Abstract
Cultures of epithelial cells are limited by the proliferative capacity of primary cells and cell senescence. Herein we show that primary human epithelial cell sheets cultured without dermal equivalents maintained homeostasis in vitro for at least 1 year. Transparency of these sheets enabled live observation of pigmented melanocytes and Fluorescent Ubiquitination-based Cell Cycle Indicator (FUCCI) labeled epithelial cells during wound healing. Cell turn over and KRT15 expression pattern stabilized within 3 months, when KRT15 bright clusters often associated with niche-like melanocytes became apparent. EdU labels were retained in a subset of epithelial cells and melanocytes after 6 months chasing, suggesting their slow cell cycling property. FUCCI-labeling demonstrated robust cell migration and proliferation following wounding. Transparency and long-term (1 year) homeostasis of this model will be a powerful tool for the study of wound healing and cell linage tracing.
Highlights
Weeks, long-term observation was not performed[27]
In order to obtain an ideal in vitro model of the human epithelial stem cell niche, we previously reported that replacing epidermal growth factor (EGF) with fibroblast growth factor 7 (FGF7 or keratinocyte growth factor; KGF) combined with the rho kinase inhibitor Y27632 can extend the culture life of a confluent epithelial cell sheet for up to 3 months[29]
Wound healing ability of KY sheets. Since both continuous cell turnover and wound healing are required for epithelial homeostasis, we examined the wound healing ability of 3-month KY sheets in a steady state
Summary
Weeks, long-term observation was not performed[27]. Transplanted epithelium can reconstruct damaged ocular surface over a long term, especially when autologous cells sources are used[28]. Cultured epithelial cell sheets may maintain homeostasis in vivo after transplantation, it is difficult to perform detailed examination of these cells due to ethical issues. We further show that KY sheets can maintain homeostasis for over 1 year, and can undergo wound healing demonstrated by live fluorescence imaging. We report changes in cell turnover and the expression pattern of the epithelial stem cell marker during the one-year culture period
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
