Understanding the skin sensitization capacity of ascaridole: a combined study of chemical reactivity and activation of the innate immune system (dendritic cells) in the epidermal environment.

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™ 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.

Non-invasive monitoring of immunization progress in mice via IgG from feces.

A non-invasive method to monitor the humoral immune response in mice after immunization is described. From fecal pellets of an individual mouse, a sufficient amount of active immunoglobulins or their fragments can be extracted to perform a regular examination of the status of the immune response by immunoassay. Hapten-specific antibodies from the feces of mice from three immunization trials showed very similar characteristics to those obtained from serum at a given date. Therefore, it can be suspected that some serum IgG enters the intestinal lumen and ends up in the feces, where they appear to be considerably stable. Hapten-specific IgAs were not found in the feces. Being able to analyze antibody titers in feces could be an interesting animal welfare refinement to standard practice that does not entail repeated blood sampling.

Free radicals in antigen formation: reduction of contact allergic response to hydroperoxides by epidermal treatment with antioxidants.

BACKGROUND: For patients with allergic contact dermatitis, the main therapy is anti-inflammatory steroids, a non-specific and symptomatic treatment. In contact allergy, the antigen formation is considered to be the binding of a chemical (hapten) to a biological macromolecule, e.g. a protein. Limonene-2-hydroperoxide (Lim-OOH) is a hapten with a known allergenic effect. It is likely to bind to proteins in the skin via a radical mechanism. It might be possible to inhibit the allergic reaction by epidermal application of substances that can trap free radicals, e.g. antioxidants such as ascorbic acid or alpha-tocopherol, prior to the application of the hapten. OBJECTIVES: To study the influence of antioxidants on the allergenic effect of Lim-OOH in sensitization experiments on guinea pigs. METHODS: Pretreatment with the antioxidants was performed before induction to study the effect on sensitization as well as before challenge testing to study the effect on elicitation. RESULTS: A reduction in the response rate was found both at sensitization and at elicitation. The antioxidants had no effect on cobalt allergy or on the allergenic effect of haptens that form antigens via nucleophilic-electrophilic reactions. No reduction of the effect was seen for irritants. CONCLUSIONS: The protective effect of antioxidants in elicitation could be of practical therapeutic value, as it indicates a possibility for the treatment of patients who have become sensitized to haptens that form full antigens via a radical mechanism.

Validation and subsequent development of the DEREK skin sensitization rulebase by analysis of the BgVV list of contact allergens.

The DEREK knowledge-based computer system contains a subset of approximately 50 rules describing chemical substructures (toxophores) responsible for skin sensitization. This rulebase, based originally on Unilever historical in-house guinea pig maximization test data, has been subject to extensive validation and is undergoing refinement as the next stage of its development. As part of an ongoing program of validation and testing, the predictive ability of the sensitization rule set has been assessed by processing the structures of the 84 chemical substances in the list of contact allergens issued by the BgVV (German Federal Institute for Health Protection of Consumers). This list of chemicals is important because the biological data for each of the chemicals have been carefully scrutinized and peer reviewed, a key consideration in an area of toxicology in which much unreliable and potentially misleading data have been published. The existing DEREK rulebase for skin sensitization identified toxophores for skin sensitization in the structures of 71 out of the 84 chemicals (85%). The exercise highlighted areas of chemistry where further development of the rulebase was required, either by extension of the scope of existing rules or by generation of new rules where a sound mechanistic rationale for the biological activity could be established. Chemicals likely to be acting as photoallergens were identified, and new rules for photoallergenicity have subsequently been written. At the end of the exercise, the refined rulebase was able to identify toxophores for skin sensitization for 82 of the 84 chemicals in the BgVV list.

Inhibition of lymphocyte proliferation and antibody production in vitro by silica, talc, bentonite or Corynebacterium parvum: involvement of peroxidative processes.

This study was undertaken to determine whether and by what means particles which induce granulomata in vivo can affect murine spleen lymphoproliferative and antibody responses in vitro. Particles of silica, talc, Bentonite or C. parvum cells inhibited lipopolysaccharide- or concanavalin A-stimulated proliferation and sheep red blood cell-induced antibody response in vitro. The inhibition required at least 48 hours exposure of the cells to the particles. The late onset of inhibition and its reproducibility at different cell or mitogen concentrations implicated particle-induced injury to both phagocytes and lymphocytes. Either alpha-tocopherol or 2-mercaptoethanol prevented the particle-induced inhibition of spleen cell responses. alpha-Tocopherol and 2-mercaptoethanol have in common the capacity to protect cells against membrane lipid peroxidation. The inhibitory peroxidative process(es) implicated by these studies are most likely attributable to: (a) stimulation of oxidative metabolism of phagocytic cells by particles; and (b) iron-catalyzed peroxidation directly by the particles. These data may be relevant in understanding the pathogenesis of and devising therapeutic approaches toward various granulomatous conditions.

Benzoyl peroxide: lack of sensitization.

A prospective study of 445 patients using benzoyl peroxide, observed regularly over a 12-month period, uncovered only a very low incidence of moderate or severe local reaction. There was, however, a significant level of mild primary irritant dermatitis which settled with continued use. In only 6 patients was there severe local reaction; one patient was lost to follow-up but in the 5 patients patch tested, only one had a contact allergic dermatitis. The study indicates the overall cutaneous safety of benzoyl peroxide.