De rm at op at ho lo gy The epidermal barrier function and antimicrobial peptides in atopic dermatitis : the role of topical modulation

Introduction The epidermal barrier is a crucial component of the defensive function of the skin. The main skin protective functions are related to water loss modulation/prevention, UV-protection, anti-oxidant and antimicrobial actions. It is well established that epidermal barrier function is abnormal in subjects suffering from atopic dermatitis. Several evidences have shown that this alteration could be considered as the initial pathogenic factor starting the pathological process of atopic dermatitis. The aim of this review was to discuss the role of topical modulation in the epidermal barrier function and antimicrobial peptides in atopic dermatitis. Conclusion The use of emollients has shown to reduce the risk of atopic dermatitis flares and to reduce the need for topical corticosteroid. New emollient and moisturizing products seem promising not only in controlling symptoms and signs of atopic dermatitis but also in improving the skin barrier functions acting specifically in the ceramide component and in normalising antimicrobial peptides production of keratynocites. Introduction Atopic dermatitis (AD) is characterised by reduced skin hydration and an impaired skin barrier mainly due to a deficit in moisturizing property, alteration in lipid content and in the production of antimicrobial substances1–4. It is important to note that in AD subjects’ epidermal barrier dysfunction could be observed also in the skin not involved in active lesions5. Skin barrier defect in AD is mainly due to reduced lipid (i.e. ceramide) content of the epidermis; a reduced or genetically altered filaggrin synthesis and finally a reduced synthesis of antimicrobial peptides (AMPs)6. The deficit of the innate immunity could explain the increased risk of pathogenic bacterial colonisation and infection observed in AD subjects7. For these reasons, treatments with the aim to improve skin barrier properties of AD subjects could be a relevant approach in the strategic treatment of this skin condition. In view of the fact that AD is a chronic condition, caring for atopic skin must be a day-to-day task. Efficient skin care may reduce acute flares by improving the compromised skin barrier and reducing trans-epidermal water loss (TEWL)8. The mainstay of basic AD management, supported by several international guidelines, is the regular use of moisturisers together with good skin hydration and the avoidance of known triggers9. New moisturising products could contain additional compounds helping in the recovery of the normal skin barrier (i.e. ceramide topical supplementation) or showing anti-inflammatory action10. When used as mono-therapy, the use of moisturising creams is associated with a clinical improvement in AD. A particular interest could be found in the use of topical anti-inflammatory and moisturising products containing compounds, which could improve the innate immunological system of the skin such as isoleucine. Topical isoleucine has shown to increase production at the skin level of AMPs11. Therefore, the use of these kinds of products could have a strong rationale in AD coadiuvant treatment. In patients with mild to moderate AD chronic lesion, the use of non-steroidal cream containing Lisoleucine has shown to significantly reduce the eczema area and severity index (EASI) score of children with AD of the face with a clinical efficacy significantly greater than the control emollient cream12. These data, therefore, suggest that it could be possible to modulate and improve the defective skin barrier system in AD patients through topical specific compounds. This paper discusses topical modulation in the epidermal barrier function and AMPs in AD. Discussion Epidermal barrier alteration in AD: focus on filaggrin The superficial layer of the skin (epidermis) plays an important role in several physiologic critical functions such as control and modulation of water loss, antimicrobial defence, the hydration of skin, UV defence, antioxidant defence and the formation of a mechanical barrier13. The normal epidermal barrier is formed mainly by flattened keratinocytes, holding together by corneodesmosomes, a water resistant layer of lipid lamellae which prevent water loss, and finally by natural moisturizing factor, a mixture of aminoacids, urea and other substances, which are contained inside the keratinocyte cell. It is well known that atopic dermatitis is a clinical condition characterised by a deficit of the barrier function of the * Corresponding author Email: masmilan2000@yahoo.it Via A. Nota 18, 20126 Milan, Italy


Introduction
Atopic dermatitis (AD) is characterised by reduced skin hydration and an impaired skin barrier mainly due to a deficit in moisturizing property, alteration in lipid content and in the production of antimicrobial substances [1][2][3][4] .It is important to note that in AD subjects' epidermal barrier dysfunction could be observed also in the skin not involved in active lesions 5 .Skin barrier defect in AD is mainly due to reduced lipid (i.e.ceramide) content of the epidermis; a reduced or genetically altered filaggrin synthesis and finally a reduced synthesis of antimicrobial peptides (AMPs) 6 .The deficit of the innate immunity could explain the increased risk of pathogenic bacterial colonisation and infection observed in AD subjects 7 .For these reasons, treatments with the aim to improve skin barrier properties of AD subjects could be a relevant approach in the strategic treatment of this skin condition.In view of the fact that AD is a chronic condition, caring for atopic skin must be a day-to-day task.Efficient skin care may reduce acute flares by improving the compromised skin barrier and reducing trans-epidermal water loss (TEWL) 8 .The mainstay of basic AD management, supported by several international guidelines, is the regular use of moisturisers together with good skin hydration and the avoidance of known triggers 9 .New moisturising products could contain additional compounds helping in the recovery of the normal skin barrier (i.e.ceramide topical supplementation) or showing anti-inflammatory action 10 .When used as mono-therapy, the use of moisturising creams is associated with a clinical improvement in AD.A particular interest could be found in the use of topical anti-inflammatory and moisturising products containing compounds, which could improve the innate immunological system of the skin such as isoleucine.Topical isoleucine has shown to increase production at the skin level of AMPs 11 .Therefore, the use of these kinds of products could have a strong rationale in AD coadiuvant treatment.In patients with mild to moderate AD chronic lesion, the use of non-steroidal cream containing Lisoleucine has shown to significantly reduce the eczema area and severity index (EASI) score of children with AD of the face with a clinical efficacy significantly greater than the control emollient cream 12 .These data, therefore, suggest that it could be possible to modulate and improve the defective skin barrier system in AD patients through topical specific compounds.This paper discusses topical modulation in the epidermal barrier function and AMPs in AD.

Epidermal barrier alteration in AD: focus on filaggrin
The superficial layer of the skin (epidermis) plays an important role in several physiologic critical functions such as control and modulation of water loss, antimicrobial defence, the hydration of skin, UV defence, antioxidant defence and the formation of a mechanical barrier 13

Epidermal barrier alteration in AD: the role of moisturizing topical treatment
Caring for atopic skin must be continuous as long as the disease is a chronic condition.Efficient skin care may reduce acute flares by improving the compromised skin barrier and reducing TEWL.The mainstay of basic AD management, supported by several international guidelines, is the regular use of moisturisers together with good skin hydration and the avoidance of known triggers.As stated by international guidelines, emollients should form the basis of atopic eczema management and should always be used, even when the atopic eczema is clear.This statement is supported by several clinical studies demonstrating the therapeutic role of emollient in AD.The multicentre barrier enhancement for eczema prevention (BEEP study) has shown that once-daily application of an emollient from birth through to the age of 6 months is able to prevent atopic dermatitis flares 33 .The 6-month cumulative incidence of eczema was 21.8% in the emollient group, in comparison with 43.3% in the control group.This means a 67% reduction in risk.In children with moderate AD, Szczepanowska et al. 34 have evaluated whether adding emollients to the standard topical corticosteroid therapy could influence the clinical outcome.Concomitant usage of emollients significantly improves xerosis and pruritus during corticosteroid treatment of atopic dermatitis, and enables to maintain clinical improvement after therapy discontinuation.Therefore, dry-skin care is very beneficial for patients with AD reducing trans-epidermal water loss and reducing skin barrier compromise.
Antimicrobial peptides in AD: is it possible to modulate their production with topical product?Pharmacological induction of AMPs at epithelial barriers could have and have a capacity to inhibit the growth of microbes and/or direct to kill micro-organisms.AMPs in general are a sequence of 10-50 aminoacids containing positive charges and hydrophobic parts.Defensins are AMPs that are considered as a prototype family of mediators of innate immunity.Defensins have efficient antimicrobial activity on a broad spectrum of micro-organisms 27 .In particular, ß-defensins present in virtually all epithelial surfaces, including those of the skin, airways, gut and urogenital tracts, contribute to the antimicrobial barrier at the epithelial surface 28 .The filaggrin deficiency observed in AD patients seems to be responsible for the reduction in the synthesis of AMP such as the defensin and cathelicidin 29 .Impairment of defencing functions increases the susceptibility to infection.Subjects with AD have a reduced synthesis of AMP and as a consequence AD is characterised by Staphylococcus aureus colonisation and recurrent skin infections 30 .However, the role of AMP in AD is still controversial.Harder et al. 31  skin 14 .Dryness of the skin, caused by an increased trans-epidermal water loss, is in fact a hallmark of patients with AD 15 .Furthermore, the disease is considered to be initiated by the impairment of the skin barrier function.In fact, as pointed out by Elias et al. 16 , breakdown of the skin barrier could be considered the initial event in the development of AD.This alteration is exemplified by loss-of-function mutations in the filaggrin (FLG) gene observed in a relevant percentage of atopic eczema patients 17 (but not in the entire population of AD patients 18 ).Filaggrin is considered as the major structural protein in the skin stratum corneum 19 .Filaggrin is a pivotal protein modulating several functions of the skin and in particular contributing to the formation of skin natural moisturising factor and also in regulating skin pH, alteration in the synthesis of filaggrin or the defective production of ceramides, which are considered as main pathophysiological mechanisms responsible of skin barrier alteration observed in this pathology 20 .When the skin barrier is compromised, the penetration of antigens, pathogens and other nonspecified irritants could be facilitated 21 .This could lead to the activation of inflammatory processes, which from a clinical point of view are characterised by flare appearance.In addition, patients with a genetic defect in filaggrin production/expression present an increased risk of allergic diseases such as asthma 22,23 , contact eczema 24 and latex allergy 25 .

Epidermal barrier alteration in AD: focus on antimicrobial peptides deficiency
As an antimicrobial barrier against invading micro-organisms, skin acts as a physical barrier but also produces AMPs and proteins, including human defensins and cathelicidins, 26   and this alteration even exists in non-lesional skin.In addition, barrier dysfunction results in an increase in protein allergen penetration through the epidermis and predisposes to secondary skin infections, commonly seen in AD.This is supported by the fact that the majority (i.e.80%) of AD patients classified as "no-atopic" subsequently showed an increase in IgE,

Conclusion
Atopic eczema (AE) is a very common skin condition mainly seen in the paediatric population.AD affects 1 out of 10 children, and is associated with significant morbidity.AD is characterised by a chronic impairment of skin barrier functions.Defective permeability and defective antimicrobial barrier are in fact a well-known aspect of AD.Skin barrier alteration and a reduction of innate immune mechanisms (low production of AMPs) are considered the hallmarks of AE.It is recognised that skin barrier dysfunction precedes eczema development, have demonstrated that after superficial skin injury an enhanced expression and secretion of human beta-defensins and psoriasin is observed in AD.Bacterial adhesion and colonisation of the skin is a complex phenomenon and different mechanisms could be involved.Skin xerosis and the alteration in the innate immunity are probably responsible for increased bacterial adhesion and the increased susceptibility of the skin of the person with AD to develop bacterial infections 7 .Even if AD is non-infective skin disease, flare-ups could be related to changes in cutaneous microbes.The penetration of bacterial antigens (superantigens), facilitated by the lack of a defective skin barrier, is viewed as another mechanism of aggravation of the DA.Ong et al. have shown that a deficiency in the expression of AMPs may account for the susceptibility of patients with atopic dermatitis to skin infection with S. aureus 32 .
developing a "true" AD.Skin barrier alteration in AD is manifested mainly with xerosis, inflammation and an increase in tendency to pathogen bacteria colonisation and infections due to the innate immune mechanisms alteration.From a molecular point of view, defective epidermal barrier function is correlated, at least in 1 out of 3 AD patients, with reduced or defective synthesis of the filamentaggregating protein filaggrin.Other alterations responsible of the skin barrier defect could be found in a reduction of ceramide levels in the skin and increased levels of endogenous proteolytic enzymes.These alterations cause an increased transepidermal water loss.Skin xerosis is a common clinical manifestation, together with eczema, of AD.Daily emollient treatment is considered a mainstay of AD treatment alone or in combination with other therapies.The use of emollients has shown to reduce the risk of AD flares and to reduce the need for topical corticosteroid.New emollient and moisturising products seem promising not only in controlling symptoms and signs of AD but also in improving the skin barrier functions acting specifically in the ceramide component and in normalising AMP production of keratynocites.therapeutic utility.It has been recently reported that the discovery of substances with low molecular weight can induce epithelial antimicrobial peptide production in cell-based assays.In particular L-isoleucine and its analogues are highly specific ß-defensin inducers in epithelial cells.In 2000, Fehlbaum et al.35 have demonstrated that L-isoleucine, in a dose-dependent manner, could induce defensin production in AMPs constitute an important component of the mammalian innate immune response.These results were confirmed by Santiago et al.In an animal model, they found that Isoleucine is an inducer of Beta-defensin production.This action of Isoleucine was observed when the aminoacid was applied directly on thracheal epithelia mucosa.Therefore, Isoleucine could be considered an AMP-promoter at skin level.In a recent multicentre trial conducted in children with mild-moderate AD of the face, Marseglia et al.12  have shown that a topical anti-inflammatory moisturising cream containing rhamnosoft, ceramides and L-Isoleucine was able to significantly reduce the EASI score by 71% in comparison with baseline after 6 weeks of treatment.This clinical effect was statistically superior to the effect on EASI score obtained in the control group treated with a simple emollient cream.