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Comparison of xenobiotic metabolizing enzyme activities in ex vivo human skin and reconstructed human skin models from SkinEthic.

Archives of toxicology (2014-03-25)
Joan Eilstein, Guillaume Léreaux, Natali Budimir, Georges Hussler, Simon Wilkinson, Daniel Duché
ABSTRACT

Skin function is not limited to a physical barrier. According to its total surface area, it is also considered as an extra-hepatic metabolizing organ. In vitro engineered human skins have been developed to replace limited ex vivo normal human skin samples (NHS). Thus, assessing and comparing skin models from SkinEthic [Episkin™, RHE™ and the full thickness model (FTM)] with NHS in terms of metabolic capability are essential. The apparent activities of main cutaneous isoforms of cytochrome P450-dependent monooxygenases (CYP1A1/1B1, 2B6/2C18/2E1, 3A5/3A7), esterase, glutathione-S-[(GST), A, M, P, T], N-acetyl-(NAT1), uridinyl-diphosphate glucuronyl-(UDPGT 1A family) and sulfo-(SULT1A1) transferases were determined using probe substrates. Mean activities indicative of CYP1A1/1B1 (expressed as pmol/mg protein/6 h) in RHE™ (2.8) and FTM (2.6) were very similar to NHS (3.0) while Episkin™ showed a higher activity (9.1). Activities of CYP3A5/3A7 in FTM (3.3) and Episkin™ (3.6) were similar to NHS (3.8) while activity in RHE™ (13.3) was higher. CYP2B6/2C18/2E1 activity was below LOQ (0.5) in all skin models and NHS. Comparable intrinsic metabolic clearances were measured between NHS and skin models for esterase, UDPGT, GST and NAT1 activities. SULT1A1 activity toward probe substrates was not detected in skin models and observed at the limit of detection in NHS. Weak cytochrome P450-dependent monooxygenases, high esterase and transferase activities suggested that NHS and skin models exhibited limited functionalization and much greater detoxification (hydrolytic and conjugating) capacities. These results demonstrate that skin models are similar to NHS in terms of metabolic functionality toward xenobiotics investigated and useful tools to assess both the local efficiency and safety of cosmetics.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Sodium dodecyl sulfate, BioReagent, suitable for electrophoresis, for molecular biology, ≥98.5% (GC)
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Trichloroacetic acid, ACS reagent, ≥99.0%
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Sodium dodecyl sulfate, ACS reagent, ≥99.0%
Supelco
Ammonium acetate, suitable for mass spectrometry (MS), LiChropur, eluent additive for LC-MS
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Sodium dodecyl sulfate, ReagentPlus®, ≥98.5% (GC)
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Iodoacetamide, BioUltra
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Ammonium acetate, for molecular biology, ≥98%
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Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 10% in H2O
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Trichloroacetic acid solution, 6.1 N
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Iodoacetamide, Single use vial of 56 mg
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Sodium dodecyl sulfate, ≥99.0% (GC), dust-free pellets
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Iodoacetamide, ≥99% (NMR), crystalline
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4-Nitrophenol, ReagentPlus®, ≥99%
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Sodium dodecyl sulfate, BioUltra, for molecular biology, ≥99.0% (GC)
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Ammonium acetate, reagent grade, ≥98%
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Ammonium acetate, ACS reagent, ≥97%
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4-Methylumbelliferone, ≥98%
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Ammonium acetate solution, for molecular biology, 7.5 M
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Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 20% in H2O
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Sodium dodecyl sulfate, BioXtra, ≥99.0% (GC)
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Coumarin, ≥99% (HPLC)
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Ammonium acetate, ≥99.99% trace metals basis
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Sodium dodecyl sulfate, 92.5-100.5% based on total alkyl sulfate content basis
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Trichloroacetic acid, ≥99.0% (titration)
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Trichloroacetic acid, BioXtra, ≥99.0%
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1-Chloro-2,4-dinitrobenzene, ≥99%
Supelco
Sodium dodecyl sulfate, dust-free pellets, suitable for electrophoresis, for molecular biology, ≥99.0% (GC)
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1,3-Dinitrobenzene, 97% anhydrous basis
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Ammonium acetate, BioUltra, for molecular biology, ≥99.0%
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4-Nitrophenol, spectrophotometric grade