Hair follicle stem cell cultures reveal self‐organizing plasticity of stem cells and their progeny

Understanding how complex tissues are formed, maintained, and regenerated through local growth, differentiation, and remodeling requires knowledge on how single‐cell behaviors are coordinated on the population level. The self‐renewing hair follicle, maintained by a distinct stem cell population, represents an excellent paradigm to address this question. A major obstacle in mechanistic understanding of hair follicle stem cell (HFSC) regulation has been the lack of a culture system that recapitulates HFSC behavior while allowing their precise monitoring and manipulation. Here, we establish an in vitro culture system based on a 3D extracellular matrix environment and defined soluble factors, which for the first time allows expansion and long‐term maintenance of murine multipotent HFSCs in the absence of heterologous cell types. Strikingly, this scheme promotes de novo generation of HFSCs from non‐HFSCs and vice versa in a dynamic self‐organizing process. This bidirectional interconversion of HFSCs and their progeny drives the system into a population equilibrium state. Our study uncovers regulatory dynamics by which phenotypic plasticity of cells drives population‐level homeostasis within a niche, and provides a discovery tool for studies on adult stem cell fate.     

毛囊干细胞及其隆突微环境

毛囊隆突(bulge)是毛囊干细胞特定的微环境,它维持并调节干细胞的特性,使干细胞在静息态、自我更新和分化上保持平衡。现重点从毛囊隆突的物理结构上来简要揭示微环境对干细胞的调控。     

Immunohistological Localization of Endogenous Unlabeled Stem Cells in Wounded Skin

Various types of endogenous stem cells (SCs) participate in wound healing in the skin at different anatomical locations. SCs need to be identified through multiple markers, and this is usually performed using flow cytometry. However, immunohistological identification of endogenous stem cells in the skin at different anatomical locations by co-staining multiple SC markers has been seldom explored. We examined the immunohistological localization of four major types of SCs in wounded skin by co-staining for their multiple markers. Hematopoietic SCs were co-stained for Sca1 and CD45; mesenchymal SCs for Sca1, CD29, and CD106; adipose SCs for CD34, CD90, and CD105; and endothelial progenitor cells and their differentiated counterparts were co-stained for CD34, Tie2, and von Willebrand factor. We found Sca1⁺CD45⁺ SCs in the epidermis, dermis and hypodermis of wounded skin. Sca1⁺CD29⁺ and Sca1⁺CD106⁺ mesenchymal SCs, CD34⁺CD105⁺, CD34⁺CD90⁺, and CD90⁺CD105⁺ adipose SCs, as well as CD34⁺Tie2⁺ endothelial progenitor cells were also located in the epidermis, dermis, and hypodermis. This study demonstrates the feasibility of using immunohistological staining to determine the location of SCs in wounded skin and the intracellular distribution of their molecular markers.     

Hair follicle and interfollicular epidermal stem cells make varying contributions to wound regeneration

Upon wounding, multiple stem cell populations in the hair follicle (HF) and interfollicular epidermis (IFE) converge at the site of injury. Although these cells can contribute permanently to the regenerating epithelium, it remains unclear whether these contributions vary among cells originating from diverse compartments in the skin. By comparing the fates of several keratinocyte lineages, we observed here an initial decrease in both HF- and IFE-derived cells within the transient acanthotic layers of the regenerating epithelium. At the same time, the relative abundance of early-arriving IFE-derived cells specifically in the wound basal layer declined as later-arriving HF-derived cells entered the site of injury. Although laggard bulge-derived cells were typically constrained at the regenerative periphery, these cells persisted in the wound basal layer. Finally, suppressing Notch enabled IFE-derived cells to out-compete HF-derived cells. Taken together, these findings indicate that IFE-, HF- and bulge-derived cells make distinct contributions to regeneration over time. Furthermore, we speculate that extrinsic, non-genetic factors such as spatial constraint, distance from the wound, and basal versus suprabasal position may largely determine whether a cell ultimately persists.     

Early-age-dependent selective decrease of differentiation potential of hair-follicle-associated pluripotent (HAP) stem cells to beating cardiac-muscle cells

We have previously discovered nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells and have shown that they can differentiate to neurons, glia, and many other cell types. HAP stem cells can be used for nerve and spinal cord repair. We have recently shown the HAP stem cells can differentiate to beating heart-muscle cells and tissue sheets of beating heart-muscle cells. In the present study, we determined the efficiency of HAP stem cells from mouse vibrissa hair follicles of various ages to differentiate to beating heart-muscle cells. We observed that the whiskers located near the ear were more efficient to differentiate to cardiac-muscle cells compared to whiskers located near the nose. Differentiation to cardiac-muscle cells from HAP stem cells in cultured whiskers in 4-week-old mice was significantly greater than in 10-, 20-, and 40-week-old mice. There was a strong decrease in differentiation potential of HAP stem cells to cardiac-muscle cells by 10 weeks of age. In contrast, the differentiation potential of HAP stem cells to other cell types did not decrease with age. The possibility of rejuvenation of HAP stem cells to differentiate at high efficiency to cardiac-muscle cells is discussed.     

Dietary vitamin A regulates wingless-related MMTV integration site signaling to alter the hair cycle

Alopecia areata (AA) is an autoimmune hair loss disease caused by a cell-mediated immune attack of the lower portion of the cycling hair follicle. Feeding mice 3–7 times the recommended level of dietary vitamin A accelerated the progression of AA in the graft-induced C3H/HeJ mouse model of AA. In this study, we also found that dietary vitamin A, in a dose dependent manner, activated the hair follicle stem cells (SCs) to induce the development and growth phase of the hair cycle (anagen), which may have made the hair follicle more susceptible to autoimmune attack. Our purpose here is to determine the mechanism by which dietary vitamin A regulates the hair cycle. We found that vitamin A in a dose-dependent manner increased nuclear localized beta-catenin (CTNNB1; a marker of canonical wingless-type Mouse Mammary Tumor Virus integration site family (WNT) signaling) and levels of WNT7A within the hair follicle bulge in these C3H/HeJ mice. These findings suggest that feeding mice high levels of dietary vitamin A increases WNT signaling to activate hair follicle SCs.     

PRC1 Fine-tunes Gene Repression and Activation to Safeguard Skin Development and Stem Cell Specification

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Human hair-follicle associated pluripotent (hHAP) stem cells differentiate to cardiac-muscle cells

We have previously demonstrated that nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells are located in the bulge area. HAP stem cells have been previously shown to differentiate to neurons, glial cells, keratinocytes, smooth-muscle cells, melanocytes and cardiac-muscle cells in vitro. Subsequently, we demonstrated that HAP stem cells could effect nerve and spinal cord regeneration in mouse models, differentiating to Schwann cells and neurons. In previous studies, we established an efficient protocol for the differentiation of cardiac-muscle cells from mouse HAP stem cells. In the present study, we isolated the upper part of human hair follicles containing human HAP (hHAP) stem cells. The upper parts of human hair follicles were suspended in DMEM containing 10% FBS where they differentiated to cardiac-muscle cells as well as neurons, glial cells, keratinocytes and smooth-muscle cells. This method is appropriate for future use with human hair follicles to produce hHAP stem cells in sufficient quantities for future heart, nerve and spinal cord regeneration in the clinic.     

Genetic mosaic analysis indicates that the bulb region of coat hair follicles contains a resident population of several active multipotent epithelial lineage progenitors.

The hair follicle represents an excellent model system for exploring the properties of lineage-forming units in a dynamic epithelium containing multiple cell types. During its growth (anagen) phase, the proximal-distal axis of the mouse coat hair (pelage) follicle provides a historical record of all epithelial lineages generated from its resident stem cell population. An unresolved question in the field is whether the bulb region of anagen pelage follicles contains multipotential progenitors and whether their individual contribution to cellular census fluctuates over time. To address this issue, chimeric follicles were harvested in midanagen from three types of genetic mosaic mouse models. Analysis of the distribution of genotypic markers, including digital three-dimensional reconstruction of serially sectioned chimeric follicles, revealed that on average the bulb contains four or fewer active progenitors, each capable of giving rise to all six follicular epithelial fates. Moreover, analysis of mosaic pelage, as well as cultured whisker follicles provided evidence that bulb-associated progenitors can give rise to expanding descendant clones during midanagen, leading to the conclusion that the bulb contains dormant or symmetrically dividing stem cells. This latter feature resembles the behavior of hematopoietic stem cells after bone marrow transplantation, and raises the question of whether this property may be shared by stem cells in other self-renewing epithelia.     

干细胞巢研究进展

干细胞巢即干细胞周围的微环境构成,一般包括干细胞的相邻细胞、粘附分子及基质等,但不同的干细胞有不同的巢结构。干细胞巢通过不同信号途径调控着干细胞的行为,使干细胞的自我更新和分化处于平衡状态。根据近年来有关干细胞巢的研究,本文从果蝇生殖系干细胞巢、哺乳动物造血干细胞巢、肠干细胞巢、毛囊表皮干细胞巢和神经干细胞巢等五个系统分别综述了干细胞巢的构成及其对干细胞的调节作用,探讨了干细胞巢作用于干细胞的内在机制。     

Activity of mesenchymal stem cells in therapies for chronic skin wound healing

Chronic or non-healing skin wounds present an ongoing challenge in advanced wound care, particularly as the number of patients increases while technology aimed at stimulating wound healing in these cases remains inefficient. Mesenchymal stem cells (MSCs) have proved to be an attractive cell type for various cell therapies due to their ability to differentiate into various cell lineages, multiple donor tissue types, and relative resilience in ex-vivo expansion, as well as immunomodulatory effects during transplants. More recently, these cells have been targeted for use in strategies to improve chronic wound healing in patients with diabetic ulcers or other stasis wounds. Here, we outline several mechanisms by which MSCs can improve healing outcomes in these cases, including reducing tissue inflammation, inducing angiogenesis in the wound bed, and reducing scarring following the repair process. Approaches to extend MSC life span in implant sites are also examined.