RESUMO
For unknow reasons, the melanocyte stem cell (McSC) system fails earlier than other adult stem cell populations1, which leads to hair greying in most humans and mice2,3. Current dogma states that McSCs are reserved in an undifferentiated state in the hair follicle niche, physically segregated from differentiated progeny that migrate away following cues of regenerative stimuli4-8. Here we show that most McSCs toggle between transit-amplifying and stem cell states for both self-renewal and generation of mature progeny, a mechanism fundamentally distinct from those of other self-renewing systems. Live imaging and single-cell RNA sequencing revealed that McSCs are mobile, translocating between hair follicle stem cell and transit-amplifying compartments where they reversibly enter distinct differentiation states governed by local microenvironmental cues (for example, WNT). Long-term lineage tracing demonstrated that the McSC system is maintained by reverted McSCs rather than by reserved stem cells inherently exempt from reversible changes. During ageing, there is accumulation of stranded McSCs that do not contribute to the regeneration of melanocyte progeny. These results identify a new model whereby dedifferentiation is integral to homeostatic stem cell maintenance and suggest that modulating McSC mobility may represent a new approach for the prevention of hair greying.
Assuntos
Desdiferenciação Celular , Folículo Piloso , Melanócitos , Nicho de Células-Tronco , Células-Tronco , Animais , Humanos , Camundongos , Folículo Piloso/citologia , Melanócitos/citologia , Células-Tronco/citologia , Microambiente Celular , Linhagem da Célula , Envelhecimento , Homeostase , Cor de Cabelo/fisiologiaRESUMO
Nedd4 and Itch are E3 ubiquitin ligases that ubiquitinate similar targets in vitro and thus are thought to function similarly. T cells lacking Itch show spontaneous activation and T helper type 2 polarization. To test whether loss of Nedd4 affects T cells in the same way, we generated Nedd4(+/+) and Nedd4(-/-) fetal liver chimeras. Nedd4(-/-) T cells developed normally but proliferated less, produced less interleukin 2 and provided inadequate help to B cells. Nedd4(-/-) T cells contained more of the E3 ubiquitin ligase Cbl-b, and Nedd4 was required for polyubiquitination of Cbl-b induced by CD28 costimulation. Our data demonstrate that Nedd4 promotes the conversion of naive T cells into activated T cells. We propose that Nedd4 and Itch ubiquitinate distinct target proteins in vivo.
Assuntos
Ativação Linfocitária/imunologia , Proteínas Proto-Oncogênicas c-cbl/metabolismo , Linfócitos T/imunologia , Ubiquitina-Proteína Ligases/imunologia , Ubiquitinação/imunologia , Animais , Antígenos CD28/imunologia , Antígenos CD28/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte , Citometria de Fluxo , Immunoblotting , Imunoprecipitação , Camundongos , Camundongos Endogâmicos C57BL , Ubiquitina-Proteína Ligases Nedd4 , Proteínas Proto-Oncogênicas c-cbl/imunologia , Linfócitos T/metabolismo , Quimeras de Transplante , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Skin wound healing is a complex regenerative phenomenon that can result in hair follicle neogenesis. Skin regeneration requires significant contribution from the immune system and involves substantial remodelling of both epidermal and dermal compartments. In this viewpoint, we consider epigenetic regulation of reepithelialization, dermal restructuring and hair neogenesis. Because little is known about the epigenetic control of these events, we have drawn upon recent epigenetic mapping and functional studies of homeostatic skin maintenance, epithelial-mesenchymal transition in cancer, and new works on regenerative dermal cell lineages and the epigenetic events that may shape their conversion into myofibroblasts. Finally, we speculate on how these various healing components might converge for wound-induced hair follicle neogenesis.
Assuntos
Epigênese Genética , Cabelo/fisiologia , Homeostase/genética , Reepitelização/genética , Pele/lesões , Animais , Transição Epitelial-Mesenquimal/genética , Humanos , Miofibroblastos/fisiologiaRESUMO
Activation of epithelial stem cells and efficient recruitment of their proliferating progeny plays a critical role in cutaneous wound healing. The reepithelialized wound epidermis has a mosaic composition consisting of progeny that can be traced back both to epidermal and several types of hair follicle stem cells. The contribution of hair follicle stem cells to wound epidermis is particularly intriguing as it involves lineage identity change from follicular to epidermal. Studies from our laboratory show that hair follicle-fated bulge stem cells commit only transient amplifying epidermal progeny that participate in the initial wound re-epithelialization, but eventually are outcompeted by other epidermal clones and largely disappear after a few months. Conversely, recently described stem cell populations residing in the isthmus portion of hair follicle contribute long-lasting progeny toward wound epidermis and, arguably, give rise to new interfollicular epidermal stem cells. The role of epithelial stem cells during wound healing is not limited to regenerating stratified epidermis. By studying regenerative response in large cutaneous wounds, our laboratory uncovered that epithelial cells in the center of the wound can acquire greater morphogenetic plasticity and, together with the underlying wound dermis, can engage in an embryonic-like process of hair follicle neogenesis. Future studies should uncover the cellular and signaling basis of this remarkable adult wound regeneration phenomenon.
Assuntos
Células Epidérmicas , Células Epiteliais/citologia , Folículo Piloso/citologia , Reepitelização/fisiologia , Regeneração/fisiologia , Glândulas Sebáceas/citologia , Células-Tronco/citologia , Adulto , Medula Óssea/fisiologia , Diferenciação Celular , Linhagem da Célula , Proliferação de Células , Cicatriz/prevenção & controle , Epiderme/lesões , Epiderme/fisiologia , Células Epiteliais/fisiologia , Folículo Piloso/fisiologia , Humanos , Glândulas Sebáceas/fisiologia , Células-Tronco/fisiologia , Ferimentos Penetrantes/patologia , Ferimentos Penetrantes/reabilitaçãoRESUMO
During wound healing in adult mouse skin, hair follicles and then adipocytes regenerate. Adipocytes regenerate from myofibroblasts, a specialized contractile wound fibroblast. Here we study wound fibroblast diversity using single-cell RNA-sequencing. On analysis, wound fibroblasts group into twelve clusters. Pseudotime and RNA velocity analyses reveal that some clusters likely represent consecutive differentiation states toward a contractile phenotype, while others appear to represent distinct fibroblast lineages. One subset of fibroblasts expresses hematopoietic markers, suggesting their myeloid origin. We validate this finding using single-cell western blot and single-cell RNA-sequencing on genetically labeled myofibroblasts. Using bone marrow transplantation and Cre recombinase-based lineage tracing experiments, we rule out cell fusion events and confirm that hematopoietic lineage cells give rise to a subset of myofibroblasts and rare regenerated adipocytes. In conclusion, our study reveals that wounding induces a high degree of heterogeneity among fibroblasts and recruits highly plastic myeloid cells that contribute to adipocyte regeneration.
Assuntos
Adipócitos/citologia , Adipócitos/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Análise de Célula Única/métodos , Pele/citologia , Células-Tronco/citologia , Animais , Western Blotting , Células Cultivadas , Feminino , Masculino , Camundongos , Análise de Sequência de RNA , Células-Tronco/metabolismo , Cicatrização/fisiologiaRESUMO
Melanoma, the deadliest skin cancer, remains largely incurable at advanced stages. Currently, there is a lack of animal models that resemble human melanoma initiation and progression. Recent studies using a Tyr-CreER driven mouse model have drawn contradictory conclusions about the potential of melanocyte stem cells (McSCs) to form melanoma. Here, we employ a c-Kit-CreER-driven model that specifically targets McSCs to show that oncogenic McSCs are a bona fide source of melanoma that expand in the niche, and then establish epidermal melanomas that invade into the underlying dermis. Further, normal Wnt and Endothelin niche signals during hair anagen onset are hijacked to promote McSC malignant transformation during melanoma induction. Finally, molecular profiling reveals strong resemblance of murine McSC-derived melanoma to human melanoma in heterogeneity and gene signatures. These findings provide experimental validation of the human melanoma progression model and key insights into the transformation and heterogeneity of McSC-derived melanoma.
Assuntos
Carcinogênese/patologia , Melanócitos/patologia , Melanoma/patologia , Células-Tronco Neoplásicas/patologia , Animais , Carcinogênese/metabolismo , Transformação Celular Neoplásica/patologia , Derme/patologia , Modelos Animais de Doenças , Epiderme/patologia , Homeostase , Humanos , Melanócitos/metabolismo , Camundongos , Mutação/genética , Células-Tronco Neoplásicas/metabolismo , Fenótipo , Regiões Promotoras Genéticas/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Microambiente Tumoral , Via de Sinalização WntRESUMO
Engagement of the T cell receptor (TCR) with its cognate peptide/MHC initiates a cascade of signaling events that results in T cell activation. Limiting the extent and duration of TCR signaling ensures a tightly constrained response, protecting cells from the deleterious impact of chronic activation. In order to limit the duration of activation, T cells must adjust levels of key signaling proteins. This can be accomplished by altering protein synthesis or by changing the rate of protein degradation. Ubiquitination is a process of 'tagging' a protein with ubiquitin and is one means of initiating protein degradation. This process is activated when an E3 ubiquitin ligase mediates the transfer of ubiquitin to a target protein. Accordingly, E3 ubiquitin ligases have recently emerged as key regulators of immune cell function. This review will explore how a small group of E3 ubiquitin ligases regulate T cell responses and thus direct adaptive immunity.
Assuntos
Tolerância Imunológica/imunologia , Imunidade/imunologia , Proteínas Proto-Oncogênicas c-cbl/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Animais , Complexos Endossomais de Distribuição Requeridos para Transporte , Humanos , Ativação Linfocitária/imunologia , Modelos Imunológicos , Ubiquitina-Proteína Ligases Nedd4 , Linfócitos T/enzimologia , Linfócitos T/imunologiaRESUMO
Although regeneration through the reprogramming of one cell lineage to another occurs in fish and amphibians, it has not been observed in mammals. We discovered in the mouse that during wound healing, adipocytes regenerate from myofibroblasts, a cell type thought to be differentiated and nonadipogenic. Myofibroblast reprogramming required neogenic hair follicles, which triggered bone morphogenetic protein (BMP) signaling and then activation of adipocyte transcription factors expressed during development. Overexpression of the BMP antagonist Noggin in hair follicles or deletion of the BMP receptor in myofibroblasts prevented adipocyte formation. Adipocytes formed from human keloid fibroblasts either when treated with BMP or when placed with human hair follicles in vitro. Thus, we identify the myofibroblast as a plastic cell type that may be manipulated to treat scars in humans.
Assuntos
Adipócitos/fisiologia , Reprogramação Celular , Miofibroblastos/fisiologia , Regeneração , Cicatrização , Animais , Proteína Morfogenética Óssea 2/farmacologia , Proteína Morfogenética Óssea 4/farmacologia , Proteínas Morfogenéticas Ósseas/metabolismo , Células Cultivadas , Cicatriz/patologia , Proteínas de Ligação a DNA/metabolismo , Fibroblastos/patologia , Folículo Piloso/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Miofibroblastos/efeitos dos fármacos , Proteínas Recombinantes/farmacologia , Transdução de Sinais , Fatores de Transcrição/metabolismoRESUMO
Genetic studies suggest that the major events of human hair follicle development are similar to those in mice, but detailed analyses of this process are lacking. In mice, hair follicle placode "budding" is initiated by invagination of Wnt-induced epithelium into the underlying mesenchyme. Modification of adherens junctions (AJs) is clearly required for budding. Snail-mediated downregulation of AJ component E-cadherin is important for placode budding in mice. Beta-catenin, another AJ component, has been more difficult to study owing to its essential functions in Wnt signaling, a prerequisite for hair follicle placode induction. Here, we show that a subset of human invaginating hair placode cells expresses the stem cell marker CD133 during early morphogenesis. CD133 associates with membrane beta-catenin in early placodes, and its continued expression correlates with loss of beta-catenin and E-cadherin from the cell membrane at a time when E-cadherin transcriptional repressors Snail and Slug are not implicated. Stabilization of CD133 via anti-CD133 antibody treatment of human fetal scalp explants depresses beta-catenin and E-cadherin membrane localization. We discuss this unique correlation and suggest a hypothetical model whereby CD133 promotes morphogenesis in early hair follicle placodes through the localized removal of membrane beta-catenin proteins and subsequent AJ dissolution.
Assuntos
Antígenos CD/genética , Caderinas/metabolismo , Glicoproteínas/genética , Folículo Piloso/citologia , Folículo Piloso/fisiologia , Peptídeos/genética , beta Catenina/metabolismo , Antígeno AC133 , Animais , Antígenos CD/metabolismo , Diferenciação Celular/fisiologia , Proliferação de Células , Transição Epitelial-Mesenquimal/fisiologia , Técnicas de Introdução de Genes , Glicoproteínas/metabolismo , Folículo Piloso/crescimento & desenvolvimento , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos Mutantes , Morfogênese/fisiologia , Peptídeos/metabolismo , Couro Cabeludo/citologia , Transcriptoma , Via de Sinalização Wnt/fisiologiaRESUMO
Wound induced hair follicle neogenesis (WIHN) describes a regenerative phenomenon in adult mammalian skin, wherein fully functional hair follicles regenerate de novo in the center of large excisional wounds. Originally described in rats, rabbits, sheep, and humans in 1940-60, the WIHN phenomenon was reinvestigated in mice only recently. The process of de novo hair regeneration largely duplicates the morphological and signaling features of normal embryonic hair development. Similar to hair development, WIHN critically depends on the activation of canonical WNT signaling. However, unlike hair development, WNT activation in WIHN is dependent on Fgf9 signaling generated by the immune system's gamma delta (γδ) T cells. The cellular bases of WIHN remain to be fully characterized, however, the available evidence leaves open the possibility for a blastema-like mechanism, wherein epidermal and/or dermal wound cells undergo epigenetic reprogramming toward a more plastic, embryonic-like state. De novo hair follicles do not regenerate from preexisting hair-fated bulge stem cells. This suggests that hair neogenesis is not driven by preexisting lineage-restricted progenitors, as is the case for amputation-induced mouse digit tip regeneration, but rather may require a blastema-like mechanism. The WIHN model is characterized by several intriguing features, which await further explanation. These include: (i) minimum wound size requirement for activating neogenesis, (ii) restriction of hair neogenesis to the wound's center, (iii) imperfect patterning outcomes, both in terms of neogenic hair positioning within the wound and in terms of their orientation. Future inquires into the WIHN process, made possible by a wide array of the available skin-specific genetic tools, will undoubtedly expand our understanding of the regeneration mechanisms in adult mammals.