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Biotransformation of 2,4-toluenediamine in human skin and reconstructed tissues

  • Toxicokinetics and Metabolism
  • Published:
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Abstract

Reconstructed human epidermis (RHE) is used for risk assessment of chemicals and cosmetics and RHE as well as reconstructed human full-thickness skin (RHS) become important for e.g., the pre-clinical development of drugs. Yet, the knowledge regarding their biotransformation capacity is still limited, although the metabolic activity is highly relevant for skin sensitization, genotoxicity, and the efficacy of topical dermatics. The biotransformation of the aromatic amine 2,4-toluenediamine (2,4-TDA) has been compared in two commercially available RHS to normal human skin ex vivo, and in primary epidermal keratinocytes and dermal fibroblasts as well as in vitro generated epidermal Langerhans cells and dermal dendritic cells. The mono N-acetylated derivative N-(3-amino-4-methyl-phenyl)acetamide (M1) was the only metabolite detectable in substantial amounts indicating the predominance of N-acetylation. RHS exceeded human skin ex vivo in N-acetyltransferase activity and in cell cultures metabolite formation ranked as follows: keratinocytes > fibroblasts ~ Langerhans cells ~ dendritic cells. In conclusion, our results underline the principal suitability of RHS as an adequate test matrix for the investigation of N-acetylation of xenobiotics which is most relevant for risk assessment associated with cutaneous exposure to aromatic amines.

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Abbreviations

2,4-TDA:

2,4-Toluenediamine

DMSO:

Dimethylsulfoxide

ESI-TOF:

Electrospray ionization-time of flight

FCS:

Fetal calve serum

HPLC:

High-performance liquid chromatography

IL:

Interleukin

IUPAC:

International Union of Pure and Applied Chemistry

LC-MS:

Liquid chromatography-mass spectrometry

LDH:

Lactate dehydrogenase

M1:

N-(3-Amino-4-methyl-phenyl)acetamide

M2:

N-(5-Amino-2-methylphenyl)acetamide

MoDC:

Monocyte-derived dendritic cells

MoLC:

Monocyte-derived Langerhans cells

MTT:

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NAT:

N-Acetyltransferase

NMR:

Nuclear magnetic resonance

OD:

Optical density

PBS:

Phosphate buffered saline

PPD:

Para-phenylenediamine

REACh:

Registration, Evaluation, Authorization and Restriction of Chemicals

RHE:

Reconstructed human epidermis

RHS:

Reconstructed human full-thickness skin

SD:

Standard deviation

SDS:

Sodium dodecyl sulfate

TRIS:

Tris(hydroxymethyl)aminomethane

UV:

Ultraviolet light

XME:

Xenobiotic metabolizing enzyme

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Acknowledgements

This work was part of a joint project pre-validating RHS for comet assay and its financial support by the German Federal Ministry for Education and Research (project no. 0316008B) is gratefully acknowledged. We thank Dr Maria Parr for critical discussion and Ruth Hesse, Gerrit Müller, and the NMR core facility of the Freie Universität Berlin for excellent technical support.

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Correspondence to Günther Weindl.

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Grohmann, L., Becker, D., Rademann, J. et al. Biotransformation of 2,4-toluenediamine in human skin and reconstructed tissues. Arch Toxicol 91, 3307–3316 (2017). https://doi.org/10.1007/s00204-017-1954-5

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  • DOI: https://doi.org/10.1007/s00204-017-1954-5

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