Dataset Information

Ficoll density gradient sedimentation isolation of pelage hair follicle mesenchymal stem cells from adult mouse back skin: a novel method for hair follicle mesenchymal stem cells isolation

ABSTRACT:

Background

Hair follicle mesenchymal stem cells (HF-MSCs) have great potential for cell therapy. Traditional method to isolate whisker HF-MSC is time-consuming and few in cell numbers. How to quickly and conveniently obtain a large number of HF-MSC for experimental research is a problem worth exploring.

Methods

Two-step Ficoll Density Gradient Sedimentation (FDGS) was performed to isolate pelage HF-MSC from adult mice. The characteristic of the isolated cells was identified and compared with whisker HF-MSC by immunofluorescence staining, flow cytometry, three-lineage differentiation and hair follicle reconstruction. Pelage HF-MSC and exosomes were injected into the dorsal skin of mice as well as hair follicle organ culture to explore its role in promoting hair growth. The cells and exosomes distribution were located by immunofluorescence staining.

Results

Isolated pelage HF-MSC expressed similar markers (ALP, Versican, NCAM, Nestin), showed similar growth pattern, possessed similar mesenchymal stem cells function and hair follicle induction ability as whisker HF-MSC. A large number of cells can be obtained with fewer mice compared to traditional method. Injected pelage HF-MSC promoted hair growth by secreting exosomes.

Conclusion

A large number of Pelage HF-MSC can be isolated by FDGS, which can promote hair growth by secreting exosomes which may target the dermal papilla and hair matrix region of host hair follicle.

Supplementary Information

The online version contains supplementary material available at 10.1186/s13287-022-03051-3.

SUBMITTER: Gan Y

PROVIDER: S-EPMC9330686 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundAlopecia affects millions of individuals globally, with hair loss becoming more common among young people. Various traditional Chinese medicines (TCM) have been used clinically for treating alopecia, however, the effective compounds and underlying mechanism are less known. We sought to investigate the effect of Alpinetin (AP), a compound extracted from Fabaceae and Zingiberaceae herbs, in hair regeneration.MethodsAnimal model for hair regeneration was mimicked by depilation in C57BL/6J mice. The mice were then topically treated with 3 mg/ml AP, minoxidil as positive control (PC), or solvent ethanol as vehicle control (VC) on the dorsal skin. Skin color changes which reflected the hair growth stages were monitored and pictured, along with H&E staining and hair shaft length measurement. RNA-seq analysis combined with immunofluorescence staining and qPCR analysis were used for mechanism study. Meanwhile, Gli1CreERT2; R26RtdTomato and Lgr5EGFP-CreERT2; R26RtdTomato transgenic mice were used to monitor the activation and proliferation of Gli1+ and Lgr5+ HFSCs after treatment. Furthermore, the toxicity of AP was tested in keratinocytes and fibroblasts from both human and mouse skin to assess the safety.ResultsWhen compared to minoxidil-treated and vehicle-treated control mice, topical application of AP promoted anagen initiation and delayed catagen entry, resulting in a longer anagen phase and hair shaft length. Mechanistically, RNA-seq analysis combined with immunofluorescence staining of Lef1 suggested that Lgr5+ HFSCs in lower bulge were activated by AP via Wnt signaling. Other HFSCs, including K15+, Lef1+, and Gli1+ cells, were also promoted into proliferating upon AP treatment. In addition, AP inhibited cleaved caspase 3-dependent apoptosis at the late anagen stage to postpone regression of hair follicles. More importantly, AP showed no cytotoxicity in keratinocytes and fibroblasts from both human and mouse skin.ConclusionThis study clarified the effect of AP in promoting hair regeneration by activating HFSCs via Wnt signaling. Our findings may contribute to the development of a new generation of pilatory that is more efficient and less cytotoxic for treating alopecia.