Abstract
Background: Wound healing is a dynamic, sequential,and complex physiological process, including a variety of cellular events, such as proliferation, adhesion, chemotaxis, and apoptosis. Skin fibroblasts and keratinocytes are the two most important cells involved in wound repair, and Relying on the proliferation and differentiation of keratinocytes to form epithelium to completely cover the wound is the most ideal result for wound repair, so expanding the source of keratinocytes is a huge challenge. In this study, we examined the phenomenon that fetal skin fibroblasts spontaneously transdifferentiated into keratinocyte-like cells in conventional culture, and evaluated the characteristics of KLCs and the potential mechanisms of the transdifferentiation process.Methods: HFF-1 were routinely cultured in ordinary DMEM medium for more than 40 days,and observed the cell morphology. The cytological properties of KLCs at the cellular and molecular levels were detected by RT-PCR, Western-blot, immunofluorescence, Transwell, and cell scratch experiments.The functionality and safety of KLCs were determined through wound healing and tumorigenicity experiments. And high-throughput transcriptome sequencing (RNA-seq) was performed to explore the mechanism underlying HFF-1 transdifferentiation.Results: The transdifferentiation process started on the 25th day and was completed by the 40th day. KLCs and KCs had similar expressions at the molecular and protein levels, both functioned similarly in wound healing and were non-tumorigenic.RNA-seq revealed that the transdifferentiation process was regulated by the activation of the classical Wnt/β-catenin signaling pathway, which could shorten the process to 10 days.Conclusion: This study demonstrates that HFF-1 can spontaneously transdifferentiate into KLCs with conventional culture conditions, and the Wnt/β-catenin signaling pathway regulates the transdifferentiation process.
GMP-Compliant Human Fetal Skin Fibroblasts for Wound Healing
