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- Andresen Bergström, Moa, 1978, et al.
(författare)
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A skin-like cytochrome P450 cocktail activates prohaptens to contact allergenic metabolites.
- 2007
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Ingår i: The Journal of investigative dermatology. - : Elsevier BV. - 1523-1747 .- 0022-202X. ; 127:5, s. 1145-53
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Tidskriftsartikel (refereegranskat)abstract
- Allergic contact dermatitis is a complex syndrome representing immunological responses to cutaneous exposure to protein-reactive chemicals. Although many contact sensitizers directly can elicit this disorder, others (prohaptens) require activation. Knowledge regarding the activating mechanisms remains limited, but one possibility is metabolic activation by cytochrome P450 (CYP) enzymes in the skin. We have, after quantitative reverse transcriptase-PCR studies of the CYP content in 18 human skin samples, developed an enriched skin-like recombinant human (rh) CYP cocktail using CYP1A1, 1B1, 2B6, 2E1, and 3A5. To validate the rhCYP cocktail, a prohaptenic conjugated diene ((5R)-5-isopropenyl-2-methyl-1-methylene-2-cyclohexene) was investigated using: the skin-like rhCYP cocktail, a liver-like rhCYP cocktail, single rhCYP enzymes, liver microsomes, keratinocytes, and a dendritic cell (DC) assay. The diene was activated to sensitizing epoxides in all non-cell-based incubations including the skin-like rhCYP cocktail. An exocyclic epoxide metabolite ((7R)-7-isopropenyl-4-methyl-1-oxaspiro[2.5]oct-4-ene) was found to be mainly responsible for the allergenic activity of the diene. This epoxide also induced pronounced DC activation indicated by upregulation of IL-8. The skin-like rhCYP cocktail provides a simplified alternative to using skin tissue preparations in mechanistic studies of CYP-mediated skin metabolism of prohaptens and offers the future possibility of designing in vitro predictive assays for assessment of allergenic activity of prohaptens.
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| 3. |
- Ott, Hagen, et al.
(författare)
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Cutaneous metabolic activation of carvoxime, a self-activating, skin-sensitizing prohapten.
- 2009
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Ingår i: Chemical Research in Toxicology. - : American Chemical Society (ACS). - 0893-228X .- 1520-5010. ; 22:2, s. 399-405
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Tidskriftsartikel (refereegranskat)abstract
- Bioactivation of low molecular weight compounds in the skin can cause contact sensitization. We have previously shown that the alpha, beta-R-unsaturated oxime R-carvoxime [1, (R)-2-methyl-5-isopropenylcyclohex-2-enone oxime] is bioactivated to two diastereomeric highly reactive and strongly sensitizing alpha, beta-epoxy oxime metabolites. To investigate if this metabolic activation is catalyzed by the major cytochrome P450 (P450) enzymes found in human skin, incubations of 1 with a skinlike P450 cocktail in the presence of glutathione were carried out. We identified three glutathione conjugates in the incubation mixture arising from two diasteomeric alpha, beta-epoxy oxime metabolites of 1, thus showing that the metabolic activation of 1 is P450-mediated. A P450 identification study using the individual P450 enzymes present in the skinlike P450 cocktail showed the involvement of P450 1A1 and 1B1 and also to some extent 2B6. P450 1B1 metabolism of 1 was found to be stereoselective as glutathione conjugates from only one of the alpha, beta-epoxyoxime metabolites were identified (metabolite 2). Additionally, 1 was found to be an inducer of P450 1B1 (but not 1A1) in human monocyte-derived dendritic cells (moDCs) and to some extent in normal human epidermal keratinocytes. A further transcriptional gene expression change observed in moDCs was a 44-fold upregulation of IL-8, a marker often used for assessment of sensitizing potential of contact allergens. The autoinduction of P450 1B1 by 1 may be a key event in the development of contact allergy to 1 and may also explain why only metabolite 2, and not 3, was found to elicit an allergic response in mice sensitized to 1. Our data show that the alpha, beta-unsaturated oxime 1 is bioactivated by human cutaneous P450, thus forming highly allergenic metabolites, and has the potential to induce its own bioactivation pathway, particularly in antigen-presenting cells.
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