topical treatment with PPARalpha activators resulted in decreased epidermal thickness. Expression of structural proteins of the upper spinous/granular layers (involucrin, profilaggrin-filaggrin, loricrin) increased following topical treatment with PPARalpha activators. Furthermore, topically applied PPARalpha activators also increased apoptosis, decreased cell proliferation, and accelerated recovery of barrier function following acute barrier abrogation. Experiments with PPARalpha-/- knockout mice showed that these effects are specifically mediated via PPARalpha. Compared with the epidermis of PPARalpha+/+ mice, involucrin, profilaggrin-filaggrin, and loricrin expression were slightly decreased in PPARalpha-/- mice. Moreover, topical clofibrate treatment did not increase epidermal differentiation in PPARalpha-/- mice. Furthermore, in cultured human keratinocytes we have demonstrated that PPARalpha activators induce an increase in involucrin mRNA levels. We have also shown that this increase in gene expression requires an intact AP-1 response element at -2117 to -2111 bp. Thus, stimulation of PPARalpha stimulates keratinocyte/epidermal differentiation and inhibits proliferation.     

Impaired sphingomyelinase activity and epidermal differentiation in atopic dermatitis

A defective permeability barrier leads to the penetration of environmental allergens into the skin and initiates immunological reactions and inflammation crucially involved in the pathogenesis of atopic dermatitis (AD). Decreased stratum corneum ceramide content may cause the defect in permeability barrier function consistently found in AD. Acid and neutral sphingomyelinase (A- and N-SMase) generate ceramides with structural and signal transduction functions in epidermal proliferation and differentiation. We determined epidermal SMase activities, DNA synthesis, involucrin, loricrin, filaggrin, and keratin expression in lesional and non-lesional skin of AD patients. We found decreased epidermal A-SMase activity in lesional and non-lesional skin, correlating with reduced stratum corneum ceramide content and disturbed barrier function. N-SMase activity was reduced in non-lesional skin and more significantly reduced in lesional skin, correlating with impaired expression of cornified envelope proteins and keratins, important for skin barrier function. Changes in involucrin, loricrin, filaggrin, keratin K 5 (basal) and K 16 (proliferation associated) were noticed in non-lesional and lesional skin, whereas changes in K 10 (suprabasal), K 6 (proliferation associated), and K 17 (inflammation associated) were found only in lesional skin. In summary, reduction in SMase-generating ceramides and impaired differentiation are involved in the defective barrier function found in AD.     

Modulations in epidermal calcium regulate the expression of differentiation-specific markers

Mammalian epidermis normally displays a distinctive calcium gradient, with low levels in the basal/spinous layers and high levels in the stratum granulosum. Although changes in stratum granulosum calcium regulate the lamellar body secretory response to permeability barrier alterations, whether modulations in calcium also regulate the expression of differentiation-specific proteins in vivo remains unknown. As acute barrier perturbations reduce calcium levels in stratum granulosum, we studied the regulation of murine epidermal differentiation after loss of calcium accompanying acute barrier disruption and by exposure of such acutely perturbed skin sites to either low (0.03 M) or high (1.8 M) calcium. Three hours after acute barrier disruption, coincident with reduced calcium and ultrastructural evidence of accelerated lamellar body secretion, both northern analyses and in situ hybridization revealed decreased mRNA levels for loricrin, profilaggrin, and involucrin in the outer epidermis, but protein levels did not change significantly. Moreover, exposure of acutely disrupted skin sites to low calcium solutions sustained the reduction in mRNA levels, whereas exposure to high calcium solutions restored normal mRNA levels (blocked by the L-type calcium channel inhibitor, nifedipine). Finally, with prolonged exposure to a low (<10% relative humidity) or high (>80% relative humidity) humidity, calcium levels increased and declined, respectively. Accordingly, mRNA and protein levels of the differentiation-specific markers increased and decreased at low and high relative humidity, respectively. These results provide direct evidence that acute and sustained fluctuations in epidermal calcium regulate expression of differentiation-specific proteins in vivo, and demonstrate that modulations in epidermal calcium coordinately regulate events late in epidermal differentiation that together form the barrier.     

Barrier function, epidermal differentiation, and human beta-defensin 2 expression in tinea corporis

Tinea corporis is a superficial mycotic infection resulting in substantial epidermal changes. We determined skin barrier function, epidermal differentiation, and human-beta-defensin 2 (hBD-2) protein expression in 10 patients with tinea corporis caused by Trichophyton rubrum (T. rubrum). We found disturbed skin barrier function as shown by a significant increase in transepidermal water loss (TEWL) and specific ultrastructural changes including disturbed formation of extracellular lipid bilayers, lamellar body extrusion, and deposit of clotted material at the stratum granulosum/stratum corneum interface. Epidermal proliferation in tinea increased several fold and accordingly, proliferation and inflammation-associated keratins K6, K16, and K17 were expressed. Expression of basal keratins K5 and K14 increased, whereas differentiation-associated K10 was reduced. Reduction of the cornified envelope proteins involucrin, loricrin, and the S100 protein filaggrin was also seen. Reduced filaggrin expression correlated with reduced skin hydration; protein breakdown products of filaggrin have been shown to be important for water binding. Surprisingly, we found pronounced epidermal protein expression of hBD-2, which may be related to disturbed epidermal differentiation and inflammation. hBD-2 showed a weak, although significant, antifungal activity against T. rubrum in the turbidimetric assay and the immunohistological staining was somewhat less pronounced in areas directly underneath fungal hyphae in the stratum corneum. Together, we describe profound changes in skin barrier structure and function, epidermal proliferation, and differentiation including pronounced protein expression of hBD-2 in tinea corporis.     

Structural basis for the barrier abnormality following inhibition of HMG CoA reductase in murine epidermis.

Recent studies have shown that increased epidermal 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG CoA) reductase activity is crucial for the barrier recovery response that follows solvent-induced barrier perturbation. Upregulation of this enzyme leads to increased cholesterologenesis, formation and secretion of cholesterol-enriched lamellar bodies, and barrier repair. Topical lovastatin-induced inhibition of HMG CoA reductase activity both delays the acute barrier-repair response, as well as leading to a chronic barrier abnormality when applied repeatedly to intact skin. Presently, we assessed the effects of repeated topical applications of two different specific inhibitors of HMG CoA reductase on barrier function, the lamellar body-secretory system, and stratum corneum intercellular domains, with functional and morphologic parameters. Once-daily applications of lovastatin or fluindostatin (XU62-320; Sandoz) for 4-8 d to intact hairless mouse epidermis produced a progressive abnormality in barrier function (transepidermal water loss greater than 2.0-5.0 in treated versus less than 0.25 mg/cm2/h for weakly active analogues or vehicle controls). The barrier defect was preceded by alterations in lamellar body internal structure and a partial failure of lamellar body secretion into the stratum corneum interstices, further confirmed by enzyme cytochemistry. Moreover, the deposition of abnormal lamellar body contents resulted in the formation of clefts in the intercellular spaces at the stratum granulosum-stratum corneum interface, resulting in increased permeability through these domains shown by lanthanum perfusion. Applications of irritants, even when producing a barrier abnormality, did not alter the lamellar body secretory system. Co-applications of cholesterol with the inhibitors reversed both the barrier abnormality and the abnormalities in the lamellar body secretory system that occur with the inhibitor alone. Finally, membrane bilayer structures in the mid-to-outer stratum corneum of inhibitor-treated specimens appeared normal, but the intercellular domains displayed enormously expanded lacunae. However, because similar dilatations also occurred in vehicle-treated samples, they can be attributed to the vehicle alone. These studies provide further evidence that the inhibitor-induced defect in barrier function a) is initiated by inhibition of HMG CoA reductase; b) can be attributed to defects in both lamellar body structure and deposition with resultant abnormalities in intercellular membrane domains in the lower stratum corneum; and c) is further enhanced by permissive effects of the vehicle on the permeability of the outer stratum corneum.     

Effect of endothelial nitric oxide synthase on epidermal permeability barrier recovery after disruption

Background We previously demonstrated that neuronal nitric oxide synthase (nNOS) in epidermal keratinocytes is associated with epidermal permeability barrier homeostasis. Objectives In the present study, we examined the contributions of inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) to epidermal permeability barrier homeostasis. Methods We measured the barrier recovery rate after tape stripping of the epidermis of iNOS and eNOS knockout mice, and carried out electron‐microscopic observation of the epidermis after acetone treatment. Results The barrier recovery rate of eNOS knockout mice was significantly faster than that of the wild‐type control, while no significant difference was observed between iNOS knockout mice and wild‐type mice. Electron‐microscopic observation at 1 h after acetone treatment indicated that barrier recovery of both nNOS and eNOS mice was faster than that of wild‐type mice, and lamellar body secretion was accelerated in both types of knockout mice. Conclusions These results suggested that both nNOS and eNOS play roles in epidermal barrier homeostasis and lamellar body secretion.     

Simultaneous effect of ursolic acid and oleanolic acid on epidermal permeability barrier function and epidermal keratinocyte differentiation via peroxisome proliferator-activated receptor-alpha

Ursolic acid (UA) and oleanolic acid (ONA) are pentacyclic triterpenoids, which naturally occur in many medicinal herbs and plants. Recent research revealed that several pharmacological effects could be attributed to UA and ONA, such as anti-tumor, anti-inflammatory and anti-microbial activities. To evaluate the effects of UA and ONA on epidermal permeability barrier recovery and normal skin, both flanks of hairless mice were topically treated with either 0.01-0.1 mg/mL UA or 0.1-1.0 mg/mL ONA after tape stripping and transepidermal water loss (TEWL) were assessed, and then hydration and TEWL were measured for 3 weeks with application of UA and ONA (2 mg/mL). We also investigated the morphological changes using light (LM) and electron microscopic (EM) examination. Finally, we observed that UA and ONA stimulated epidermal keratinocyte differentiation via peroxisome proliferator-activated receptor (PPAR)-alpha using Western immunoblotting. The recovery rate of epidermal permeability barrier after tape stripping increased in the UA- and ONA-treated groups (0.1 mg/mL UA and 0.5 mg/mL ONA) at 6 h to more than 20% when compared to the vehicle-treated group (P < 0.05). In both groups, hydration was increased compared to the vehicle group from 1 week without TEWL alteration (P < 0.05). An LM finding showed that epidermal thickening was frequently observed (UA > ONA > vehicle). EM examination revealed an increase in secretion and in the number of lamellar bodies in treated groups and that complete formation of lipid bilayers was also prominent (ONA > UA > vehicle). Protein expression of PPAR-alpha, involucrin, loricrin and filaggrin increased twofold and threefold in HaCaT cells treated for 24 h with either ONA (10 micromol/L) or UA (10 micromol/L), respectively, reflecting that the UA and ONA can improve the recovery of skin barrier function and induce epidermal keratinocyte differentiation via PPAR-alpha. Taken together, these results suggest that UA and ONA will be pertinent candidates for the improvement of epidermal permeability barrier function.     

表皮通透的屏障功能及其调节

保护机体、防止过多的水分丢失是皮肤的主要功能之一,角质层角质形成细胞及其细胞间脂类的组分对此功能起决定作用。角质形成细胞间的复层板层膜是皮肤屏障功能的主要结构。表皮通透的屏障功能影响皮肤的水分丢失量,且对表皮及真皮的生物活性(如角质形成细胞的增生、皮肤的炎症等)均有调节作用。诸多因素如脂类、性别、年龄、酸碱度、离子及精神等均可以作用于表皮通透的屏障功能。因此,深入地了解表皮通透的屏障功能及其影响因素有助于皮肤病的防治。