Abstract
Lip vermilion is unique and can be distinguished from the adjacent skin and oral mucosa. However, because of the lack of appropriate evaluation tools, skin and/or oral mucosa substitutes such as in vitro vermilion epithelial models have been used for lip product testing. We aimed to develop and characterize a lip vermilion epithelium reconstruction model (LVERM) using skin and oral keratinocytes. LVERM was manufactured by co-culturing primary skin and oral keratinocytes, using a device that allowed the separation of cell seeding, and created an intercalated cell-free zone, referred to as the vermilion part. After removing the device, LVERM construction was completed in 8 days, in a submerged condition. Subsequently, they were placed in an air–liquid interface for 7 days. To determine the epithelial characteristics of LVERM, keratin 2e (KRT2) and small proline-rich protein 3 (SPRR3) expression patterns were examined. The in vivo expression profiles of KRT2 and SPRR3 genes in vermilion were also examined. We found that a continuous multi-layered epithelium was generated in the LVERM that exhibited ortho- and para-keratinization in the skin and oral mucosa parts, respectively. Although an intermediate keratinization pattern was observed in the vermilion part, KRT2 and SPRR3 were co-expressed in the suprabasal layer, consistent with the expression pattern of a single vermilion epithelial model. Clustering analysis revealed that KRT2 and SPRR3 gene expression in vermilion was location-dependent within the sample. Therefore, LVERM can be used as an evaluation tool for lip products and has great importance in innovative approaches for cosmetic testing.
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Data availability
The microarray data that support the findings of this study are openly available in [the NCBI Gene Expression Omnibus database] at [https://www.ncbi.nlm.nih.gov/ (accessed on 26 Jan 2023)], reference number [GSE222604, GSE222605].
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Acknowledgements
The authors are grateful to Akihiko Iida and staff member of Nagaoka Red Cross Hospital, Katsuhiko Honma and staff member of Tsuruoka Municipal Shonai Hospital, and Hanako Wakatsuki and staff member of the Department of Plastic and Reconstructive Surgery, Niigata University School of Medicine, for their assistance during harvesting lip tissues from patients. The authors also thank Yasumitsu Kodama, Division of Oral and Maxillofacial Surgery, Faculty of Dentistry & Graduate School of Medical and Dental Sciences, Niigata University, for the ethical issue management, and Hiroko Kato, School of Pharmacology, Osaka University, for remarkable technical assistance.
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This research was supported by KOSÉ Corporation.
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EK, EI, OS, EN and KI contributed to the study conception and design. EK, YL, RK, EH, EI, OS, SO and EN contributed reagents/materials/analysis tools. EK, YL, RK, EH and EI conducted experiments. EK, YL, EI, OS, SO, EN and KI collected and analyzed the data. EK, YL, EH, EI, SO and EN visualized data. EK and KI wrote the first draft of the paper. YL, EI, OS, SO, EN and KI commented on previous versions of the paper, edited and reviewed the paper. All authors have read and agreed to the published version of the manuscript.
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Division of Biomimetic (Ryota Kobayashi, Emi Hoshikawa, and Kenji Izumi) in Niigata University, is financially supported by a grant/research funding from KOSÉ Corporation. Eri Kobayashi, Eriko Itai, Osamu Sakata and Eiji Naru are employees of KOSÉ Corporation. Yiwei Ling and Shujiro Okuda declare no conflict of interest.
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Kobayashi, E., Ling, Y., Kobayashi, R. et al. Development of a lip vermilion epithelium reconstruction model using keratinocytes from skin and oral mucosa. Histochem Cell Biol 160, 349–359 (2023). https://doi.org/10.1007/s00418-023-02206-4
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DOI: https://doi.org/10.1007/s00418-023-02206-4