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Updated: Aug 8, 2026
Isolation and Culture of Primary Mouse Keratinocytes from Neonatal and Adult Mouse Skin
Published on: July 14, 2017
Differentiation of human keratinocytes is associated with a progressive loss of interferon gamma-induced
M C Little1, D J Gawkrodger, S Mac Neil
1University Department of Medicine, Northern General Hospital, Sheffield, U.K.
Insights
Differentiated keratinocytes lose their ability to increase intercellular adhesion molecule-1 (ICAM-1) expression in response to interferon-gamma (IFN-gamma). This suggests only basal keratinocytes are responsive to inflammatory cytokines in skin inflammation.
Area of Science:
- Dermatology
- Immunology
- Cell Biology
Background:
- Intercellular adhesion molecule-1 (ICAM-1) is crucial for leukocyte-keratinocyte interactions in inflammatory skin conditions.
- Keratinocyte upregulation of ICAM-1 is observed in inflammatory dermatoses, influenced by cytokines like interferon-gamma (IFN-gamma).
- The impact of keratinocyte differentiation state on ICAM-1 inducibility by IFN-gamma requires further investigation.
Purpose of the Study:
- To test if keratinocyte differentiation diminishes their capacity to upregulate ICAM-1 expression upon IFN-gamma stimulation.
- To correlate changes in ICAM-1 responsiveness with keratinocyte differentiation markers.
Main Methods:
- Keratinocytes from 13 donors were cultured in differentiation-permissive (Green's medium) or differentiation-inhibiting (MCDB153) conditions.
- Interferon-gamma (IFN-gamma) induced ICAM-1 expression was quantified using a cell-based ELISA.
- Keratinocyte differentiation was assessed by measuring involucrin expression.
Main Results:
- Keratinocytes in MCDB153 consistently responded to IFN-gamma over 4 days.
- Keratinocytes in Green's medium showed a decreased response to IFN-gamma, with significant upregulation only at high concentrations by day 3.
- By day 4, keratinocytes in Green's medium no longer upregulated ICAM-1 in response to IFN-gamma, paralleling increased involucrin expression.
Conclusions:
- Keratinocyte differentiation is associated with a progressive loss of responsiveness to IFN-gamma-induced ICAM-1 upregulation.
- Basal, proliferative keratinocytes are likely the primary responders to inflammatory cytokines in vivo.
- This finding supports the role of basal keratinocytes in initiating and amplifying inflammatory skin reactions.
Abstract:
Intercellular adhesion molecule-1 (ICAM-1) expression is a necessary requirement for leucocyte/keratinocyte interactions, and keratinocyte upregulation of ICAM-1 is seen in several inflammatory dermatoses. While keratinocytes have a very low constitutive expression of ICAM-1, they can be induced to upregulate ICAM-1 in response to several inflammatory cytokines, such as interferon-gamma (IFN gamma). There is some evidence to suggest that the state of keratinocyte differentiation may influence the level of ICAM-1 expression induced by IFN gamma. The purpose of this study was to investigate the hypothesis that keratinocytes, as they differentiate, may lose their ability to increase their expression of ICAM-1 in response to IFN gamma. Keratinocytes from 13 donors were cultured under conditions which either permitted (Green's medium) or inhibited (MCDB153) differentiation. The ICAM-1 expression in response to IFN gamma was assessed by a sensitive, cell-based, ELISA, and related to the state of differentiation of the cells (as assessed by a quantitative assay for involucrin). While keratinocytes grown in MCDB153 remained responsive to IFN gamma throughout 4 days of culture, the response of keratinocytes grown in Green's medium progressively decreased, so that, by day 3, only a high concentration of IFN gamma (500 U/ml) led to a significant increase in ICAM-1 expression. After 4 days in culture, no upregulation of ICAM-1 was seen. The deterioration in the response of Green's-cultured keratinocytes to IFN gamma mirrored an increase in their expression of involucrin in parallel cultures. Overall, our data demonstrate a progressive loss in the ability of keratinocytes to upregulate ICAM-1 in response to IFN gamma, which is associated with differentiation of these cells. This would support the view that only basal proliferative keratinocytes are responsive to inflammatory cytokines, and play a part in the initiation and amplification of inflammatory reactions in vivo.
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