pyrimidinones and Dermatitis--Contact

Although extracellular adenosine 5'-triphosphate (eATP) has a crucial role in the sensitization phase of contact hypersensitivity (CHS), the mechanism by which hapten causes keratinocyte cell death and ATP release is unknown. We examined the time course of cell death, reactive oxygen species (ROS) production, and ATP release in HaCaT cells and in normal human keratinocytes after exposure to nonmetal haptens, NiCl2, or irritants. Both haptens and irritants caused cell death of keratinocytes but with different time courses. N-acetylcysteine (NAC) significantly reduced only nonmetal hapten-induced cell death as assessed by propidium iodide exclusion. We examined the effects of antioxidants and pannexin (Panx) inhibitors on cell death, ROS production, and ATP release by chemical-treated HaCaT cells. Nonmetal hapten-induced cell death, but not NiCl2- or irritant-related cell death, was dependent on reactivity to thiol residues in the cells. NAC reduced cell death and ATP release, whereas antioxidants and Panx inhibitors did not inhibit cell death but significantly attenuated ATP release. Panx1 small interfering RNA (siRNA) also suppressed ATP release from hapten-exposed HaCaT cells. Intraperitoneal injection of a Panx1 inhibitor attenuated murine CHS. These findings suggest that nonmetal hapten reactivity to thiol residues causes membrane disruption of keratinocytes and ROS production that leads to ATP release through opening of Panx hemichannels.

Topics: Adenosine Triphosphate; Animals; Antioxidants; Benzopyrans; Cell Death; Cells, Cultured; Connexins; Cystine; Dermatitis, Contact; Dinitrochlorobenzene; Disease Models, Animal; Female; Haptens; Humans; Irritants; Keratinocytes; Mice; Mice, Inbred C57BL; Nerve Tissue Proteins; Nickel; Protein Structure, Quaternary; Protein Structure, Tertiary; Pyrimidinones; Reactive Oxygen Species; RNA, Small Interfering

We studied the involvement of sensory neurons in skin sensitization to allergens using a mouse model in which the T-helper type 2 response is essential. Skin sensitization to fluorescein isothiocyanate (FITC) has been shown to be enhanced by several phthalate esters, including dibutyl phthalate (DBP). For different types of phthalate esters, we found a correlation between the ability of transient receptor potential (TRP) A1 activation and that of enhancing skin sensitization. A TRPA1-specific antagonist, HC-030031, was shown to suppress skin sensitization in the presence of DBP. However, since phthalate esters also activate TRPV1, phthalate esters could activate other types of TRP channels non-selectively. Furthermore, sensitization to FITC is also enhanced by menthol, which activates TRPA1 and TRPM8. Here we established an in vitro system for measuring TRPM8 activation. The selectivity for TRPM8 was established by the fact that two TRPM8 agonists (menthol and icilin) induced calcium mobilization, whereas agonists of TRPA1 and TRPV1 did not. We demonstrated that phthalate esters do not activate TRPM8. TRPA1-antagonist HC-030031 did not inhibit TRPM8 activation induced by menthol or icilin. These results show that phthalate esters activate TRPA1 and TRPV1 with selectivity. TRPM8 activation is not likely to be involved in the sensitization to FITC.

Topics: Acetanilides; Animals; Calcium; CHO Cells; Cricetinae; Dermatitis, Contact; Dose-Response Relationship, Drug; Menthol; Mice; Phthalic Acids; Plasticizers; Purines; Pyrimidinones; TRPM Cation Channels; TRPV Cation Channels