Abstract
ObjectiveEpidermal stem cells (EpSCs) can self-renew, which are responsible for the long-term maintenance of the skin, and it also plays a critical role in wound re-epithelization, but the mechanism underlying EpSCs proliferation is unclear. GDF-5, also known as BMP-14, is a member of the BMP family and can be used as a self-renewal supporter. Here, we studied the effects of GDF-5 on mouse EpSCs proliferation mechanism in wound healing.MethodsFirstly, the effects of GDF-5 on EpSCs proliferation was tested by using CCK8 reagent and PCNA expression was analyzed by Western blotting. Secondly, we screened genes that promote EpSCs proliferation in the FOX and cyclin family by qPCR, and then the protein expression level of the selected genes was further analyzed by Western blotting. Thirdly, siRNA plasmids and pAdEasy adenovirus were transfected or infected, respectively, into mouse EpSCs to detect the effect of target genes on GDF-5-induced cell proliferation. Furthermore, we injected GDF-5 to a deep partial thickness burn mouse model for finding out whether EpSCs proliferation can be detected by immunohistochemical. Finally, the relevant target genes were analyzed by qPCR, immunoblotting, and dual-luciferase reporter gene detection.ResultsWe discovered that 100 ng/ml recombinant mouse GDF-5 was the optimal concentration for promoting mouse EpSCs proliferation. Through preliminary screened by qPCR, we found that Foxg1 and cyclin D1 could be the downstream molecules of GDF-5, and the results were confirmed by Western blotting. And the effect of GDF-5 on mouse EpSCs proliferation was adjusted by Foxg1/cyclin D1 in vitro and in vivo. Besides, GDF-5-induced transcription of cyclin D1 was regulated by Foxg1-mediated cyclin D1 promoter activity.ConclusionThis paper showed that GDF-5 promotes mouse EpSCs proliferation via Foxg1-cyclin D1 signal pathway. It is suggested that GDF-5 may be a new approach to make EpSCs proliferation which can be used in wound healing.
Highlights
The epidermis is derived from ectoderm cells during embryonic development, these cells go through a layering process to form basal, spinous, and granular layers [1]
We screened downstream genes in the FOX and cyclins family, the results discovered that Foxg1 and cyclin D1 were a dose-dependent relationship with Growth/differentiation factor 5 (GDF-5)
The protein levels of Foxg1 and cyclin D1 in mouse Epidermal stem cells (EpSCs) treated by GDF-5 were detected by Western blotting (WB)
Summary
The epidermis is derived from ectoderm cells during embryonic development, these cells go through a layering process to form basal, spinous, and granular layers [1]. The skin epidermis contains different appendages including sweat glands, hair follicles, and sebaceous glands, †. The self-renewal and damage repairing of skin tissue mainly depend on the compensatory proliferation and differentiation of EpSCs [3]. After Billingham and Reynolds firstly reported skin cell transplantation for wound healing in 1952 [4], EpSCs have been used in clinical practice, repairing of burns, acute trauma, and skin damage caused by certain diseases [5,6,7]. The expansion of EpSCs always is a choke point in its clinical application. The physiological state of EpSCs can be affected by different signaling
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