PI3K/Akt signaling pathway is essential for de novo hair follicle regeneration

Abstract

Background: Cultured epidermal stem cells (Epi-SCs) and skin-derived precursors (SKPs) were capable of reconstituting functional hair follicles after implantation, while the signaling pathways that regulate neogenic hair follicle formation are poorly investigated. In this study, we aimed to understand the interactions between Epi-SCs and SKPs during skin organoid formation and to uncover key signal pathways crucial for de novo hair follicle regeneration. Methods: To track their fate after transplantation, Epi-SCs derived from neonatal C57BL/6 mice were labeled with tdTomato, and SKPs were isolated from neonatal C57BL/6/GFP mice. A mixture of Epi-SCs-tdTomato and SKPs-EGFP in Matrigel was observed under two-photon microscope in culture and after implantation into excisional wounds in nude mice, to observe dynamic migrations of the cells during hair follicle morphogenesis. Signaling communications between the two cell populations were examined by RNA-Seq analysis. Potential signaling pathways revealed by the analysis were validated by targeting the pathways using specific inhibitors to observe a functional loss in de novo hair follicle formation. Results: Two-photon microscopy analysis indicated that when Epi-SCs and SKPs were mixed in Matrigel and cultured, they underwent dynamic migrations resulting in the formation of a bilayer skin-like structure (skin organoid), where Epi-SCs positioned themselves in the outer layer; when the mixture of Epi-SCs and SKPs was grafted into excisional wounds in nude mice, a bilayer structure resembling the epidermis and the dermis formed at the 5th day, and de novo hair follicles generated subsequently. RNA-Seq analysis of the two cell types after incubation in mixture revealed dramatic alterations in gene transcriptome, where PI3K-Akt signaling pathway in Epi-SCs was significantly upregulated; meanwhile, elevated expressions of several growth factors and cytokine potentially activating PI3K were found in SKPs, suggesting active reciprocal communications between them. In addition, inhibition of PI3K or Akt by specific inhibitors markedly suppressed the hair follicle regeneration mediated by Epi-SCs and SKPs. Conclusions: Our data indicate that the PI3K-Akt signaling pathway plays a crucial role in de novo hair follicle regeneration, and the finding may suggest potential therapeutic applications in enhancing hair regeneration.

Identification of Key Pathways and Genes Related to the Development of Hair Follicle Cycle in Cashmere Goats

“…Focal adhesions can engage with the surrounding ECM [ 64 ] and relate to the migration of HF stem cells towards the bulb region [ 65 ]. The effect of the PI3K/AKT signaling pathway on the regeneration of hair follicles has also been reported [ 66 ]. All these results prove that COL1A1 and COL1A2 are essential for regeneration of cashmere hair and can be used as important candidate genes for in-depth study of its function.…”

Genetic Signatures of Selection for Cashmere Traits in Chinese Goats

“…The Wnt signalling pathway has emerged as a potential regulator of hematopoietic stem cell self-renewal [ 69 ]. The PI3K/Akt signaling pathway is essential for de novo hair follicle regeneration [ 70 ]. Moreover, the role of melatonin in promoting Cashmere yield has been demonstrated for decades [ 71 ], and melatonin also participates in stem cell differentiation pathways.…”

Functional hair follicle regeneration: an updated review

“…A mixture of Epi-SCs and SKPs was grafted into excisional wounds in nude mice, and a bilayer structure resembling the epidermis and the dermis formed on the fifth day, followed by de novo HF. 80 In the regeneration process, the SKPs formed DPCs in neogenic HFs and abundant dermal cells in the dermis, and the Epi-SCs formed the epidermis and trunk of the HF. More importantly, this experiment also demonstrates that the PI3K-Akt signalling pathway plays a crucial role in the interactions between Epi-SCs and SKPs and de novo HF regeneration, which may suggest potential therapeutic applications in enhancing hair regeneration.…”

“…More importantly, this experiment also demonstrates that the PI3K-Akt signalling pathway plays a crucial role in the interactions between Epi-SCs and SKPs and de novo HF regeneration, which may suggest potential therapeutic applications in enhancing hair regeneration. 80 During the hair growth cycle, HFSCs periodically switch between the active and inactive stages to maintain stem cell populations and generate new HFs, while this potential is impaired in aged HFSCs. 81 However, in transplantation assays in vivo, aged HFSCs could still regenerate HFs when supported with the young dermis, while young HFSCs failed to regenerate HFs when combined with the aged dermis, which shows that the ageing skin microenvironment dictates stem cell behaviour and illustrates the dominant role of the niche microenvironment.…”