Effect of IFN-γ on the kynurenine/tryptophan ratio in monolayer-cultured keratinocytes and a 3D reconstructed human epidermis model.
Cells, Cultured
Chromatography, High Pressure Liquid
Culture Media
/ metabolism
Epidermis
/ immunology
Humans
Immunohistochemistry
Indoleamine-Pyrrole 2,3,-Dioxygenase
/ analysis
Interferon-gamma
/ metabolism
Interleukin-6
/ metabolism
Interleukin-8
/ metabolism
Keratinocytes
/ immunology
Kynurenine
/ analysis
Metabolic Networks and Pathways
/ immunology
Primary Cell Culture
/ methods
Recombinant Proteins
/ metabolism
Tryptophan
/ analysis
IDO
Kynurenine
Pro-inflammatory cytokines
Reconstructed human epidermis
Tryptophan
Journal
Journal of dermatological science
ISSN: 1873-569X
Titre abrégé: J Dermatol Sci
Pays: Netherlands
ID NLM: 9011485
Informations de publication
Date de publication:
Sep 2020
Sep 2020
Historique:
received:
16
03
2020
revised:
10
07
2020
accepted:
15
07
2020
pubmed:
13
8
2020
medline:
16
6
2021
entrez:
13
8
2020
Statut:
ppublish
Résumé
Interferon-gamma (IFN-γ) represents a potent inducer for keratinocyte inflammatory and immune activation in vitro. Since tryptophan (trp) conversion to kynurenine (kyn) is involved in inflammation, the topical kyn/trp ratio may serve as a biomarker of skin inflammation. However, the trp metabolism in keratinocytes exposed to IFN-γ is not yet fully understood. The aim of this study was to establish a human epidermis model in order to quantify cytokine and kyn/trp secretion from IFN-γ stimulated cells and tissues. Moreover, to compare the cell response of 2D-cultured keratinocytes and the 3D epidermis model. Polycarbonate filters were used on which primary keratinocytes could attach and stratify in order to form the typical layers of reconstructed human epidermis (RHE). After IFN-γ treatment, secretion of kyn/trp was measured by high performance liquid chromatography. Gene and protein expression of indoleamine 2,3-dioxygenase 1 (IDO) was analyzed with real-time PCR and immunohistochemistry. The secretion of cytokines was quantified with ELISA. Trp catabolism to kyn was significantly increased (P < 0.01) in the 2D culture in response to IFN-γ treatment. Before kyn secretion, IDO was strongly upregulated (P < 0.001). IFN-γ treatment also increased the secretion of IL-6 and IL-8 from the keratinocytes. In the RHE, IDO was upregulated by IFN-γ, and kyn secretion could be detected. Interestingly, IDO expression was only present in the basal cells of the RHE. Our results suggest that IFN-γ acts as an inducer of trp degradation preferentially in undifferentiated keratinocytes, indicated by the IDO expression in the basal layer of the RHE.
Sections du résumé
BACKGROUND
BACKGROUND
Interferon-gamma (IFN-γ) represents a potent inducer for keratinocyte inflammatory and immune activation in vitro. Since tryptophan (trp) conversion to kynurenine (kyn) is involved in inflammation, the topical kyn/trp ratio may serve as a biomarker of skin inflammation. However, the trp metabolism in keratinocytes exposed to IFN-γ is not yet fully understood.
OBJECTIVE
OBJECTIVE
The aim of this study was to establish a human epidermis model in order to quantify cytokine and kyn/trp secretion from IFN-γ stimulated cells and tissues. Moreover, to compare the cell response of 2D-cultured keratinocytes and the 3D epidermis model.
METHODS
METHODS
Polycarbonate filters were used on which primary keratinocytes could attach and stratify in order to form the typical layers of reconstructed human epidermis (RHE). After IFN-γ treatment, secretion of kyn/trp was measured by high performance liquid chromatography. Gene and protein expression of indoleamine 2,3-dioxygenase 1 (IDO) was analyzed with real-time PCR and immunohistochemistry. The secretion of cytokines was quantified with ELISA.
RESULTS
RESULTS
Trp catabolism to kyn was significantly increased (P < 0.01) in the 2D culture in response to IFN-γ treatment. Before kyn secretion, IDO was strongly upregulated (P < 0.001). IFN-γ treatment also increased the secretion of IL-6 and IL-8 from the keratinocytes. In the RHE, IDO was upregulated by IFN-γ, and kyn secretion could be detected. Interestingly, IDO expression was only present in the basal cells of the RHE.
CONCLUSION
CONCLUSIONS
Our results suggest that IFN-γ acts as an inducer of trp degradation preferentially in undifferentiated keratinocytes, indicated by the IDO expression in the basal layer of the RHE.
Identifiants
pubmed: 32782183
pii: S0923-1811(20)30234-6
doi: 10.1016/j.jdermsci.2020.07.005
pii:
doi:
Substances chimiques
CXCL8 protein, human
0
Culture Media
0
IDO1 protein, human
0
IFNG protein, human
0
IL6 protein, human
0
Indoleamine-Pyrrole 2,3,-Dioxygenase
0
Interleukin-6
0
Interleukin-8
0
Recombinant Proteins
0
Kynurenine
343-65-7
Interferon-gamma
82115-62-6
Tryptophan
8DUH1N11BX
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
177-184Informations de copyright
Copyright © 2020 The Authors. Published by Elsevier B.V. All rights reserved.
Déclaration de conflit d'intérêts
Declaration of Competing Interest The authors have no conflict of interest to declare.