Genotoxicity of micro- and nano-particles of kaolin in human primary dermal keratinocytes and fibroblasts

INTRODUCTION: Kaolin is a clay mineral with the chemical composition Al(2)Si(2)O(5)(OH)(4). It is an important industrial material, and is also used as a white cosmetic pigment. We previously reported that fine particles of kaolin have genotoxic potency to Chinese hamster ovary CHO AA8 cells, and to the lungs of C57BL/6 J and ICR mice. In the present study, we evaluated the genotoxicity of different particle sizes of kaolin using primary normal human diploid epidermal keratinocytes and primary normal human diploid dermal fibroblasts, in addition to a CHO AA8 cell line. FINDINGS: After 6-h treatment with kaolin micro- and nano-particles of particle sizes 4.8 μm and 0.2 μm (200 nm), respectively, the frequencies of micronucleated cells increased in a dose-dependent manner. The frequency increased 3- to 4-fold by exposure to the particles at 200 μg/mL (i.e., 31.4 μg/cm(2)) in all cells tested. Two-way ANOVA revealed a significant main effect of particle size, and the nano-particles tended to have a higher potency of micronucleus (MN) induction. However, the cell type did not significantly affect the MN frequencies. In addition, one-hour treatment with the kaolin particles increased DNA damage in a dose-dependent manner in a comet assay. The %tail DNA was increased 8- to 20-fold by exposure to the particles at 200 μg/mL, for all cells tested. The kaolin nano-particles had higher DNA-damaging potency than the micro-particles. Furthermore, treatment with kaolin particles dose-dependently increased the production of reactive oxygen species (ROS) in all cells. Again, we observed that kaolin nano-particles induced more ROS than the micro-particles in all cells. CONCLUSION: Kaolin particles demonstrated genotoxicity in primary normal human diploid epidermal keratinocytes and fibroblasts as well as in CHO AA8 cells. Although no significant difference was observed among these three types of cells, fine particles of kaolin tended to have higher genotoxic potency than coarse particles. Since studies on its genotoxicity to skin have been scarce, the findings of the present study could contribute to safety evaluations of kaolin particles when used as a white cosmetic pigment.