during the acquisition of a fully terres- trialized life style (Strasser et al., 2014), our results have uncovered an important property of LOR, photoprotective func- tion, that is indispensable for adapting to terrestrial conditions. All procedures were under the pro- tocols approved by the University of Colorado Animal Care and Use Program. CONFLICT OF INTEREST The authors state no conflict of interest. ACKNOWLEDGMENTS This research was supported by a grant from NIAMS (AR47898) to DRR. The authors thank the University of Colorado Skin Disease Research Center (P30AR057212) Morphology Phenotyping Core for the assistance and the Biophysics Core Facility at the University of Colorado for the technical assistance with the HPLC analysis. Yosuke Ishitsuka 1,* and Dennis R. Roop 1 1 Department of Dermatology and Charles C. Gates Center for Regenerative Medicine, Anschutz Medical Campus, University of Colorado, Aurora, Colorado, USA * Corresponding author e-mail: yosuke. ishitsuka@md.tsukuba.ac.jp SUPPLEMENTARY MATERIAL Supplementary material is linked to the online version of the paper at www.jidonline.org, and at https://10.1016/j.jid.2018.06.164. REFERENCES Barresi C, Stremnitzer C, Mlitz V, Kezic S, Kammeyer A, Ghannadan M, et al. Increased sensitivity of histidinemic mice to UVB radia- tion suggests a crucial role of endogenous urocanic acid in photoprotection. J Invest Der- matol 2011;131:188e94. Denecker G, Hoste E, Gilbert B, Hochepied T, Ovaere P, Lippens S, et al. Caspase-14 pro- tects against epidermal UVB photodamage and water loss. Nat Cell Biol 2007;9:666e74. Elias PM. Stratum corneum defensive functions: an integrated view. J Invest Dermatol 2005;125: 183e200. Hoste E, Kemperman P, Devos M, Denecker G, Kezic S, Yau N, et al. 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Biosynthetic pathways of filaggrin and lor- icrin—two major proteins expressed by termi- nally differentiated epidermal keratinocytes. J Struct Biol 1990;104:150e62. Strasser B, Mlitz V, Hermann M, Rice RH, Eigenheer RA, Alibardi L, et al. Evolutionary origin and diversification of epidermal barrier proteins in amniotes. Mol Biol Evol 2014;31: 3194e205. Yuspa SH, Kilkenny AE, Steinert PM, Roop DR. Expression of murine epidermal differentiation markers is tightly regulated by restricted extra- cellular calcium concentrations in vitro. J Cell Biol 1989;109:1207e17. Laser Irradiation of Organic Tattoo Pigments Releases Carcinogens with 3,3 0 -Dichlorobenzidine Inducing DNA Strand Breaks in Human Skin Cells Journal of Investigative Dermatology (2018) 138, 2687e2690; doi:10.1016/j.jid.2018.05.031 TO THE EDITOR Laser irradiation is the most common way to remove unwanted tattoos. Nowadays, primarily organic pigments are used for brilliant color shades (Laux et al., 2016). Laser irradiation of organic pigments was shown to produce carcinogenic compounds after laser irradiation (Hauri and Hohl, 2015; Schreiver et al., 2015; Vasold et al., 2004). Because of the detection of car- cinogens, a potential connection between tattoos and skin cancer is being controversially discussed (Kluger and Koljonen, 2012). Necessary long- term cancer studies are currently unfeasible because animal testing for tattoo-related health issues was rated unethical in Germany due to its voluntary application. Alternatively, in vitro methods such as the staining of phosphorylated histone H2AX are widely used to investigate genotoxicity in terms of DNA double-strand breaks and concurrent genome instability, both of which are crucial events during cancer development. Here we quantified decomposition products of the diazo pigment orange Abbreviations: DCBD, 3,3 0 -dichlorobenzidine; HCB, hexachlorobenzene; P.O., pigment orange; P.Y., pigment yellow Accepted manuscript published online 20 June 2018; corrected proof published online 20 August 2018 ª 2018 The Authors. Published by Elsevier, Inc. on behalf of the Society for Investigative Dermatology. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by- nc-nd/4.0/). H Hering et al. Carcinogens Formed by Tattoo Laser Irradiation www.jidonline.org 2687