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EVALUATION OF DAILY PHOTOPROTECTION USING RECONSTRUCTED SKIN MODEL :  Relevance of UVA absorption profile and SPF value
 

EVALUATION OF DAILY PHOTOPROTECTION USING RECONSTRUCTED SKIN MODEL : Relevance of UVA absorption profile and SPF value

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One of the major long term consequences of chronic non extreme sun exposures is the development of photoaging. The recent definition of a standard daily ultraviolet radiation (DUVR) spectrum has ...

One of the major long term consequences of chronic non extreme sun exposures is the development of photoaging. The recent definition of a standard daily ultraviolet radiation (DUVR) spectrum has allowed us to reproduce non-zenithal sun exposure conditions. Exposure to simulated DUVR induces biological damage in human skin, suggesting the need for an appropriate daily photoprotection. The reconstructed human skin in vitro, including a living dermal equivalent and a fully differentiated epidermis represents a predictive tool to study the effects of UV. In the present study, sunscreen products were evaluated after topical application on human reconstructed skin in vitro. Two commercial sunscreens (A and B) having similar SPF value (~15) but different profile of transmission over the UVA range were tested on skin models exposed to increasing doses of DUVR. Another pair of sunscreens was also tested. Product C had a SPF ~18 with a well-balanced UVB/UVA absorption profile and product D, with SPF ~27 and low UVA absorption. Biological parameters were assessed by (i) histology, (ii) immunostaining of dermal fibroblasts and (iii) MMP-1 production. Products A and C gave a better protection from DUVR with regard to fibroblast alterations and MMP-1 release compared to products B and D respectively. These results support that a well balanced profile of absorption and the level of UVA protection is of higher relevance than the SPF value to ensure an efficient daily photoprotection from DUVR with regard to end points evaluated in this study, especially the photoaging related biological markers.

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    EVALUATION OF DAILY PHOTOPROTECTION USING RECONSTRUCTED SKIN MODEL :  Relevance of UVA absorption profile and SPF value EVALUATION OF DAILY PHOTOPROTECTION USING RECONSTRUCTED SKIN MODEL : Relevance of UVA absorption profile and SPF value Presentation Transcript

    • EVALUATION OF DAILY PHOTOPROTECTION USINGRECONSTRUCTED SKIN MODEL :Relevance of UVA absorption profile and SPF value F Lejeune, F Christiaens and F BernerdL’Oré al Research & Innovation, Clichy, Chevilly-Larue, France
    • ABSTRACT One of the major long term consequences of chronic non extreme sun exposures is the development of photoaging. The recent definition of a standard daily ultraviolet radiation (DUVR) spectrum has allowed us to reproduce non-zenithal sun exposure conditions. Exposure to simulated DUVR induces biological damage in human skin, suggesting the need for an appropriate daily photoprotection. The reconstructed human skin in vitro, including a living dermal equivalent and a fully differentiated epidermis represents a predictive tool to study the effects of UV. In the present study, sunscreen products were evaluated after topical application on human reconstructed skin in vitro. Two commercial sunscreens (A and B) having similar SPF value (~15) but different profile of transmission over the UVA range were tested on skin models exposed to increasing doses of DUVR. Another pair of sunscreens was also tested. Product C had a SPF ~18 with a well- balanced UVB-UVA absorption profile and product D, with SPF ~27 and low UVA absorption. Biological parameters were assessed by (i) histology, (ii) immunostaining of dermal fibroblasts and (iii) MMP-1 production. Products A and C gave a better protection from DUVR with regard to fibroblast alterations and MMP-1 release compared to products B and D respectively. These results support that a well balanced profile of absorption and the level of UVA protection is of higher relevance than the SPF value to ensure an efficient daily photoprotection from DUVR with regard to end points evaluated in this study, especially the photoaging related biological markers.
    • INTRODUCTIONOne of the major long term consequences of chronic sun exposure is the development of photoaging clinical signsincluding wrinkles and sagging. These deleterious events mostly result from alterations of extracellular matrix andfibroblasts in the dermal compartment. It is now well recognized that UVA wavelengths play a major role inphotoaging process indicating a predominant involvement of sub-erythemal sun exposures. It is therefore ofimportance to design the most efficient photoprotection for all kinds of solar exposure.The reconstructed human skin in vitro which includes a living dermal equivalent and a fully differentiated epidermisrepresents a highly reliable predictive model to characterize wavelength and cell type specific biological damagetogether with tissue distribution. Using this 3D skin model, we previously showed that while UVB directly affectedepidermis with sunburn related biological damage (1), UVA radiation directly impacted the dermal compartmentthrough fibroblast impairment and extracellular matrix modifications, including MMP-1 production (2).Based on the identification of these biological end-points, evaluation of sunscreen efficiency could be assessed.Test products can be topically applied to the skin model alike in vivo due to the presence of horny layers (3).OBJECTIVEHere we studied the efficiency of two pairs of sunscreen products (A/B, and C/D) having differentcharacteristics. o The pair A/B was composed of two commercial sunscreen products. They have the same labeled SPF of 15 but different UVA absorption profile. o The pair C/D was composed as follow: Suncreen C had a SPF of 15 and a UVA-PF of 10 as measured by the PPD method. Suncreen D had a SPF of 30 and a UVA-PF of 4.Test product was applied to the surface of reconstructed skin which was then exposed to increasing dosesof daily UV radiation (DUVR).
    • MATERIAL AND METHODS1 Cells : Human fibroblasts and keratinocytes were obtained from normal human breast skin2 Reconstructed human skin in vitro Dermal equivalents were prepared with 106 dermal fibroblasts and bovine type I collagen. Epidermal keratinocytes were seeded at 33000/cm². The culture was performed during 7 days in submerged conditions, then raised on a grid at the air liquid interface for subsequent 7 days of culture. Culture medium was changed every 2 days. KERATINOCYTES 3 days 7days 7d Collagen, Fibroblasts Dermal equivalent Seeding of Normal human skin « Monolayered Epidermis » Air-liquid interface Medium Keratinocytes Epidermal differentiation Human skin in vitro 0,5 SSR 0,4 Daily UV (DUVR) Experimental source for Daily UV (ref 4) 0,33 UV source 0,2 SSR (Solar Simulated Radiation) ( UVA/UVB = 12 ) Erythemal exposure DUVR (Daily UV) UVA/UVB = 27. Non erythemal 0,1 exposure 290 300 310 320 330 340 350 360 370 380 390 400 Wavelength (nm)
    • MATERIAL AND METHODS4 Sunscreen products and treatment : Products C and D were prepared in the same vehicle (oil in water emulsion). Sunscreen products or vehicle were applied topically at ~ 2mg/cm² 10 minutes prior UV exposure. Samples were then rinsed and analyzed 48 hours after UV exposure for classical histology. Culture medium were collected and MMP1 protein level was assessed using ELISA (Amersham). Product A Product B Product A 0.5 Product BAvobenzone : 2% Octinoxate : 6% 0.4Terephthalylidene Dicamphor Sulfonic Acid : 2% Zinc Oxide : 3% Absorbance(Mexoryl SX ™) 0.3 (AU)Octrocrylene 10 % 0.2Drometrizole Trisiloxane 1% 0.1 0 300 320 340 360 380 Wavelength (nm) 400 1 Product C Product D Product C 0,8Butyl Methoxydibenzoylmethane: 3% Octocrylene : 2,5% Product D AbsorbanceTerephthalylidene Dicamphor Sulfonic Acid : 3% Ethylhexyl Methoxycinnamate 0,6Octrocrylene 5% : 7,5%Drometrizole Trisiloxane 1% Ethylhexyl Salicylate: 2,5% 0,4 Zinc Oxide : 6% 0,2 SPF 18 ± 2 27 ± 2 0 290 300 310 320 330 340 350 360 370 380 390 400 UVA PF (PPD) 10 ± 2 4±1 Wavelength (nm)
    • RESULTS 1 Effects of DUVR exposure Histology Control 12 J/cm² DUVR 30 J/cm² DUVR A B C Vimentin D E F Visualization of DUVR induced damage in human reconstructed skin Histological section and vimentin immunostaining allowed to revealed that DUVR exposure induced the disappearance of dermal fibroblasts and alterations of epidermal granular layers at 48 h.
    • RESULTS 2 Evaluation of products A and B Histology of reconstructed skins treated Vimentin staining of reconstructed with sunscreen products A and B skins treated with sunscreen products A and B 48 hours post exposure to DUVR, topical application of sunscreen product A could prevent morphological alterations induced by exposure to increasing doses of DUVR, i.e. disappearance of dermal fibroblasts and modifications of epidermal granular layers, while this protection was not afforded by sunscreen product B.
    • RESULTS 3 Evaluation of products A and B 50 40 MMP1 ng/ml 30 20 10 0 0J/cm² 12J/cm² 0J/cm² 30J/cm² 50J/cm² 0J/cm² 30J/cm² 50J/cm² No product Sunscreen A Sunscreen B MMP1 production
    • RESULTS 4 Evaluation of products C and D Histology of reconstructed skins treated with sunscreen products C and D 48 hours post exposure to DUVR, topical application of sunscreen product C could prevent morphologicalalterations induced by exposure to increasing doses of DUVR, i.e. disappearance of dermal fibroblasts and modifications of epidermal granular layers, while this protection was not afforded by sunscreen product D.
    • RESULTS 5 Evaluation of products C and D MMP1 ng/ml MMP1 ng/ml 40 40 35 35 30 30 25 25 20 20 15 15 10 10 5 5 0 0 0J/cm² 30J/cm² 0J/cm² 30J/cm² 0J/cm² 30J/cm² 0J/cm² 30J/cm² Vehicle Product C Vehicle Product D MMP1 production
    • CONCLUSIONS Our results showed that product A afforded a better protection against biological damage mostly located within the dermal equivalent and previously identified after UVA exposure compared to product B.  These results indicate that the SPF value is not sufficient for evaluation of daily UV protection and emphasize the need for a high UVA absorption with regard to the prevention of photoaging-related biological end-points. Our results showed that product C afforded a better protection against biological damage mostly located within the dermal equivalent compared to product D. These results indicate that absorption profile (well balanced UVB/UVA level of protection) is of higher relevance than SPF value for daily UV photoprotection as regards the prevention of UV-induced photodamage.REFERENCE LIST 1- Bernerd F. and Asselineau D. Successive alteration and recovery of epidermal differentiation and morphogenesis after specific UVB damages in skin reconstructed in vitro. Dev. Biol., 1997, 183:123-138 2- Bernerd F. and Asselineau D. UVA exposure of human skin reconstructed in vitro induces apoptosis of dermal fibroblasts: subsequent connective tissue repair and implications in photoaging. Cell Death Diff., 1998, 5:792-802 3- Bernerd F, Vioux C, Lejeune F. and Asselineau D. The sun protection factor (SPF) inadequately defines broad spectrum photoprotection : demonstration using skin reconstructed in vitro exposed to UVA, UVB or UV-solar simulated radiation. Eur. J. Dermatol. 2003, 13: 242-249 4- Christiaens F, Chardon A, Fourtanier A, Frederick JE. Standard ultraviolet daylight for non-extreme exposure conditions. Photochem Photobiol. 2005, 81(4):874-8.