DOI: 10.1111/exd.12388
Review
www.wileyonlinelibrary.com/journal/EXD
New insights in photoaging, UVA induced damage and skin types Claire Battie, Setsuko Jitsukawa, Francßoise Bernerd, Sandra Del Bino, Claire Marionnet and le Verschoore Miche L’Oreal Research and Innovation, Asnieres, France Correspondence: Claire Battie, L’Oreal Research and Innovation, Asnieres, France, Tel.: +33 1 47 56 71 98, e-mail: [email protected] Abstract: UVA radiation is the most prevalent component of solar UV radiation; it deeply penetrates into the skin and induces profound alterations of the dermal connective tissue. In recent years, the detrimental effects of UVA radiation were more precisely demonstrated at cellular and molecular levels, using adequate methods to identify biological targets of UVA radiation and the resulting cascade impairment of cell functions and tissue degradation. In particular gene expression studies recently revealed that UVA radiation induces modulation of several genes confirming the high sensitivity of dermal fibroblasts to UVA radiation. The major visible damaging effects of UVA radiation only appear after years of exposure: it has been clearly evidenced
that they are responsible for more or less early signs of photoageing and photocarcinogenesis. UVA radiation appears to play a key role in pigmented changes occurring with age, the major sign of skin photoaging in Asians. Skin susceptibility to photoaging alterations also depends on constitutive pigmentation. The skin sensitivity to UV light has been demonstrated to be linked to skin color type.
Introduction
around midday. Another interesting characteristic of UVA radiation is that it comes through glass, whereas UVB rays are almost entirely absorbed. Thus, high UVA doses may be received even in indoor conditions while erythemal UVB radiation is filtered out. Seventy percent of UVB radiation that reaches the skin is absorbed by the stratum corneum, 20% reaches viable epidermis, and only 10% penetrates the uppermost part of the dermis. On the other hand, UVA radiation is partly absorbed by the epidermis, but 20–30% of it reaches deep dermis. UVB radiation consequently has a major action on the epidermis compared to UVA radiation which can also target the dermis. That’s one of the reasons why UVA radiation is now considered to be a major factor in the process of skin photoaging but also participates in photocarcinogenesis. The contribution of diffuse UV radiation is also important and should not be underestimated. Indeed a recent study suggests that diffuse irradiation may explain a large part of the cumulative annual exposure dose (3). The consequences of UVA exposure on skin are particularly important in Asia, since UVA exposure appear to be more important in principal cities in Asia compared to Europe because of the low latitude. The calculation of UVA fluence received in Asia relative to that received in Paris on a winter day showed that in Asia, the exposure to UVA light is much more than in Paris (Fig. 1) (2).
Solar UV radiation reaching the earth is a combination of UVB (290–320 nm) and UVA (320–400 nm) wavelengths. Since UVA radiation is less energetic than UVB radiation, UVB radiation has long been thought to be the factor responsible for the damaging effects of solar radiation. But it is now proved that UVA radiation plays a major role. In fact UVA radiation is the most prevalent component of solar UV radiation, it penetrates deeper than UVB radiation into the skin and induces profound alterations of the dermal connective tissue. In recent years, the detrimental effects of UVA radiation were more precisely demonstrated at cellular and molecular levels, using adequate methods to identify biological targets of UVA radiation and the resulting cascade impairment of cell functions and tissue degradation.
Skin photoaging: UVA radiation plays a major role Cutaneous UV exposure UVB radiation reaches the earth in relatively low amounts (about 0.5% of solar spectral irradiance at ground level, integrated over 290–2500 nm range) and is highly energetic. In contrast, UVA rays are lower in energy, but they are at least 20 times more abundant. Ninety five percent of UV rays reaching the ground level are UVA (1). Solar UV irradiance depends on many geo-orbital and environmental parameters including latitude, time of the year (season), hour of the day, meteorological conditions and the thickness of ozone layer. UVA radiation is less affected by those parameters and varied to a lesser extent than UVB radiation. For example UVA irradiance is less affected by seasons, and decreases to a lesser extent in winter (2). The time of the day plays an important role too. Indeed UVA and UVB radiations both raise from the beginning of the day, peak at noon and decrease at the end of the day but UVA radiation is present for most part of the day as it follows the variation of visible light whereas UVB rays are at the highest between 10 a.m. and 4 p.m., especially
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
Key words: photoageing – photoprotection – skin sensitivity to UV – UVA
Accepted for publication 24 March 2014
UVA radiation induces biological alterations related to photoaging – in vivo proofs In vivo studies have now given clear evidence that UVA radiation significantly contributes to long-term dermal structure deterioration and clinical signs of photoaging. It is responsible for the early appearance of signs of photoageing, which include patchy/mottled pigmentation, wrinkling, laxity, sagging, dryness, etc. One of the major modes of action of UVA radiation is the generation of –
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(a)
(b)
Figure 1. UVA dose received in Asian cities compared to that in Paris: Harbin, Beijing, Seoul and Tokyo respectively receive 1.29, 1.84, 1.93 and 1.97 times more UVA doses than Paris.
oxidative stress which is a well known factor in the pathogenesis of photoaging (4). The biological effects of repetitive, suberythemal UVA exposure were explored in vivo (5–7). For example, in a reported clinical study, 14 volunteers were exposed three times per week for 13 weeks to increasing doses of UVA radiation (5). Different biological markers related to skin photoaging were studied: ferritin and tenascin expression as well as lysozyme quantification. The results showed that UVA exposure induced an increased expression of ferritin in the basal and suprabasal keratinocyte layers and in interstitial dermal cells (Fig. 2a). As ferritin is an important marker of antioxidant activity, its increase proved that UVA radiation induced oxidative stress. UVA radiation also induced a marked increase in tenascin expression, just below the dermo-epidermal junction (Fig. 2b). Tenascin is a large glycoprotein of the extracellular matrix, interacting with collagens, proteoglycans and fibronectin. It is expressed at low levels in normal skin while it is upregulated in skin tumours and in a number of skin diseases, which reveals the damaging effects of UVA radiation. UVA radiation induced a significant (P < 0.05) increase in deposit of lysozyme (Fig. 2c). Lysozyme has been shown to be associated with damaged elastic fibers in many tissues (8,9). In skin, lysozyme deposits on elastic fibers are seen in sun-damaged regions, and the number of lysozyme-containing elastin fibers appears to be correlated with the degree of sun damage. It has been suggested that lysozyme binds specifically to damaged elastin fibers and thereby inhibits the proteolytic degradation of altered fibers by elastases (10). This may be linked to solar elastosis. The same study also investigated the effects of repetitive, suberythemal UVA exposure on skin pigmentation. It was shown that this regimen of exposure induced a strong increase in pigmentation. Pigmented changes are particularly important in Asia since it is the major sign of skin photoaging in Asians (11–13). An ethnic group-related variation in melanosome distribution was reported showing a mix of individual (about 60%) and aggregated (about
8
(c)
Figure 2. UVA exposure induces alterations of epidermal and dermal protein expression. Immunofluorescence microscopy using antisera against ferritin (a), tenascin (b) and lysosyme (c).
40%) melanosomes in Asian skins, whereas aggregated melanosomes (85%) are prevailing in European skins (14–19). The density and highly variable size of melanosomes in Asian skins could account for irregular, spotty pigmentation associated with photoageing. It has been proved that in darker-skinned individuals, UVA radiation induces greater pigmenting effects than UVB (20).
UVA radiation induces biological alterations related to photoaging – in vitro proofs In vitro organotypic skin models have been developed providing a three-dimensional tissue structure and a complete epidermal differentiation like in vivo. Such models allow to study the early biological events occurring after a single or repeated UV exposure and to investigate the biological effects of UVA and UVB radiations. Unlike UVB radiation, UVA radiation, due to its high penetration properties, can reach deeper parts of the skin and affects the dermal compartment. In the reconstructed skin model, UVA radiation actually induces major alterations in the dermal compartment through the generation of ROS. As a result of exposure to
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
UVA induced damage and skin types
UVA radiation (25 J/cm²), the dermal fibroblasts located in the upper part of the dermal equivalent disappear within 48 h following exposure through an apoptotic process (Fig. 3) (21). On the other hand, the epidermal structure and organization are not significantly affected, indicating that the survival ability of dermal fibroblasts after exposure to pure UVA radiation is lower compared to that of epidermal keratinocytes. These results confirm previous experiments showing that dermal fibroblasts are more sensitive to UVA-induced oxidative stress than keratinocytes (21,22). The increase in MMP-1 after UV exposure was also observed in reconstructed skin model (23,24). MMP-1 is an interstitial collagenase able to hydrolyze type I collagen, the major component of the dermis, and it seems to play a crucial role in the disorganization and progressive degeneration of dermal extracellular matrix (25–27). Under UVA exposure, MMP-1 production was directly induced in the dermal fibroblasts. Removal of epidermis immediately after UVA exposure did not alter this induction. These results confirmed other data on UVA-induced MMP-1 in cultured fibroblasts (28,29). In contrast, UVB-induced MMP-1 production required the presence of the epidermis. More recently the impact of UVA radiation was studied at the molecular level by evaluating UVA radiation-induced gene expression in reconstructed skin in vitro and in human skin biopsies in vivo. The results showed that the expression of genes related to
oxidative stress response and relevant for photoageing were affected by UVA exposure (30). For example, in vitro and in vivo, UVA exposure induced an increase of MMP-1, haeme oxygenase-1, and superoxide dismutase-2 gene expression. The use of the reconstructed skin model allowed the authors to precise the contribution of fibroblast and keratinocytes in gene expression modulation. Another transcriptomic study on reconstructed skin model (31) focused more particularly on the impact of UVA radiation during a non zenithal UV exposure, occurring in most outdoors activities, called daily UV radiation (DUVR) (32). To characterize the contribution of UVA wavelengths in the biological impact of DUVR, a reconstructed skin model was exposed either to 13 J/cm2 DUVR or to 25 J/cm2 UVA radiation. Gene modulation of several genes related to functional families was quantified (oxidative stress, extracellular matrix, transcription, signaling, heat shock proteins, cytokines). Out of the 60 genes found modulated in fibroblasts, 92% of them were common to DUVR and to UVA radiation. Five percent and three percent only were specific to UVA radiation or to DUVR. In keratinocytes, the vast majority of the modulated genes were identical in DUVR or UVA exposure conditions. Twenty percent were specific to DUVR, underlying the impact of UVB and short UVA radiation included in DUVR spectrum and received by the keratinocytes (Fig. 4). This transcriptomic study confirmed that UVA radiation included in DUVR has a strong impact in fibroblasts and keratinocytes of reconstructed skin.
Figure 3. Biological changes observed in vitro in reconstructed model without or after UVA exposure (21) Oxidative stress was revealed by DCFH-DA probe, fibroblast alterations correspond to apoptotic process (TUNEL reaction),and disappearance of dermal fibroblasts was revealed by histology (HES staining).
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
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Figure 4. Percentages of modulated genes in fibroblasts and keratinocytes of reconstructed skin exposed to UVA radiation or to daily UV radiation (DUVR).
Altogether these phenomena may be involved in early events occurring during photoaging that lead to drastic alterations of dermal structure and ‘solar elastosis’. Both in vivo and in vitro studies confirm the fundamental role of UVA radiation in skin photoaging.
Skin color types in Japan and sensitivity of skin to UV light and photodamage Skin color types Skin color type essentially depends on constitutive pigmentation. Melanin is the major determinant of skin color. There is more than one type of melanin: melanin is a mixture of pigments that includes diffuse, unprotective yellow-red pheomelanins, and more or less dense, protective brown-black eumelanins. These combine in different proportions which tend to vary with ethnicity and photoexposure – dark skin having higher melanin and eumelanin content than light skin. Variation in skin pigmentation is strongly influenced by the amount of melanin, its composition and its distribution through the epidermis (33–35). Assessment of skin color according to Fitzpatrick’s phototype classification based on self reported erythema sensitivity and tanning ability has been extensively used but it has turned out to show some limitations in terms of quantitative reliability, reproducibility and relevance in some populations, especially in Asia (36,37). In order to cope with such problem another skin color classification has been proposed based on colorimetric parameters L* (luminance) and b* (yellow/blue component), Individual Typology Angle (ITA) that is calculated according to the formula °ITA = (Arc Tan (L* 50)/b*) 9 180/3.14159 (38). This allowed skin color types to be classified objectively into six groups, from very light to dark skin. A study was conducted in Japan to explore the diversity of skin color among Japanese with 464 Japanese women (39). Their skin color types were evaluated based on the determination of the ITA. Figure 5 illustrates the results obtained: Japanese skin types belong for a majority to intermediate group but also to light, and tan groups.
Skin color types and sensitivity to UV light Very few experimental data are available showing the relationship between UV-induced damage and skin type (40,41). Del Bino
10
Figure 5. Japanese skin color distribution in the Individual Typology Angle (ITA) field (n = 464).
et al. analyzed 42 ex vivo skin samples to investigate the relationship between skin color and UV-induced skin response. Skin color type was determined through the measurement of ITA. The skin samples were exposed to increasing doses of solar-simulated radiation. An immunolabeling method was used to detect the formation of typical DNA lesions, cyclobutane pyrimidine dimers (CPD). The results showed an inverse relationship between constitutive pigmentation and CPD formation. Immunolabeling of pyrimidine dimers showed a dose dependent accumulation of DNA damage throughout the whole epidermal layers including the basal layer as well as in the upper dermis of light, intermediate and tan skins. In contrast, in brown and dark skins, DNA damage was only detected in suprabasal epidermal layers (40). This confirms the results of another study which revealed a significant inverse correlation between the absolute amount and distribution of melanin and the amount of photoprotection from UV-induced DNA damage (42). Lesions in the upper dermal cells may be relevant to greater susceptibility to photoaging (43,44). Melanocytes, also localized within the basal epidermal layer may also be targeted in such light, intermediate and tan skin groups. By crossing the results with the classification of Japanese skin depending on ITA value, it revealed that Japanese skin is particularly sensitive to UV light. From light to tan skin types group there are particular high risks of DNA damages which could lead to a photocancer proneness and an early onset of photoaging.
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
UVA induced damage and skin types
How to prevent photoaging? Daily well-balanced photoprotection can prevent hyperpigmentation As detailed above, Asian skin is sensitive to UV-induced pigmentation and in particular to UVA-induced pigmentation. Some studies have shown that by broadening the absorption spectrum from UVB towards UVA radiation the sunscreen efficacy is increased against sun-induced pigmentation. Fourtanier et al. demonstrated that sunscreens with appropriate UV protection efficacy were effective for UVR induced pigmentation. Six different sunscreen products containing UVA + UVB absorbers with different SPF/UVAPF ratios were tested on volunteers’ skin exposed to solar radiation mimicking standard daily UVR (45). Healthy male or female volunteers originated from Asia with Fitzpatrick skin type III and ITA° values from 28° to 41° (Intermediate skin color) were included. The results showed that for the same level of SPF, products having the highest UVAPF provided higher protection against pigmentation, and products having a SPF/UVAPF ratio below 3 were more effective than those with a ratio above 3. The product with a well-balanced photoprotection is more efficient in the prevention of UV-induced pigmentation. The importance of daily care sunscreen in prevention of pigmentation was investigated in a recent double-blind randomized study (46). 21 Japanese women, 30–45 years with pigmented spots on the upper back were recruited. Areas on the upper back, with and without spot, were exposed to suberythemal UVA-1 doses (340–400 nm). Fifteen minutes before each exposure, a broadspectrum sunscreen (SPF50, PPD 18) or its vehicle were applied. The results showed that pigmentation score was significantly lower in the daily care treated area than in the vehicle treated one at day 42 (Fig. 6). This confirms that the use of a daily care product containing a broad-spectrum filtration offers an efficient protection of Japanese skin after chronic UVA-1 realistic exposure conditions.
Daily well-balanced photoprotection can prevent dermal alterations In vitro and in vivo studies have now demonstrated that UVA radiation contributes significantly in long-term dermal structure deterioration and clinical signs of photoaging. We have seen previously that 3-dimensional organotypic models have been developed to identify and understand the early biological events induced by UV light, but also to evaluate the efficiency of sunscreens to prevent those biological events. Bernerd et al. (47) demonstrated that the product filtering both UVB and UVA radiations afforded a better protection than the product absorbing UVB with regards to biological markers related to photo-aging. The value of SPF alone seemed not to be sufficient to predict the protective effect of the sunscreens in solar simulated exposure conditions. An in vitro study using a skin reconstructed model was also conducted to assess the protection afforded by two different sunscreens under standard daily ultraviolet radiation exposure conditions (48). The two sunscreens had the same SPF value but different profile of UVA-PF (ratio SPF/UVA-PF (PPD) 3 for sunscreen B). Dose response experiments showed that the sunscreen with the highest UVA-PF (A) provided a better protection against dermal damage. The results showed that the sunscreen having the ratio SPF/UVA-PF (PPD) 3, with regard to photoaging related biomarkers, i.e. dermal fibroblasts alteration and matrix metalloproteinase production. It thus demonstrates that for a given SPF value, efficient photoprotection required a significant UVA absorption potency. The efficacy of well-balanced photoprotection has also been evaluated on reconstructed skin model at the molecular level (30). The protection afforded by a broad spectrum sunscreen (SPF: 67.5 6.2 and UVA-PF (PPD method): 31.1 6.4) was evaluated thanks to a semi-global gene expression analysis. 244 genes in keratinocytes and 227 in fibroblasts were analyzed separately in the reconstructed skin after UVA exposure with or without prior application of the sunscreen. In both skin compartments, UVA radiation induced modulation of several genes involved in extracellular matrix, oxidative stress response, heat shock response, cell growth, inflammation and epidermal differentiation. Sunscreen pre-application abrogated these effects or reduced them significantly. The application of the sunscreen before UVA exposure kept a gene expression profile very close or similar to unexposed samples. These data indicated that a broad spectrum sunscreen was able to prevent skin cells from UVA-induced gene responses corresponding to cellular events beyond the in vivo protection factor determination.
Conclusion
Figure 6. Effect of a daily care sunscreen on the skin pigmentation induced by UVA radiation (light blue: vehicle; dark blue: daily care).
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
UVA radiation has been clearly evidenced to be responsible for more or less onset of photoageing and photocarcinogenesis in early stage. Gene expression studies recently revealed that UVA radiation induces modulation of several genes confirming the high sensitivity of dermal fibroblasts to UVA radiation. UVA radiation also appears to play a key role in pigmentary changes occurring with age, the major sign of skin photoaging in Asians. It has been shown that skin susceptibility to photoaging alterations also depends on constitutive pigmentation. The skin sensitivity to UV light has been demonstrated to be linked to skin color type. Our
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data emphasized more particularly the high sensitivity of Japanese skin to UV light. UVA radiation effects may be more insidious than UVB- induced, with sunburn alarm, and therefore they need to be taken into account very seriously. An effective photoprotection against photoaging and hyperpigmentation is only provided by a real broad spectrum sunscreen providing potent absorption
in both UVB and UVA radiations and referred to as well-balanced.
Conflict of interest The authors have no conflicts of interest that are directly relevant to the content of this article.
References
1 Commission Internationale de l’ Eclairage (International Commission on Illumination) 1989, http://www.cie.co.at/Publications/index.php?i_ca_ id=360, Publ. CIE No. 85, ISBN 978 3 900734 22 0. 2 Sabziparvar A A, Shine K P, Forster P M. Photochem Photobiol 1999: 69: 193–202. 3 Vernez D, Milon A, Vuilleumier L et al. Br J Dermatol 2012: 167: 383–390. 4 Ou-Yang H, Stamatas G, Saliou C et al. J Invest Dermatol 2004: 122: 1020–1029. 5 Seite S, Moyal D, Richard S et al. J Photochem Photobiol, B 1998: 44: 69–76. 6 Lavker R M, Gerberick G F, Veres D et al. J Am Acad Dermatol 1995: 32: 53–62. 7 Lowe N J, Meyers D P, Wieder J M et al. J Invest Dermatol 1995: 105: 739–743. 8 Mera S L, Lovell C R, Jones R R et al. Br J Dermatol 1987: 117: 21–27. 9 Albrecht S, From L, Kahn H J. J Cutan Pathol 1991: 18: 75–80. 10 Park P W, Biedermann K, Mecham L et al. J Invest Dermatol 1996: 106: 1075–1080. 11 Zhao P, Zhu X, Liu Y et al. Arch Environ Health 1998: 53: 405–409. 12 Chung J H. Photodermatol Photoimmunol Photomed 2003: 19: 109–121. 13 Goh S H. Br J Dermatol 1990: 122: 105–109. 14 Alaluf S, Atkins D, Barrett K et al. Pigment Cell Res 2002: 15: 112–118. 15 Szabo G, Gerald A B, Pathak M A et al. Nature 1969: 222: 1081–1092. 16 Toda K, Pathak M A, Parrish J A et al. Nat New Biol 1972: 236: 143–145.
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17 Konrad K, Wolff K. Arch Dermatol 1973: 107: 853–860. 18 Thong H Y, Jee S H, Sun C C et al. Br J Dermatol 2003: 149: 498–505. 19 Jimbow M, Jimbow K. Clin Dermatol 1989: 7: 11–27. 20 Kollias N, Malallah Y H, al-Ajmi H et al. Photodermatol Photoimmunol Photomed 1996: 12: 183–188. 21 Bernerd F, Asselineau D. Cell Death Differ 1998: 5: 792–802. 22 Bernerd F, Asselineau D. Dev Biol 1997: 183: 123–138. 23 Fagot D, Asselineau D, Bernerd F. Photochem Photobiol 2004: 79: 499–505. 24 Fagot D, Asselineau D, Bernerd F. Arch Dermatol Res 2002: 293: 576–583. 25 Lahmann C, Young A R, Wittern K P et al. Photochem Photobiol 2001: 73: 657–663. 26 Fisher G J, Datta S C, Talwar H S et al. Nature 1996: 379: 335–339. 27 Chung J H, Seo J Y, Choi H R et al. J Invest Dermatol 2001: 17: 1218–1224. 28 Scharffetter K, Wlaschek M, Hogg A et al. Arch Dermatol Res 1991: 283: 506–511. 29 Wlaschek M, Bolsen K, Herrmann G et al. J Invest Dermatol 1993: 101: 164–168. 30 Marionnet C, Grether-Beck S, Seite S et al. Exp Dermatol 2011: 20: 477–482. 31 Marionnet C, Lejeune F, Pierrard C et al. J Dermatol Sci 2012: 66: 238–240. 32 Christiaens F J, Chardon A, Fourtanier A et al. Photochem Photobiol 2005: 81: 874–878. 33 Alaluf S, Atkins D, Barrett K et al. Pigment Cell Res 2002: 15: 112–118.
34 Wakamatsu K, Kavanagh R, Kadekaro A L et al. Pigment Cell Res 2006: 19: 154–162. 35 Nielsen K P, Zhao L, Stamnes J J et al. J Photochem Photobiol, B 2006: 82: 194–198. 36 Taylor S C. J Am Acad Dermatol 2002: 46: S41– S62. 37 Rawlings A V. Int J Cosmet Sci 2006: 28: 79–93. 38 Chardon A, Cretois I, Hourseau C. Int J Cosmet Sci 1991: 13: 191–208. 39 Del Bino S, Bernerd F. Br J Dermatol 2013: 169 (Suppl S3): 38–45. 40 Del Bino S, Sok J, Bessac E et al. Pigment Cell Res 2006: 19: 606–614. 41 Ueda M, Matsunaga T, Bito T et al. Photodermatol Photoimmunol Photomed 1996: 12: 22–26. 42 Tadokoro T, Kobayashi N, Zmudzka B Z et al. FASEB J 2003: 17: 1177–1179. 43 Fisher G J, Kang S, Varani J et al. Arch Dermatol 2002: 138: 1462–1470. 44 Rijken F, Bruijnzeel P L, Van Weelden H et al. J Invest Dermatol 2004: 122: 1448–1455. 45 Fourtanier A, Moyal D, Seite S. Photochem Photobiol Sci 2012: 11: 81–89. 46 Morita A, Rougier A, Seite S. Poster World Congress of Dermatology 2011, Seoul, Korea 47 Bernerd F, Vioux C, Lejeune F et al. Eur J Dermatol 2003: 13: 242–249. 48 Lejeune F, Christiaens F, Bernerd F. Photodermatol Photoimmunol Photomed 2008: 24: 249–255.
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
Review
www.wileyonlinelibrary.com/journal/EXD
New insights in photoaging, UVA induced damage and skin types Claire Battie, Setsuko Jitsukawa, Francßoise Bernerd, Sandra Del Bino, Claire Marionnet and le Verschoore Miche L’Oreal Research and Innovation, Asnieres, France Correspondence: Claire Battie, L’Oreal Research and Innovation, Asnieres, France, Tel.: +33 1 47 56 71 98, e-mail: [email protected] Abstract: UVA radiation is the most prevalent component of solar UV radiation; it deeply penetrates into the skin and induces profound alterations of the dermal connective tissue. In recent years, the detrimental effects of UVA radiation were more precisely demonstrated at cellular and molecular levels, using adequate methods to identify biological targets of UVA radiation and the resulting cascade impairment of cell functions and tissue degradation. In particular gene expression studies recently revealed that UVA radiation induces modulation of several genes confirming the high sensitivity of dermal fibroblasts to UVA radiation. The major visible damaging effects of UVA radiation only appear after years of exposure: it has been clearly evidenced
that they are responsible for more or less early signs of photoageing and photocarcinogenesis. UVA radiation appears to play a key role in pigmented changes occurring with age, the major sign of skin photoaging in Asians. Skin susceptibility to photoaging alterations also depends on constitutive pigmentation. The skin sensitivity to UV light has been demonstrated to be linked to skin color type.
Introduction
around midday. Another interesting characteristic of UVA radiation is that it comes through glass, whereas UVB rays are almost entirely absorbed. Thus, high UVA doses may be received even in indoor conditions while erythemal UVB radiation is filtered out. Seventy percent of UVB radiation that reaches the skin is absorbed by the stratum corneum, 20% reaches viable epidermis, and only 10% penetrates the uppermost part of the dermis. On the other hand, UVA radiation is partly absorbed by the epidermis, but 20–30% of it reaches deep dermis. UVB radiation consequently has a major action on the epidermis compared to UVA radiation which can also target the dermis. That’s one of the reasons why UVA radiation is now considered to be a major factor in the process of skin photoaging but also participates in photocarcinogenesis. The contribution of diffuse UV radiation is also important and should not be underestimated. Indeed a recent study suggests that diffuse irradiation may explain a large part of the cumulative annual exposure dose (3). The consequences of UVA exposure on skin are particularly important in Asia, since UVA exposure appear to be more important in principal cities in Asia compared to Europe because of the low latitude. The calculation of UVA fluence received in Asia relative to that received in Paris on a winter day showed that in Asia, the exposure to UVA light is much more than in Paris (Fig. 1) (2).
Solar UV radiation reaching the earth is a combination of UVB (290–320 nm) and UVA (320–400 nm) wavelengths. Since UVA radiation is less energetic than UVB radiation, UVB radiation has long been thought to be the factor responsible for the damaging effects of solar radiation. But it is now proved that UVA radiation plays a major role. In fact UVA radiation is the most prevalent component of solar UV radiation, it penetrates deeper than UVB radiation into the skin and induces profound alterations of the dermal connective tissue. In recent years, the detrimental effects of UVA radiation were more precisely demonstrated at cellular and molecular levels, using adequate methods to identify biological targets of UVA radiation and the resulting cascade impairment of cell functions and tissue degradation.
Skin photoaging: UVA radiation plays a major role Cutaneous UV exposure UVB radiation reaches the earth in relatively low amounts (about 0.5% of solar spectral irradiance at ground level, integrated over 290–2500 nm range) and is highly energetic. In contrast, UVA rays are lower in energy, but they are at least 20 times more abundant. Ninety five percent of UV rays reaching the ground level are UVA (1). Solar UV irradiance depends on many geo-orbital and environmental parameters including latitude, time of the year (season), hour of the day, meteorological conditions and the thickness of ozone layer. UVA radiation is less affected by those parameters and varied to a lesser extent than UVB radiation. For example UVA irradiance is less affected by seasons, and decreases to a lesser extent in winter (2). The time of the day plays an important role too. Indeed UVA and UVB radiations both raise from the beginning of the day, peak at noon and decrease at the end of the day but UVA radiation is present for most part of the day as it follows the variation of visible light whereas UVB rays are at the highest between 10 a.m. and 4 p.m., especially
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
Key words: photoageing – photoprotection – skin sensitivity to UV – UVA
Accepted for publication 24 March 2014
UVA radiation induces biological alterations related to photoaging – in vivo proofs In vivo studies have now given clear evidence that UVA radiation significantly contributes to long-term dermal structure deterioration and clinical signs of photoaging. It is responsible for the early appearance of signs of photoageing, which include patchy/mottled pigmentation, wrinkling, laxity, sagging, dryness, etc. One of the major modes of action of UVA radiation is the generation of –
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(a)
(b)
Figure 1. UVA dose received in Asian cities compared to that in Paris: Harbin, Beijing, Seoul and Tokyo respectively receive 1.29, 1.84, 1.93 and 1.97 times more UVA doses than Paris.
oxidative stress which is a well known factor in the pathogenesis of photoaging (4). The biological effects of repetitive, suberythemal UVA exposure were explored in vivo (5–7). For example, in a reported clinical study, 14 volunteers were exposed three times per week for 13 weeks to increasing doses of UVA radiation (5). Different biological markers related to skin photoaging were studied: ferritin and tenascin expression as well as lysozyme quantification. The results showed that UVA exposure induced an increased expression of ferritin in the basal and suprabasal keratinocyte layers and in interstitial dermal cells (Fig. 2a). As ferritin is an important marker of antioxidant activity, its increase proved that UVA radiation induced oxidative stress. UVA radiation also induced a marked increase in tenascin expression, just below the dermo-epidermal junction (Fig. 2b). Tenascin is a large glycoprotein of the extracellular matrix, interacting with collagens, proteoglycans and fibronectin. It is expressed at low levels in normal skin while it is upregulated in skin tumours and in a number of skin diseases, which reveals the damaging effects of UVA radiation. UVA radiation induced a significant (P < 0.05) increase in deposit of lysozyme (Fig. 2c). Lysozyme has been shown to be associated with damaged elastic fibers in many tissues (8,9). In skin, lysozyme deposits on elastic fibers are seen in sun-damaged regions, and the number of lysozyme-containing elastin fibers appears to be correlated with the degree of sun damage. It has been suggested that lysozyme binds specifically to damaged elastin fibers and thereby inhibits the proteolytic degradation of altered fibers by elastases (10). This may be linked to solar elastosis. The same study also investigated the effects of repetitive, suberythemal UVA exposure on skin pigmentation. It was shown that this regimen of exposure induced a strong increase in pigmentation. Pigmented changes are particularly important in Asia since it is the major sign of skin photoaging in Asians (11–13). An ethnic group-related variation in melanosome distribution was reported showing a mix of individual (about 60%) and aggregated (about
8
(c)
Figure 2. UVA exposure induces alterations of epidermal and dermal protein expression. Immunofluorescence microscopy using antisera against ferritin (a), tenascin (b) and lysosyme (c).
40%) melanosomes in Asian skins, whereas aggregated melanosomes (85%) are prevailing in European skins (14–19). The density and highly variable size of melanosomes in Asian skins could account for irregular, spotty pigmentation associated with photoageing. It has been proved that in darker-skinned individuals, UVA radiation induces greater pigmenting effects than UVB (20).
UVA radiation induces biological alterations related to photoaging – in vitro proofs In vitro organotypic skin models have been developed providing a three-dimensional tissue structure and a complete epidermal differentiation like in vivo. Such models allow to study the early biological events occurring after a single or repeated UV exposure and to investigate the biological effects of UVA and UVB radiations. Unlike UVB radiation, UVA radiation, due to its high penetration properties, can reach deeper parts of the skin and affects the dermal compartment. In the reconstructed skin model, UVA radiation actually induces major alterations in the dermal compartment through the generation of ROS. As a result of exposure to
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
UVA induced damage and skin types
UVA radiation (25 J/cm²), the dermal fibroblasts located in the upper part of the dermal equivalent disappear within 48 h following exposure through an apoptotic process (Fig. 3) (21). On the other hand, the epidermal structure and organization are not significantly affected, indicating that the survival ability of dermal fibroblasts after exposure to pure UVA radiation is lower compared to that of epidermal keratinocytes. These results confirm previous experiments showing that dermal fibroblasts are more sensitive to UVA-induced oxidative stress than keratinocytes (21,22). The increase in MMP-1 after UV exposure was also observed in reconstructed skin model (23,24). MMP-1 is an interstitial collagenase able to hydrolyze type I collagen, the major component of the dermis, and it seems to play a crucial role in the disorganization and progressive degeneration of dermal extracellular matrix (25–27). Under UVA exposure, MMP-1 production was directly induced in the dermal fibroblasts. Removal of epidermis immediately after UVA exposure did not alter this induction. These results confirmed other data on UVA-induced MMP-1 in cultured fibroblasts (28,29). In contrast, UVB-induced MMP-1 production required the presence of the epidermis. More recently the impact of UVA radiation was studied at the molecular level by evaluating UVA radiation-induced gene expression in reconstructed skin in vitro and in human skin biopsies in vivo. The results showed that the expression of genes related to
oxidative stress response and relevant for photoageing were affected by UVA exposure (30). For example, in vitro and in vivo, UVA exposure induced an increase of MMP-1, haeme oxygenase-1, and superoxide dismutase-2 gene expression. The use of the reconstructed skin model allowed the authors to precise the contribution of fibroblast and keratinocytes in gene expression modulation. Another transcriptomic study on reconstructed skin model (31) focused more particularly on the impact of UVA radiation during a non zenithal UV exposure, occurring in most outdoors activities, called daily UV radiation (DUVR) (32). To characterize the contribution of UVA wavelengths in the biological impact of DUVR, a reconstructed skin model was exposed either to 13 J/cm2 DUVR or to 25 J/cm2 UVA radiation. Gene modulation of several genes related to functional families was quantified (oxidative stress, extracellular matrix, transcription, signaling, heat shock proteins, cytokines). Out of the 60 genes found modulated in fibroblasts, 92% of them were common to DUVR and to UVA radiation. Five percent and three percent only were specific to UVA radiation or to DUVR. In keratinocytes, the vast majority of the modulated genes were identical in DUVR or UVA exposure conditions. Twenty percent were specific to DUVR, underlying the impact of UVB and short UVA radiation included in DUVR spectrum and received by the keratinocytes (Fig. 4). This transcriptomic study confirmed that UVA radiation included in DUVR has a strong impact in fibroblasts and keratinocytes of reconstructed skin.
Figure 3. Biological changes observed in vitro in reconstructed model without or after UVA exposure (21) Oxidative stress was revealed by DCFH-DA probe, fibroblast alterations correspond to apoptotic process (TUNEL reaction),and disappearance of dermal fibroblasts was revealed by histology (HES staining).
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
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Figure 4. Percentages of modulated genes in fibroblasts and keratinocytes of reconstructed skin exposed to UVA radiation or to daily UV radiation (DUVR).
Altogether these phenomena may be involved in early events occurring during photoaging that lead to drastic alterations of dermal structure and ‘solar elastosis’. Both in vivo and in vitro studies confirm the fundamental role of UVA radiation in skin photoaging.
Skin color types in Japan and sensitivity of skin to UV light and photodamage Skin color types Skin color type essentially depends on constitutive pigmentation. Melanin is the major determinant of skin color. There is more than one type of melanin: melanin is a mixture of pigments that includes diffuse, unprotective yellow-red pheomelanins, and more or less dense, protective brown-black eumelanins. These combine in different proportions which tend to vary with ethnicity and photoexposure – dark skin having higher melanin and eumelanin content than light skin. Variation in skin pigmentation is strongly influenced by the amount of melanin, its composition and its distribution through the epidermis (33–35). Assessment of skin color according to Fitzpatrick’s phototype classification based on self reported erythema sensitivity and tanning ability has been extensively used but it has turned out to show some limitations in terms of quantitative reliability, reproducibility and relevance in some populations, especially in Asia (36,37). In order to cope with such problem another skin color classification has been proposed based on colorimetric parameters L* (luminance) and b* (yellow/blue component), Individual Typology Angle (ITA) that is calculated according to the formula °ITA = (Arc Tan (L* 50)/b*) 9 180/3.14159 (38). This allowed skin color types to be classified objectively into six groups, from very light to dark skin. A study was conducted in Japan to explore the diversity of skin color among Japanese with 464 Japanese women (39). Their skin color types were evaluated based on the determination of the ITA. Figure 5 illustrates the results obtained: Japanese skin types belong for a majority to intermediate group but also to light, and tan groups.
Skin color types and sensitivity to UV light Very few experimental data are available showing the relationship between UV-induced damage and skin type (40,41). Del Bino
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Figure 5. Japanese skin color distribution in the Individual Typology Angle (ITA) field (n = 464).
et al. analyzed 42 ex vivo skin samples to investigate the relationship between skin color and UV-induced skin response. Skin color type was determined through the measurement of ITA. The skin samples were exposed to increasing doses of solar-simulated radiation. An immunolabeling method was used to detect the formation of typical DNA lesions, cyclobutane pyrimidine dimers (CPD). The results showed an inverse relationship between constitutive pigmentation and CPD formation. Immunolabeling of pyrimidine dimers showed a dose dependent accumulation of DNA damage throughout the whole epidermal layers including the basal layer as well as in the upper dermis of light, intermediate and tan skins. In contrast, in brown and dark skins, DNA damage was only detected in suprabasal epidermal layers (40). This confirms the results of another study which revealed a significant inverse correlation between the absolute amount and distribution of melanin and the amount of photoprotection from UV-induced DNA damage (42). Lesions in the upper dermal cells may be relevant to greater susceptibility to photoaging (43,44). Melanocytes, also localized within the basal epidermal layer may also be targeted in such light, intermediate and tan skin groups. By crossing the results with the classification of Japanese skin depending on ITA value, it revealed that Japanese skin is particularly sensitive to UV light. From light to tan skin types group there are particular high risks of DNA damages which could lead to a photocancer proneness and an early onset of photoaging.
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
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How to prevent photoaging? Daily well-balanced photoprotection can prevent hyperpigmentation As detailed above, Asian skin is sensitive to UV-induced pigmentation and in particular to UVA-induced pigmentation. Some studies have shown that by broadening the absorption spectrum from UVB towards UVA radiation the sunscreen efficacy is increased against sun-induced pigmentation. Fourtanier et al. demonstrated that sunscreens with appropriate UV protection efficacy were effective for UVR induced pigmentation. Six different sunscreen products containing UVA + UVB absorbers with different SPF/UVAPF ratios were tested on volunteers’ skin exposed to solar radiation mimicking standard daily UVR (45). Healthy male or female volunteers originated from Asia with Fitzpatrick skin type III and ITA° values from 28° to 41° (Intermediate skin color) were included. The results showed that for the same level of SPF, products having the highest UVAPF provided higher protection against pigmentation, and products having a SPF/UVAPF ratio below 3 were more effective than those with a ratio above 3. The product with a well-balanced photoprotection is more efficient in the prevention of UV-induced pigmentation. The importance of daily care sunscreen in prevention of pigmentation was investigated in a recent double-blind randomized study (46). 21 Japanese women, 30–45 years with pigmented spots on the upper back were recruited. Areas on the upper back, with and without spot, were exposed to suberythemal UVA-1 doses (340–400 nm). Fifteen minutes before each exposure, a broadspectrum sunscreen (SPF50, PPD 18) or its vehicle were applied. The results showed that pigmentation score was significantly lower in the daily care treated area than in the vehicle treated one at day 42 (Fig. 6). This confirms that the use of a daily care product containing a broad-spectrum filtration offers an efficient protection of Japanese skin after chronic UVA-1 realistic exposure conditions.
Daily well-balanced photoprotection can prevent dermal alterations In vitro and in vivo studies have now demonstrated that UVA radiation contributes significantly in long-term dermal structure deterioration and clinical signs of photoaging. We have seen previously that 3-dimensional organotypic models have been developed to identify and understand the early biological events induced by UV light, but also to evaluate the efficiency of sunscreens to prevent those biological events. Bernerd et al. (47) demonstrated that the product filtering both UVB and UVA radiations afforded a better protection than the product absorbing UVB with regards to biological markers related to photo-aging. The value of SPF alone seemed not to be sufficient to predict the protective effect of the sunscreens in solar simulated exposure conditions. An in vitro study using a skin reconstructed model was also conducted to assess the protection afforded by two different sunscreens under standard daily ultraviolet radiation exposure conditions (48). The two sunscreens had the same SPF value but different profile of UVA-PF (ratio SPF/UVA-PF (PPD) 3 for sunscreen B). Dose response experiments showed that the sunscreen with the highest UVA-PF (A) provided a better protection against dermal damage. The results showed that the sunscreen having the ratio SPF/UVA-PF (PPD) 3, with regard to photoaging related biomarkers, i.e. dermal fibroblasts alteration and matrix metalloproteinase production. It thus demonstrates that for a given SPF value, efficient photoprotection required a significant UVA absorption potency. The efficacy of well-balanced photoprotection has also been evaluated on reconstructed skin model at the molecular level (30). The protection afforded by a broad spectrum sunscreen (SPF: 67.5 6.2 and UVA-PF (PPD method): 31.1 6.4) was evaluated thanks to a semi-global gene expression analysis. 244 genes in keratinocytes and 227 in fibroblasts were analyzed separately in the reconstructed skin after UVA exposure with or without prior application of the sunscreen. In both skin compartments, UVA radiation induced modulation of several genes involved in extracellular matrix, oxidative stress response, heat shock response, cell growth, inflammation and epidermal differentiation. Sunscreen pre-application abrogated these effects or reduced them significantly. The application of the sunscreen before UVA exposure kept a gene expression profile very close or similar to unexposed samples. These data indicated that a broad spectrum sunscreen was able to prevent skin cells from UVA-induced gene responses corresponding to cellular events beyond the in vivo protection factor determination.
Conclusion
Figure 6. Effect of a daily care sunscreen on the skin pigmentation induced by UVA radiation (light blue: vehicle; dark blue: daily care).
© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd Experimental Dermatology, 2014, 23 (Suppl. 1), 7–12
UVA radiation has been clearly evidenced to be responsible for more or less onset of photoageing and photocarcinogenesis in early stage. Gene expression studies recently revealed that UVA radiation induces modulation of several genes confirming the high sensitivity of dermal fibroblasts to UVA radiation. UVA radiation also appears to play a key role in pigmentary changes occurring with age, the major sign of skin photoaging in Asians. It has been shown that skin susceptibility to photoaging alterations also depends on constitutive pigmentation. The skin sensitivity to UV light has been demonstrated to be linked to skin color type. Our
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data emphasized more particularly the high sensitivity of Japanese skin to UV light. UVA radiation effects may be more insidious than UVB- induced, with sunburn alarm, and therefore they need to be taken into account very seriously. An effective photoprotection against photoaging and hyperpigmentation is only provided by a real broad spectrum sunscreen providing potent absorption
in both UVB and UVA radiations and referred to as well-balanced.
Conflict of interest The authors have no conflicts of interest that are directly relevant to the content of this article.
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