Reporter system controlled by the involucrin promoter as a tool to follow epidermal differentiation

Various approaches have been explored to study skin biology, including the use of stem cells. Mesenchymal stem cells (MSCs) from the umbilical cord can be safely and easily obtained; however, a simple strategy to monitor their differentiation is essential. Involucrin is a marker of keratinocyte diff...

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Bibliographic Details
Published in:Biochimie 2022-10, Vol.201, p.33-42
Main Authors: Fernandes, Myrian Thiago Pruschinski, dos Santos, Jeniffer Farias, Freitas, Bruna Letícia, Reigado, Gustavo Roncoli, Antunes, Fernanda, Tessarollo, Nayara Gusmão, Chambergo, Felipe Santiago, Strauss, Bryan Eric, Nunes, Viviane Abreu
Format: Article
Language:English
Subjects:
Cell differentiation
Epidermis
Involucrin promoter
Keratinocytes
Mesenchymal stem cells
Reporter genes
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Summary:Various approaches have been explored to study skin biology, including the use of stem cells. Mesenchymal stem cells (MSCs) from the umbilical cord can be safely and easily obtained; however, a simple strategy to monitor their differentiation is essential. Involucrin is a marker of keratinocyte differentiation, and its promoter (pINV) directs stratum-specific expression of this protein. We designed a reporter system containing EGFP under the control of pINV to assess MSC transdifferentiation into keratinocytes. The functional sequence of pINV was inserted into a lentiviral vector, producing LeGO-GpINV. MSCs were transduced with LeGO-GpINV and induced to transdifferentiate into keratinocytes under cultivation with keratinocyte serum-free medium. MSC transdifferentiation was confirmed by morphological changes and by the expression of epidermal markers by flow cytometry, quantitative PCR, Western blot and the activity of epidermal kallikreins 5, 6 and 7. After 14 days of transdifferentiation, MSCs transduced with LeGO-GpINV showed an increase in EGFP fluorescence and expressed CK10, CK14, involucrin and filaggrin. There was also an increase in kallikrein activity. This reporter system allowed us to temporally assess epidermal differentiation, simultaneously with involucrin expression, opening possibilities for the in vivo study of skin biology and in regenerative medicine. •Involucrin promoter as a tool to follow epidermal differentiation.•A GFP-based lentiviral vector can be used to monitor MSC epidermal differentiation.•MSC induced to epidermal fate express GFP under involucrin promoter control.
ISSN:0300-9084
1638-6183
DOI:10.1016/j.biochi.2022.06.014