Anneliese Striz1, Ana DePina1, Robert Jones1, Xiugong Gao1, Jeffrey Yourick1. 1. Division of Toxicology, Office of Applied Research and Safety Assessment, Center for Food Safety and Applied Nutrition, U.S. Food and Drug Administration, Laurel, MD, USA.
Abstract
PURPOSE: Dihydroxyacetone (DHA) is the only ingredient approved by the U.S. FDA as a colour additive in sunless tanning (self-tanning) products. Consumer sunless tanning products available for retail purchase contain 1-15% DHA. Although originally thought to only interact with the stratum corneum, more recent research has shown that DHA penetrates beyond the stratum corneum to living keratinocytes indicating a possible route of exposure in the epidermis. MATERIALS AND METHODS: Normal Human Epidermal Keratinocytes (NHEK) were used to determine any potential in vitro toxicological effects of DHA in the epidermis. NHEK cells exposed to DHA concentrations up to 0.90% (100 mM) in dosing media were evaluated for viability, genotoxicity (Comet Assay), and gene expression changes by microarray analysis. RESULTS: Cell viability significantly decreased ∼50% after 3-h exposure to 50 and 100 mM DHA. DNA damage was only found to be significantly increased in cells exposed to cytotoxic DHA concentrations. A subtoxic dose of DHA induced significant gene expression changes. Particularly, expression of cyclin B1, CDK1, and six other genes associated with the G2/M cell cycle checkpoint was significantly decreased which correlates well with a G2/M block reported in the existing literature. Advanced Glycation End Product (AGE) formation significantly increased after 24 h of DHA exposure at and above 10 mM. In summary, these data show that DHA is cytotoxic above 25 mM in primary keratinocytes. Genotoxicity was detected only at cytotoxic concentrations, likely indicative of non-biologically relevant DNA damage, while subtoxic doses induce gene expression changes and glycation. CONCLUSION: DHA treatment had a significant and negative effect on primary keratinocytes consistent with in vitro cultured cell outcomes; however, more information is needed to draw conclusions about the biological effect of DHA in human skin.
PURPOSE: Dihydroxyacetone (DHA) is the only ingredient approved by the U.S. FDA as a colour additive in sunless tanning (self-tanning) products. Consumer sunless tanning products available for retail purchase contain 1-15% DHA. Although originally thought to only interact with the stratum corneum, more recent research has shown that DHA penetrates beyond the stratum corneum to living keratinocytes indicating a possible route of exposure in the epidermis. MATERIALS AND METHODS: Normal Human Epidermal Keratinocytes (NHEK) were used to determine any potential in vitro toxicological effects of DHA in the epidermis. NHEK cells exposed to DHA concentrations up to 0.90% (100 mM) in dosing media were evaluated for viability, genotoxicity (Comet Assay), and gene expression changes by microarray analysis. RESULTS: Cell viability significantly decreased ∼50% after 3-h exposure to 50 and 100 mM DHA. DNA damage was only found to be significantly increased in cells exposed to cytotoxic DHA concentrations. A subtoxic dose of DHA induced significant gene expression changes. Particularly, expression of cyclin B1, CDK1, and six other genes associated with the G2/M cell cycle checkpoint was significantly decreased which correlates well with a G2/M block reported in the existing literature. Advanced Glycation End Product (AGE) formation significantly increased after 24 h of DHA exposure at and above 10 mM. In summary, these data show that DHA is cytotoxic above 25 mM in primary keratinocytes. Genotoxicity was detected only at cytotoxic concentrations, likely indicative of non-biologically relevant DNA damage, while subtoxic doses induce gene expression changes and glycation. CONCLUSION: DHA treatment had a significant and negative effect on primary keratinocytes consistent with in vitro cultured cell outcomes; however, more information is needed to draw conclusions about the biological effect of DHA in human skin.