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160 In vivo investigation of stratum corneum thickness and epidermal barrier structure: Links with inflammatory status
ABSTRACTS | Epidermal Structure and Function 160
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In vivo investigation of stratum corneum thickness and epidermal barrier structure: Links with inflammatory status M Dumas2, J Franchi2, A Bernois2, M Juan2, E Leblanc2, CH Heusele2, A Bensussan1, M Bagot1,3, S Schnebert2 and L Michel1 1 Dermatology, INSERM U976, Paris, France, 2 LVMH RECHERCHE, Saint-Jean de Braye, France and 3 Dermatology, APHP Saint-Louis, Paris, France The stratum corneum (SC) is the first protection to environmental stress and a key interface for skin hydration and bacterial invasion. The links between SC modifications and inflammatory status in healthy skin have not been precisely investigated. The present study was carried out on 23 healthy Caucasian women aged from 30 to 40 years with dry or normal skin. Biopsies were characterized histologically, immunostained for barrier function markers, analyzed by qPCR for inflammatory proteins, and skin surface lipid extracted with hexane/ethanol for HPTLC quantification. We observed that fineness of the SC was correlated with low expression of epidermal desmoglein 1 and transglutaminase 1 both involved in SC cohesion and resistance since other epidermal proteins such as filaggrin, protein zona occludens 1, aquaporins 3, 9, 10, and clusterin were not affected. This relative cutaneous fragility was correlated with increased interleukin (IL-8), and macrophage migration inhibitory factor (MIF) since other mediators e.g. IL-6 and IL-13, arachidonate 15-lipoxygenase, peroxisome proliferator-activated receptor beta, tumor necrosis factor alpha, toll-like receptor 4 and thymic stromal lymphopoietin were non-affected. In addition, MIF expression was inversely correlated with free fatty acids and ceramids such as AP (CER 6) and EOH (CER 4) types and not with other SC lipids e.g. cholesterol, triglyceride, ceramids AP, EOS, NH/AS, NP and NS types nor with total ceramids indicating the possible relation between SC lipids composition and cutaneous inflammation. Furthermore, a low expression of beta-defensin-2 correlated with high expression of IL-8 suggests that the innate antimicrobial defense could also participate in skin reactivity. This study highlights the major role of both MIF and IL-8 in the inflammation status linked to physical and biological barrier failure and fragility.
Interest of 10-Hydroxy-2-decenoic acid and acefylline-containing-creams for hydration and nutrition of dry skin M Galliano1, C Carrasco1, V Mengeaud2, S Bessou-Touya1 and H Duplan1 1 TOULOUSE FRANCE, PIERRE FABRE DERMO-COSMETIQUE, Toulouse, France and 2 Laboratoires Dermatologiques Ducray, Lavaur, France Filaggrin proteolysis and catabolism is an important cue that allows optimal hydration of the stratum corneum (SC), by generating the natural moisturizing factor (NMF). During this process, peptidylarginine deiminases catalyse the citrullination of filaggrin, which improves its degradation. We previously showed that 10-Hydroxy-2-decenoic acid, a natural fatty acid, induced filaggrin protein production while the xanthine derivative acefylline enhanced peptidylarginine deiminase activities. Here we investigated the effect of acefylline on filaggrin production and degradation on reconstructed human epidermis (RHE) generated under dryness condition. We observed that RHE responded to dryness by upregulation of both profilaggrin and proteases involved in filaggrin catabolism, leading to an increase of NMF components and citrulline residues. Addition of acefylline further promoted filaggrin degradation as shown by a decrease of filaggrin monomers, an increase of filaggrin catabolites. We then investigated the effect of cosmetic creams containing 10-Hydroxy-2-decenoic acid or acefylline on lipids by measuring the neosynthesis of lipids in RHE and by using the highresolution X-microdiffraction technique on SC. We found that treatment of RHE with the active ingredients-containing-creams showed a significant modulation of the neosynthesis of ceramides involved in the reinforcement of the lipid barrier. X-microdiffraction analysis showed that the active ingredients-containing-creams penetrated into the SC and positively influenced the lipids organization either by enhancing hexagonal organization in the outer SC or by increasing the orthorhombic structure. Those observations suggest a reinforcement of the lipid barrier. In conclusion, our study supports the benefits of treatment of dry skin with adapted formulations containing 10-Hydroxy-2-decenoic acid and acefylline to correct hydration and favour nutrition of the skin.
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In vitro cortisol exposure leads to a stress-induced increased expression of skin aging markers A Van Summeren, M Yu, J Sun, C Pollefliet, H Corstjens and L Declercq BSR, The Este´e Lauder Companies, Oevel, Belgium With today’s busy, modern lifestyle, it is difficult to find the right balance between work and life. This lack of balance often causes stress. It is commonly accepted that psychological stress affect the appearance of facial skin. Previously, we identified panelists who have been chronically exposed to stress, mainly associated with a lack of sleep and a busy lifestyle. According to these panelists, this stress caused tired looking skin. Indeed, these panelists exhibited more wrinkles, a greater extent of skin laxity, and less skin radiance than their nonstressed counterparts of similar gender and age. It is known that as a response to stress, cortisol is released through the hypothalamic-pituitary-adrenal axis. A prolonged exposure to stress will result in persistent elevated levels of cortisol. This study was set up to evaluate the in vitro impact of cortisol, as a mimic of chronic stress, on the skin. Fibroblasts were exposed to cortisol and afterwards populated in a free floating collagen lattice. The contractility of fibroblasts decreased in a dose dependent manner. In addition, a GlasboxÒ system was used to measure the contractile forces exerted by the cells real-time. Once again, these results indicated that cortisol inhibited the cellematrix construction and disrupted fibroblast mechanical tension. To further investigate the effect of stress on skin, ex vivo skin explants were chronically exposed to cortisol. These skin explants were characterized by epidermal atrophy, an increased expression of the senescent markers p21 and progerin, and a decrease in the protein expression of elastin and fibronectin. In conclusion, in vitro conditions mimicking the impact of chronic stress on skin cells by prolonged exposure to cortisol, revealed modifications of the contractile forces of fibroblasts, increases in senescence markers and decreases in markers related to elasticity. These cortisol induced alterations might contribute to the visible signs of tired looking skin, and have been shown here to induce skin changes consistent with premature aging.
Skin microbiota alteration link to skin symptoms after a harsh cleanser AG Gueniche, C Clavaud, O Perin, D Bernard, M Thomas, F Benech, M Bayer-Vanmoen, B Renault and F Cabirol LOREAL RESEARCH AND INNOVATION, AULNAY SOUS BOIS, France Topical skin cleansers are commonly used products to clean the skin. To investigate the time needed for skin and its microbiota to recover, a group of 30 healthy volunteers from 30 to 50 years old were selected according to both photo ageing and chronological age score. We evaluated the skin before, immediately after, 3h and 6h after cleansing using a harsh soap. Especially, total bacterial load by QPCR (V1-V3 16S rRNA KAPA Biosystems kit with SYBER Green), bacterial community using Next generation sequencing (Amplification V1-V3 on Illumina MiSeq platform) in addition to skin clinical evaluation, self-assessment and instrumental methods were followed during this clinical trial. Instrumental evaluation including skin hydration, barrier function and pH are altered just after cleansing and recover within 6 hours. Clinical expert evaluation and self-assessment revealed that a harsh wash increases dryness, roughness and discomfort and these skin alterations do not recover even after 6 hours. The harsh wash decreased the quantity, diversity and changes the community structure of the skin bacterial communities. Importantly compared to before washing the alterations on skin bacteria after 3 hours are higher (load and beta diversity) than just after washing meaning that the skin bacteria has difficulty to recover. In addition, skin bacteria are not at all recovered after 6h. All this important findings show that it is important to propose a skin routine that will firstly clean by respecting the diversity of skin microbiota secondly will feed the skin microbiota and facilitate their recovery.
S220 Journal of Investigative Dermatology (2017), Volume 137
Epidermal barrier improvement after Excipial application in atopic xerosis is revealed by non-invasive proteome analysis I Carlavan Research, GALDERMA, Biot, France The stratum corneum plays a crucial role in barrier function and is under investigation in several skin pathologies. The stratum corneum (SC) of atopic xerosis (AX) reveals not only decreased hydration but also mildly impaired barrier function characterized by an increase in Trans Epidermal Water Loss (TEWL), elevated pH values, and an increased turnover rate of the SC consisting of thick layers of smaller-sized corneocytes. The mildly impaired SC functions of AX can be improved by daily repeated applications of effective moisturizers, which are effective in preventing the progression of AX to atopic dermatitis (AD). In a recent study, clinical and biophysical observations were in favour of an improvement of skin surface after 8 daily applications of Excipial (anti-itch foam). In AX the effect of moisturizers in general on the stratum corneum protein composition has not yet been fully analysed. Mass spectrometry analysis was performed using protein extracted from stratum corneum collected before and 48h after 8 days of Excipial application. More than 400 proteins were identified in SC. Several proteins were found significantly modulated after Excipial treatment. We found a significant increase in content of proteins involved in cohesion of the SC and its barrier function, such as Desmoplakin and the Keratin type II cytoskeletal 2 and in regulation of epidermal homeostasis, such as filaggrin-2. In addition, we found two abundant unique peptides of serum albumin slightly but significantly decreased after treatment. Using Luminex technology a slight decrease in serum albumin content was observed (fold: -1.5, p <0.05). Interestingly, albumin was previously described up-regulated in AD. Taken together, our proteomics data indicate that skin homeostasis and barrier function in AX was improved at the molecular level after 8 days Excipial application.
Assessment of protective role of novel ceramide microspheres on retinol stabilization and its anti-acne activity M Pasikowska1, R Debowska1, K Cal2, B Ostrowska3, K Rogiewicz1,3 and I Eris1,3 1 Centre for Science and Research Dr Irena Eris, Cosmetic Laboratories Dr Irena Eris, Piaseczno, Poland, 2 Laboratory of Molecule Engineering, Gdansk, Poland and 3 Cosmetic Laboratories Dr Irena Eris, Technology Division, Piaseczno, Poland Retinol is very unstable molecule. Here we present the method to prolongate retinol’s stability by its incorporation to novel protective ceramide microspheres in topical formulations. In addition to this stability and efficacy of topical 0,3% retinol formulation in adult acne treatement was performed. In order to measure guality and quantity of retinol in vivo, three formulations were investigated: RST (retinospheres), R (pure retinol) and P (placebo) by UV fluorescent microscopy as well as UV spectroscopy. Tape-stripping skin samples aftery applying latter formulations from 5 volunteers were collected. Moreover for the in vivo efficacy study of RST formulation, 20 adult patients with acne were enrolled. The skin parameters like porfirines, apperiance of seboceous glands, and skin moisturization were performed by Visiopor PP34 and Corneometer CM 825 (Courage-Khazakha). Assessment of retinol quantity in human stratum coreum revealled that retinol incorporated in microspheres penetrates 2,5-fold more to stratum coreum than native retinol. Also higher amount of retinol in RST formulation indicated increased stability of the molecule comapred to R formulation. Decrease in retinol content in R formulation was 3 times higher that in RST formulation. In vivo stability of retinol was also confirmed by fluorescence microscopy where RST formulation had up to 2 times higher fluorescence intensity compared to R formulation. In vivo efficacy tests on RST formulation revealled that after 4 weeks of cream usage the numer and size of porfirines decresed by 16% and 27% respectively. Moisturization of skin incresed by 20%. Retinol provides multifunctional activity for acne, photodamaged or ageing skin, however in order to provide stability of the molecule it is necessery to develope special microspheres displaying protective effect on retinol.
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In vivo investigation of stratum corneum thickness and epidermal barrier structure: Links with inflammatory status M Dumas2, J Franchi2, A Bernois2, M Juan2, E Leblanc2, CH Heusele2, A Bensussan1, M Bagot1,3, S Schnebert2 and L Michel1 1 Dermatology, INSERM U976, Paris, France, 2 LVMH RECHERCHE, Saint-Jean de Braye, France and 3 Dermatology, APHP Saint-Louis, Paris, France The stratum corneum (SC) is the first protection to environmental stress and a key interface for skin hydration and bacterial invasion. The links between SC modifications and inflammatory status in healthy skin have not been precisely investigated. The present study was carried out on 23 healthy Caucasian women aged from 30 to 40 years with dry or normal skin. Biopsies were characterized histologically, immunostained for barrier function markers, analyzed by qPCR for inflammatory proteins, and skin surface lipid extracted with hexane/ethanol for HPTLC quantification. We observed that fineness of the SC was correlated with low expression of epidermal desmoglein 1 and transglutaminase 1 both involved in SC cohesion and resistance since other epidermal proteins such as filaggrin, protein zona occludens 1, aquaporins 3, 9, 10, and clusterin were not affected. This relative cutaneous fragility was correlated with increased interleukin (IL-8), and macrophage migration inhibitory factor (MIF) since other mediators e.g. IL-6 and IL-13, arachidonate 15-lipoxygenase, peroxisome proliferator-activated receptor beta, tumor necrosis factor alpha, toll-like receptor 4 and thymic stromal lymphopoietin were non-affected. In addition, MIF expression was inversely correlated with free fatty acids and ceramids such as AP (CER 6) and EOH (CER 4) types and not with other SC lipids e.g. cholesterol, triglyceride, ceramids AP, EOS, NH/AS, NP and NS types nor with total ceramids indicating the possible relation between SC lipids composition and cutaneous inflammation. Furthermore, a low expression of beta-defensin-2 correlated with high expression of IL-8 suggests that the innate antimicrobial defense could also participate in skin reactivity. This study highlights the major role of both MIF and IL-8 in the inflammation status linked to physical and biological barrier failure and fragility.
Interest of 10-Hydroxy-2-decenoic acid and acefylline-containing-creams for hydration and nutrition of dry skin M Galliano1, C Carrasco1, V Mengeaud2, S Bessou-Touya1 and H Duplan1 1 TOULOUSE FRANCE, PIERRE FABRE DERMO-COSMETIQUE, Toulouse, France and 2 Laboratoires Dermatologiques Ducray, Lavaur, France Filaggrin proteolysis and catabolism is an important cue that allows optimal hydration of the stratum corneum (SC), by generating the natural moisturizing factor (NMF). During this process, peptidylarginine deiminases catalyse the citrullination of filaggrin, which improves its degradation. We previously showed that 10-Hydroxy-2-decenoic acid, a natural fatty acid, induced filaggrin protein production while the xanthine derivative acefylline enhanced peptidylarginine deiminase activities. Here we investigated the effect of acefylline on filaggrin production and degradation on reconstructed human epidermis (RHE) generated under dryness condition. We observed that RHE responded to dryness by upregulation of both profilaggrin and proteases involved in filaggrin catabolism, leading to an increase of NMF components and citrulline residues. Addition of acefylline further promoted filaggrin degradation as shown by a decrease of filaggrin monomers, an increase of filaggrin catabolites. We then investigated the effect of cosmetic creams containing 10-Hydroxy-2-decenoic acid or acefylline on lipids by measuring the neosynthesis of lipids in RHE and by using the highresolution X-microdiffraction technique on SC. We found that treatment of RHE with the active ingredients-containing-creams showed a significant modulation of the neosynthesis of ceramides involved in the reinforcement of the lipid barrier. X-microdiffraction analysis showed that the active ingredients-containing-creams penetrated into the SC and positively influenced the lipids organization either by enhancing hexagonal organization in the outer SC or by increasing the orthorhombic structure. Those observations suggest a reinforcement of the lipid barrier. In conclusion, our study supports the benefits of treatment of dry skin with adapted formulations containing 10-Hydroxy-2-decenoic acid and acefylline to correct hydration and favour nutrition of the skin.
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In vitro cortisol exposure leads to a stress-induced increased expression of skin aging markers A Van Summeren, M Yu, J Sun, C Pollefliet, H Corstjens and L Declercq BSR, The Este´e Lauder Companies, Oevel, Belgium With today’s busy, modern lifestyle, it is difficult to find the right balance between work and life. This lack of balance often causes stress. It is commonly accepted that psychological stress affect the appearance of facial skin. Previously, we identified panelists who have been chronically exposed to stress, mainly associated with a lack of sleep and a busy lifestyle. According to these panelists, this stress caused tired looking skin. Indeed, these panelists exhibited more wrinkles, a greater extent of skin laxity, and less skin radiance than their nonstressed counterparts of similar gender and age. It is known that as a response to stress, cortisol is released through the hypothalamic-pituitary-adrenal axis. A prolonged exposure to stress will result in persistent elevated levels of cortisol. This study was set up to evaluate the in vitro impact of cortisol, as a mimic of chronic stress, on the skin. Fibroblasts were exposed to cortisol and afterwards populated in a free floating collagen lattice. The contractility of fibroblasts decreased in a dose dependent manner. In addition, a GlasboxÒ system was used to measure the contractile forces exerted by the cells real-time. Once again, these results indicated that cortisol inhibited the cellematrix construction and disrupted fibroblast mechanical tension. To further investigate the effect of stress on skin, ex vivo skin explants were chronically exposed to cortisol. These skin explants were characterized by epidermal atrophy, an increased expression of the senescent markers p21 and progerin, and a decrease in the protein expression of elastin and fibronectin. In conclusion, in vitro conditions mimicking the impact of chronic stress on skin cells by prolonged exposure to cortisol, revealed modifications of the contractile forces of fibroblasts, increases in senescence markers and decreases in markers related to elasticity. These cortisol induced alterations might contribute to the visible signs of tired looking skin, and have been shown here to induce skin changes consistent with premature aging.
Skin microbiota alteration link to skin symptoms after a harsh cleanser AG Gueniche, C Clavaud, O Perin, D Bernard, M Thomas, F Benech, M Bayer-Vanmoen, B Renault and F Cabirol LOREAL RESEARCH AND INNOVATION, AULNAY SOUS BOIS, France Topical skin cleansers are commonly used products to clean the skin. To investigate the time needed for skin and its microbiota to recover, a group of 30 healthy volunteers from 30 to 50 years old were selected according to both photo ageing and chronological age score. We evaluated the skin before, immediately after, 3h and 6h after cleansing using a harsh soap. Especially, total bacterial load by QPCR (V1-V3 16S rRNA KAPA Biosystems kit with SYBER Green), bacterial community using Next generation sequencing (Amplification V1-V3 on Illumina MiSeq platform) in addition to skin clinical evaluation, self-assessment and instrumental methods were followed during this clinical trial. Instrumental evaluation including skin hydration, barrier function and pH are altered just after cleansing and recover within 6 hours. Clinical expert evaluation and self-assessment revealed that a harsh wash increases dryness, roughness and discomfort and these skin alterations do not recover even after 6 hours. The harsh wash decreased the quantity, diversity and changes the community structure of the skin bacterial communities. Importantly compared to before washing the alterations on skin bacteria after 3 hours are higher (load and beta diversity) than just after washing meaning that the skin bacteria has difficulty to recover. In addition, skin bacteria are not at all recovered after 6h. All this important findings show that it is important to propose a skin routine that will firstly clean by respecting the diversity of skin microbiota secondly will feed the skin microbiota and facilitate their recovery.
S220 Journal of Investigative Dermatology (2017), Volume 137
Epidermal barrier improvement after Excipial application in atopic xerosis is revealed by non-invasive proteome analysis I Carlavan Research, GALDERMA, Biot, France The stratum corneum plays a crucial role in barrier function and is under investigation in several skin pathologies. The stratum corneum (SC) of atopic xerosis (AX) reveals not only decreased hydration but also mildly impaired barrier function characterized by an increase in Trans Epidermal Water Loss (TEWL), elevated pH values, and an increased turnover rate of the SC consisting of thick layers of smaller-sized corneocytes. The mildly impaired SC functions of AX can be improved by daily repeated applications of effective moisturizers, which are effective in preventing the progression of AX to atopic dermatitis (AD). In a recent study, clinical and biophysical observations were in favour of an improvement of skin surface after 8 daily applications of Excipial (anti-itch foam). In AX the effect of moisturizers in general on the stratum corneum protein composition has not yet been fully analysed. Mass spectrometry analysis was performed using protein extracted from stratum corneum collected before and 48h after 8 days of Excipial application. More than 400 proteins were identified in SC. Several proteins were found significantly modulated after Excipial treatment. We found a significant increase in content of proteins involved in cohesion of the SC and its barrier function, such as Desmoplakin and the Keratin type II cytoskeletal 2 and in regulation of epidermal homeostasis, such as filaggrin-2. In addition, we found two abundant unique peptides of serum albumin slightly but significantly decreased after treatment. Using Luminex technology a slight decrease in serum albumin content was observed (fold: -1.5, p <0.05). Interestingly, albumin was previously described up-regulated in AD. Taken together, our proteomics data indicate that skin homeostasis and barrier function in AX was improved at the molecular level after 8 days Excipial application.
Assessment of protective role of novel ceramide microspheres on retinol stabilization and its anti-acne activity M Pasikowska1, R Debowska1, K Cal2, B Ostrowska3, K Rogiewicz1,3 and I Eris1,3 1 Centre for Science and Research Dr Irena Eris, Cosmetic Laboratories Dr Irena Eris, Piaseczno, Poland, 2 Laboratory of Molecule Engineering, Gdansk, Poland and 3 Cosmetic Laboratories Dr Irena Eris, Technology Division, Piaseczno, Poland Retinol is very unstable molecule. Here we present the method to prolongate retinol’s stability by its incorporation to novel protective ceramide microspheres in topical formulations. In addition to this stability and efficacy of topical 0,3% retinol formulation in adult acne treatement was performed. In order to measure guality and quantity of retinol in vivo, three formulations were investigated: RST (retinospheres), R (pure retinol) and P (placebo) by UV fluorescent microscopy as well as UV spectroscopy. Tape-stripping skin samples aftery applying latter formulations from 5 volunteers were collected. Moreover for the in vivo efficacy study of RST formulation, 20 adult patients with acne were enrolled. The skin parameters like porfirines, apperiance of seboceous glands, and skin moisturization were performed by Visiopor PP34 and Corneometer CM 825 (Courage-Khazakha). Assessment of retinol quantity in human stratum coreum revealled that retinol incorporated in microspheres penetrates 2,5-fold more to stratum coreum than native retinol. Also higher amount of retinol in RST formulation indicated increased stability of the molecule comapred to R formulation. Decrease in retinol content in R formulation was 3 times higher that in RST formulation. In vivo stability of retinol was also confirmed by fluorescence microscopy where RST formulation had up to 2 times higher fluorescence intensity compared to R formulation. In vivo efficacy tests on RST formulation revealled that after 4 weeks of cream usage the numer and size of porfirines decresed by 16% and 27% respectively. Moisturization of skin incresed by 20%. Retinol provides multifunctional activity for acne, photodamaged or ageing skin, however in order to provide stability of the molecule it is necessery to develope special microspheres displaying protective effect on retinol.