Skin sensitisation to fragrance hydroperoxides: interplay between dendritic cells, keratinocytes and free radicals Short title: Dendritic cell activation and radicals of terpene fragrance hydroperoxides

Background: Positive patch test reactions to mixtures of oxidized terpenes con-

To improve the prediction of the possible allergenicity of chemicals in contact with the skin, investigations of upstream events are required to better understand the molecular mechanisms involved in the initiation of allergic reactions. Ascaridole, one of the compounds responsible for skin sensitization to aged tea tree oil, degrades into intermediates that evolve via different mechanisms involving radical species. We aimed at broadening the knowledge about the contribution of radical intermediates derived from ascaridole to the skin sensitization process by assessing the reactivity profile towards amino acids, identifying whether free radicals are formed in a reconstructed human epidermis (RHE) model and their biological properties to activate the immune system, namely dendritic cells in their natural context of human HaCaT keratinocytes and RHE. Electron paramagnetic resonance combined to spin-trapping in EpiSkin TM RHE confirmed the formation of C-radicals in the epidermal tissue from 10 mM ascaridole concentration, while reactivity studies toward amino acids showed electrophilic intermediates issued from radical rearrangements of ascaridole as the main reactive species. Activation of THP-1 cells, as surrogate for dendritic cells, that were cocultured with HaCaT was significantly upregulated after treatment with low micromolar concentrations based on cell surface expression of the co-stimulatory molecule CD86 and the adhesion molecule CD54. Placing THP-1 cells underneath the RHE allowed us to monitor which of the concentrations that produce radical(s) and/or protein antigens in the epidermal skin environment promote the activation of dendritic cells. We detected no significant upregulation of CD86/CD54 after topical RHE application of concentrations up to 30 mM ascaridole (t=24 h) but clear upregulation after 60 mM.

Dermal exposure to cumene hydroperoxide (CumOOH) during manufacturing processes is a toxicological issue for the industry. Its genotoxicity, mutagenic action, ability to promote skin tumor, capacity to induce epidermal hyperplasia and aptitude to induce allergic and irritant skin contact dermatitis are well known. These toxic effects appear to be mediated through the activation to free radical species such as hydroxyl, alkoxyl and alkyl radicals characterized basically by electron paramagnetic resonance (EPR) and spin-trapping (ST) techniques. To be a skin sensitizer CumOOH needs to covalently bind to skin proteins in the epidermis to form the antigenic entity triggering the immunotoxic reaction. Cleavage of the O-O bond allows formation of unstable CumO • /CumOO • radicals rearranging to longer half-life specific carbon-centered radicals R • proposed to be at the origin of the antigen formation. Nevertheless, it is not still clear which R • are precisely formed in the epidermis and thus involved in the sensitization process. The aim of this work was to elucidate in conditions closer to real-life sensitization which specific R • are formed in a 3D reconstructed human epidermis (RHE) model by using 13 C-substituted CumOOH at carbon positions precursors of potentially reactive radicals and EPR-ST. We demonstrated that most probably methyl radicals derived from -scission of CumO • radicals occur in RHE through a one-electron reductive pathway suggesting that these could be involved in the antigen formation inducing skin sensitization. We also describe a coupling between nitroxide radicals and  position 13 C atoms that could be of an added value to the very few examples existing for the coupling of radicals with 13 C atoms.

No abstract

Blastomycosis‐like pyoderma is a rare skin disorder most commonly caused by bacterial infection. It is usually diagnosed in immunocompromised patients. We report a case of BLP in an immunocompetent woman, who presented with a 6‐week history of verrucous cutaneous plaque of the left wrist.

between positive intradermal tests and breakthrough reactions during RDD (p<0.0001). Patients that had negative intradermal test did not react during RDD. The intravenous protocol was safer than the oral protocol (p ¼ 0.047). The other 24/63 (38.1%) patients considered of low risk were challenged with penicillin and only one reacted (1.6%). CONCLUSIONS Risk stratification and RDD were safe and effective. Intradermal skin testing identified patients with increased risk of reaction during RDD. More patients should be included in the study to confirm the superiority of the intravenous RDD protocol.

Bastomycosis-like pyoderma is a rare skin disorder most commonly caused by bacterial infection. It is usually diagnosed in immunocompromised patients. We report a case of BLP in an immunocompetent woman, who presented with a 6-week history of verrucous cutaneous plaque of the left wrist.