Protective Effect of Hyperpigmented Skin on UV-Mediated Cutaneous Cancer Development Masashi Kato, Nobutaka Ohgami, Yoshiyuki Kawamoto, Toyonori Tsuzuki,

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Protective Effect of Hyperpigmented Skin on UV-Mediated Cutaneous Cancer Development Masashi Kato, Nobutaka Ohgami, Yoshiyuki Kawamoto, Toyonori Tsuzuki, Khaled Hossain, Takeshi Yanagishita, Yuichiro Ohshima, Hideo Tsuboi, Osamu Yamanoshita, Yoshinari Matsumoto, Masahide Takahashi, Izumi Nakashima Journal of Investigative Dermatology Volume 127, Issue 5, Pages 1244-1249 (May 2007) DOI: 10.1038/sj.jid.5700659 Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Macroscopic and microscopic characteristics of HL/RET-transgenic mice of line 242-hr/hr. (a) Macroscopic appearances of a 3-month-old HL/RET-transgenic mouse of line 242-hr/hr with hyperpigmented skin (bottom) and a littermate control hairless mouse (top) are presented. (a) Arrows indicate the areas without hyperpigmentation in the HL/RET-transgenic mouse. (b) Normally pigmented skin in the control mouse and (c) hyperpigmented skin in the HL/RET-transgenic mouse were stained with hematoxylin and eosin (b, c). (b, c) Bar=20μm. Journal of Investigative Dermatology 2007 127, 1244-1249DOI: (10.1038/sj.jid.5700659) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 UV irradiation-induced squamous cell carcinoma in HL/RET-transgenic mice of line 242-hr/hr (n=8). HL/RET-transgenic mice of line 242-hr/hr (n=8) and control non-transgenic hairless mice (n=5) were irradiated repeatedly with a total of 4,200kJ/m2 (30kJ/m2/day irradiation 5–6 times a week for 9 months) of UV light. Microscopic observations of squamous cell carcinoma that developed in the (b) mice of line 242-hr/hr and (a) control hairless mice are shown. (a, b) Bar=100μm. (c) Periods up to development of the skin squamous cell carcinoma (mean±SD) after starting UV irradiation in the hairless transgenic and control mice are also presented. The mean time of carcinoma development in the HL/RET-transgenic mice (n=6) was shown by the Mann–Whitney U-test to be significantly (P<0.01) delayed compared with that in the control hairless mice (n=5). HL-RET: RET-transgenic hairless mice of line 242-hr/hr; HL: littermate non-transgenic control hairless mice. Journal of Investigative Dermatology 2007 127, 1244-1249DOI: (10.1038/sj.jid.5700659) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Skin was protected by hyperpigmentation against single UV irradiation-mediated induction of skin damage in HL/RET-transgenic mice of line 242-hr/hr. (b, d, f, h, and j) Microscopic appearance of the skin from 12-week-old transgenic mice of line 242-hr/hr and (a, c, e, g, and i) littermate control non-transgenic hairless mice (a, b) before or (c, d) on day 1, (e, f) day 3, (g, h) day 5, and (i, j) day 10 after a single high dose (180kJ/m2) of UV irradiation. (h, j) Arrowheads indicate increased melanin production in the epidermis of mice after UV irradiation. (a–j) Bar=80μm. Journal of Investigative Dermatology 2007 127, 1244-1249DOI: (10.1038/sj.jid.5700659) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Single UV irradiation-mediated high-grade signal transduction in the skin was prevented by hyperpigmented skin in HL/RET-transgenic mice of line 242-hr/hr. Protein tyrosine phosphorylation (a, b), phospho-ERK, phospho-p38, and phospho-JNK levels and protein expression levels of ERK, p38, and JNK (c, d) of the skin from (b, d) 6-week-old HL/RET-transgenic mice of line 242-hr/hr and (a, c) littermate control wild-type hairless mice. Skins without UVB irradiation (UV(−) in panels c and d) and UV-irradiated skin at 7 days (UV(+) in panels c and d) or (a, b) at indicated days after a single high dose (180kJ/m2) of UVB irradiation are shown. Journal of Investigative Dermatology 2007 127, 1244-1249DOI: (10.1038/sj.jid.5700659) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions