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1.
Mol Med Rep ; 23(5)2021 May.
Artigo em Inglês | MEDLINE | ID: mdl-33760132

RESUMO

Hair loss, including alopecia, is a common and distressing problem for men and women, and as a result, there is considerable interest in developing treatments that can prevent or reverse hair loss. Dermal papillae closely interact with epidermal cells and play a key role during hair follicle induction and hair morphogenesis. As dermal papilla cells (DPCs) lose their hair­inducing ability in monolayer cultures in vitro, it is difficult to obtain de novo hair follicle structures following DPC transplantation in vivo. The present study aimed to explore culture conditions to maintain DPC characteristics using conditioned media (CM) from the supernatant of cultured HaCaT keratinocyte cells supplemented with other components. Initially, it was observed that during passaging of in vitro monolayer DPC cultures, the Wnt/ß­catenin pathway was repressed, while the TGF­ß/Smad pathway was activated, and that HaCaT cells cultivated in 1% fetal bovine serum had higher levels of expression of Wnt3a and Wnt10b compared with normal keratinocytes. Culturing of high­passage (P7) DPCs in CM from HaCaT cells (HaCaT­CM) actively stimulated cell proliferation and maintained Sox2 and Versican expression levels. Supplementation of HaCaT­CM with SB431542 (SB, a TGF­ß receptor inhibitor), CHIR99021, (CHIR, a GSK3α/ß inhibitor and activator of Wnt signaling) and platelet­derived growth factor (PDGF)­AA further increased the expression levels of Sox2, Versican and alkaline phosphatase (ALP) in P7 DPCs. Three­dimensional culture of P7 DPCs using hanging drop cultures in HaCaT­CM supplemented with SB, CHIR and PDGF­AA resulted in larger cell aggregates and a further significant upregulation of Sox2, ALP and Versican expression levels. Taken together, these findings demonstrated that HaCaT­CM supplemented with SB, CHIR and PDGF­AA may preserve the hair­inducing ability of high­passage DPCs and may therefore be useful in reconstructing new hair follicles in vivo.


Assuntos
Alopecia/genética , Desdiferenciação Celular/efeitos dos fármacos , Derme/crescimento & desenvolvimento , Fator de Crescimento Derivado de Plaquetas/genética , Alopecia/tratamento farmacológico , Alopecia/patologia , Benzamidas/farmacologia , Técnicas de Cultura de Células , Meios de Cultivo Condicionados , Derme/citologia , Dioxóis/farmacologia , Cabelo/crescimento & desenvolvimento , Cabelo/metabolismo , Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Humanos , Queratinócitos/citologia , Queratinócitos/metabolismo , Piridinas/farmacologia , Pirimidinas/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Via de Sinalização Wnt/efeitos dos fármacos
2.
Methods Mol Biol ; 2269: 175-201, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33687680

RESUMO

Bench-to-bedside axis of therapeutic product development is currently being oriented towards minimum invasiveness on both ends-not only clinical application but harvesting of the starting biological material as well. This is particularly relevant for Advanced Therapy Medicinal Products and their specific legislative requirements, even more so in skin regeneration. It is precisely the skin equivalents and grafts that benefit from the minimum-to-noninvasive approach to a noteworthy extent, taking in account the sensitive nature of both skin harvesting and grafting.This chapter includes protocols for two separate steps of generating skin equivalent from the cells cultured from hair follicle outer root sheath. The first step is a non-pigmented epidermal equivalent generated from human keratinocytes from the outer root sheath named non-pigmented epidermal graft. The second step consists of co-cultivating human keratinocytes and human melanocytes from the outer root sheath, hereby producing a pigmented epidermal graft.


Assuntos
Derme/metabolismo , Fibroblastos/metabolismo , Folículo Piloso/metabolismo , Queratinócitos/metabolismo , Melanócitos/metabolismo , Engenharia Tecidual , Técnicas de Cocultura , Derme/citologia , Fibroblastos/citologia , Folículo Piloso/citologia , Humanos , Queratinócitos/citologia , Melanócitos/citologia
3.
Gene ; 770: 145339, 2021 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-33333220

RESUMO

Hair follicle (HF) development is characterized by periodic growth cycles regulated by numerous factors. We previously showed that SMAD2 might be involved in the HF growth cycle in Angora rabbits. However, its extra role in the HF growth and development remains obscure. In this study, we cloned the complete coding sequence (CDS) of the Angora rabbit SMAD2 gene. Within SMAD2 CDS, we identified the open reading frame (ORF) had a length of 1314 bp and encoding 437 amino acids. Bioinformatics analyses revealed that the SMAD2 protein is unstable and hydrophilic, and predominatelylocalizesin the cell nucleus. We identified that SMAD2 expression was elevated in the telogen phase of the during HF cycle. The knockdown and overexpression of SMAD2 could regulate HF growth and development related genes, such as WNT2, FGF2, and LEF1.Furthermore, SMAD2 may upregulate TGF-ß signaling pathway-related genes, including TFDP1, E2F4, and RBL1. In conclusion, our results indicate that SMAD2 plays a vital role in HF development by regulating the TGF-ß signaling pathway.


Assuntos
Folículo Piloso/metabolismo , Proteína Smad2/metabolismo , Animais , Fator 2 de Crescimento de Fibroblastos/metabolismo , Folículo Piloso/citologia , Masculino , Coelhos , Proteína p107 Retinoblastoma-Like/metabolismo , Proteína Wnt2/metabolismo
4.
Nat Commun ; 11(1): 5114, 2020 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-33037205

RESUMO

Tissue growth in the adult is an orchestrated process that often requires biological clocks to time stem cell and progenitor activity. Here, we employed the hair follicle, which cycles between growth and regression in a timely-restricted mode, to show that some components of the hair cycle clock reside within the mesenchymal niche of the hair follicle, the dermal papilla (DP), and both Fgf and Wnt signaling pathways interact within the DP to regulate the expression of these components that include Wnt agonists (Rspondins) and antagonists (Dkk2 and Notum). The levels of Wnt agonists and antagonists in the DP are progressively reduced and elevated during the growth phase, respectively. Consequently, Wnt signaling activity in the overlying epithelial progenitor cells decreases, resulting in the induction of the regression phase. Remarkably, DP properties allow Wnt activity in the DP to persist despite the Wnt-inhibiting milieu and consequently synchronize the induction and progression of the regression phase. This study provides insight into the importance of signaling crosstalk in coupling progenitors and their niche to regulate tissue growth.


Assuntos
Fatores de Crescimento de Fibroblastos/metabolismo , Regulação da Expressão Gênica , Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Via de Sinalização Wnt/fisiologia , Animais , Esterases/genética , Esterases/metabolismo , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/genética , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos Knockout , Camundongos Mutantes , Receptores de Fatores de Crescimento de Fibroblastos/genética , Receptores de Fatores de Crescimento de Fibroblastos/metabolismo , Pele/citologia , Trombospondinas/genética , Trombospondinas/metabolismo
6.
Nature ; 582(7812): 399-404, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32494013

RESUMO

The skin is a multilayered organ, equipped with appendages (that is, follicles and glands), that is critical for regulating body temperature and the retention of bodily fluids, guarding against external stresses and mediating the sensation of touch and pain1,2. Reconstructing appendage-bearing skin in cultures and in bioengineered grafts is a biomedical challenge that has yet to be met3-9. Here we report an organoid culture system that generates complex skin from human pluripotent stem cells. We use stepwise modulation of the transforming growth factor ß (TGFß) and fibroblast growth factor (FGF) signalling pathways to co-induce cranial epithelial cells and neural crest cells within a spherical cell aggregate. During an incubation period of 4-5 months, we observe the emergence of a cyst-like skin organoid composed of stratified epidermis, fat-rich dermis and pigmented hair follicles that are equipped with sebaceous glands. A network of sensory neurons and Schwann cells form nerve-like bundles that target Merkel cells in organoid hair follicles, mimicking the neural circuitry associated with human touch. Single-cell RNA sequencing and direct comparison to fetal specimens suggest that the skin organoids are equivalent to the facial skin of human fetuses in the second trimester of development. Moreover, we show that skin organoids form planar hair-bearing skin when grafted onto nude mice. Together, our results demonstrate that nearly complete skin can self-assemble in vitro and be used to reconstitute skin in vivo. We anticipate that our skin organoids will provide a foundation for future studies of human skin development, disease modelling and reconstructive surgery.


Assuntos
Cabelo/citologia , Cabelo/crescimento & desenvolvimento , Organoides/citologia , Células-Tronco Pluripotentes/citologia , Pele/citologia , Animais , Ectoderma/citologia , Feminino , Cabelo/transplante , Cor de Cabelo , Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Folículo Piloso/inervação , Folículo Piloso/transplante , Cabeça , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , Organoides/crescimento & desenvolvimento , Organoides/inervação , Organoides/transplante , RNA-Seq , Análise de Célula Única , Pele/crescimento & desenvolvimento , Pele/inervação , Transplante de Pele
7.
Nat Cell Biol ; 22(6): 640-650, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32393888

RESUMO

Tissue homeostasis and regeneration rely on resident stem cells (SCs), whose behaviour is regulated through niche-dependent crosstalk. The mechanisms underlying SC identity are still unfolding. Here, using spatiotemporal gene ablation in murine hair follicles, we uncover a critical role for the transcription factors (TFs) nuclear factor IB (NFIB) and IX (NFIX) in maintaining SC identity. Without NFI TFs, SCs lose their hair-regenerating capability, and produce skin bearing striking resemblance to irreversible human alopecia, which also displays reduced NFIs. Through single-cell transcriptomics, ATAC-Seq and ChIP-Seq profiling, we expose a key role for NFIB and NFIX in governing super-enhancer maintenance of the key hair follicle SC-specific TF genes. When NFIB and NFIX are genetically removed, the stemness epigenetic landscape is lost. Super-enhancers driving SC identity are decommissioned, while unwanted lineages are de-repressed ectopically. Together, our findings expose NFIB and NFIX as crucial rheostats of tissue homeostasis, functioning to safeguard the SC epigenome from a breach in lineage confinement that otherwise triggers irreversible tissue degeneration.


Assuntos
Alopecia/patologia , Diferenciação Celular , Cromatina/metabolismo , Folículo Piloso/citologia , Fatores de Transcrição NFI/fisiologia , Células-Tronco/citologia , Alopecia/genética , Alopecia/metabolismo , Animais , Células Cultivadas , Cromatina/genética , Feminino , Folículo Piloso/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Regeneração , Células-Tronco/metabolismo
8.
PLoS One ; 15(4): e0231376, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32298297

RESUMO

Hair follicle stem cells (HFSCs) have been shown to be essential in the development and regeneration of hair follicles (HFs). The Inner Mongolia Cashmere goat (Capra hircus) has two types of HFs, primary and secondary, with cashmere being produced from the secondary hair follicle. To identify the genes associated with cashmere growth, transcriptome profiling of anagen and telogen secondary HFSCs was performed by RNA-Seq. The RNA-Seq analysis generated over 58 million clean reads from each group, with 2717 differentially expressed genes (DEGs) detected between anagen and telogen, including 1500 upregulated and 1217 downregulated DEGs. A large number of DEGs were predominantly associated with cell part, cellular process, binding, biological regulation and organelle. In addition, the PI3K-Akt, MAPK, Ras and Rap1 signaling pathways may be involved in the growth of HFSCs cultured in vitro. The RNA-Seq results showed that the well-defined HFSC signature genes and cell cycle-associated genes showed no significant differences between anagen and telogen HFSCs, indicating a relatively quiescent cellular state of the HFSCs cultured in vitro. These results are useful for future studies of complex molecular mechanisms of hair follicle cycling in cashmere goats.


Assuntos
Cabras/genética , Folículo Piloso/citologia , Células-Tronco Mesenquimais/metabolismo , Transcriptoma , Animais , Células Cultivadas , Perfilação da Expressão Gênica/métodos , Perfilação da Expressão Gênica/normas , Folículo Piloso/metabolismo , RNA-Seq/métodos , Lã/citologia , Lã/metabolismo
9.
Life Sci ; 252: 117667, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32304761

RESUMO

AIMS: Pantothenic acid (PA) has been applied to treat alopecia, but the underlying mechanism is still unclear. Our study aims to explore the underlying mechanism of PA in regulating hair follicle (HF) growth. MAIN METHODS: Mink HFs and dermal papilla (DP) cells were isolated and cultured in vitro. HFs and DP cells were treated with 0, 10, 20, 40 µg/ml PA. The effect of PA on HF growth, DP cell proliferation, cell cycle distribution, cell migration, and insulin-like growth factor-1 (IGF-1) and vascular endothelial growth factor (VEGF) expressions in DP cells was measured. Moreover, the effect of PA on inhibitor of DNA binding 3 (ID3)/Notch signaling pathway was analyzed. Subsequently, ID3 was silenced to validate whether ID3/Notch signaling pathway was involved in regulating DP cell proliferation by PA. KEY FINDINGS: Both 20 µg/ml and 40 µg/ml PA promoted HF growth, G1/S transition of DP cells and IGF-1 and VEGF expressions in DP cells, while only 20 µg/ml PA promoted cell viability and the migration of DP cells. Thus 20 µg/ml PA was chosen for the following experiments. PA treatment was found to up-regulate ID3 expression but down-regulate Notch receptor 1 (Notch1) and Notch signaling targets expressions. Furthermore, ID3 knockdown reversed PA-induced cell proliferation and inhibition of Notch1 and Notch signaling targets expressions, indicating that PA-induced DP cell proliferation and inhibition of Notch signaling were mediated via up-regulation of ID3. SIGNIFICANCE: This study provides an underlying mechanism related to the effect of PA on stimulating DP cell proliferation.


Assuntos
Proliferação de Células/efeitos dos fármacos , Derme/efeitos dos fármacos , Folículo Piloso/efeitos dos fármacos , Ácido Pantotênico/farmacologia , Animais , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Derme/citologia , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Folículo Piloso/citologia , Proteínas Inibidoras de Diferenciação/metabolismo , Masculino , Vison , Ácido Pantotênico/administração & dosagem , Receptores Notch/metabolismo , Transdução de Sinais/efeitos dos fármacos
10.
An Bras Dermatol ; 95(3): 278-282, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32299738

RESUMO

BACKGROUND: The hair follicle is a unique structure, one of the most dynamic structures in mammalians, which can reproduce in every new cycle all the mechanism involved in its fetal development. Although a lot of research has been made about the human hair follicle much less has been discovered about the importance of the cytokeratins (CKs) in its development. OBJECTIVE: Study the immunohistochemical pattern of epithelial CKs during human hair follicle development. METHODS: We performed an immunohistochemical study using fresh post-mortem skin biopsies of human fetuses between 4 and 25 weeks of gestational age to study the expression of cytokeratins (CKs): CK1, CK10, CK13, CK14, CK16 and CK20 during human hair follicle fetal development. STUDY LIMITATIONS: Restrospective study with a good number of makers but with a small population. RESULTS/CONCLUSION: We found that, the CKs were expressed in an intermediate time during follicular development. The epithelial CKs (CK1, CK14, CK10, CK13) and the epithelial CKs with a proliferative character such as CK16 were expressed first, as markers of cellular maturation and follicular keratinization. At a later phase, CK20 was expressed in more developed primitive hair follicles as previously discussed in literature.


Assuntos
Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Queratinas Específicas do Cabelo/análise , Fatores Etários , Anticorpos Monoclonais/análise , Desenvolvimento Fetal , Idade Gestacional , Humanos , Imuno-Histoquímica , Estudos Retrospectivos
11.
Int J Mol Sci ; 21(7)2020 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-32218218

RESUMO

Increasing cashmere yield is one of the important goals of cashmere goat breeding. To achieve this goal, we screened the key genes that can improve cashmere performance. In this study, we used the RNA raw datasets of the skin and dermal papilla cells of secondary hair follicle (SHF-DPCs) samples of hair follicle (HF) anagen and telogen of Albas cashmere goats and identified a set of significant differentially expressed genes (DEGs). To explore potential associations between gene sets and SHF growth features and to identify candidate genes, we detected functional enrichment and constructed protein-protein interaction (PPI) networks. Through comprehensive analysis, we selected Thymosin ß4 (Tß4), Rho GTPase activating protein 6 (ARHGAP6), ADAM metallopeptidase with thrombospondin type 1 motif 15, (ADAMTS15), Chordin (CHRD), and SPARC (Osteonectin), cwcv and kazal-like domains proteoglycan 1 (SPOCK1) as candidate genes. Gene set enrichment analysis (GSEA) for these genes revealed Tß4 and ARHGAP6 have a close association with the growth and development of SHF-DPCs. However, the expression of Tß4 in the anagen was higher than that in the telogen, so we finally chose Tß4 as the ultimate research object. Overexpressing Tß4 promoted and silencing Tß4 inhibited the proliferation of SHF-DPCs. These findings suggest that Tß4 can promote the growth and development of SHF-DPCs and indicate that this molecule may be a valuable target for increasing cashmere production.


Assuntos
Proliferação de Células , Folículo Piloso/metabolismo , Timosina/metabolismo , Animais , Células Cultivadas , Ativação Enzimática , Perfilação da Expressão Gênica , Cabras , Folículo Piloso/citologia , Folículo Piloso/crescimento & desenvolvimento , Timosina/genética
12.
Int J Mol Sci ; 21(3)2020 Jan 25.
Artigo em Inglês | MEDLINE | ID: mdl-31991762

RESUMO

Despite advances in medical treatments, the proportion of the population suffering from alopecia is increasing, thereby creating a need for new treatments to control hair loss and prevent balding. Human hair follicle dermal papilla cells (hDPCs), a type of specialized fibroblast in the hair bulb, play an essential role in controlling hair growth and in conditions like androgenic alopecia. This study aimed to evaluate the intensity-dependent effect of extremely low-frequency electromagnetic fields (ELF-EMFs) on the expression of anagen-related molecules in hDPCs in vitro. We examined the effect of ELF-EMF on hDPCs to determine whether activation of the GSK-3ß/ERK/Akt signaling pathway improved hDPC activation and proliferation; hDPCs were exposed to ELF-EMFs at a frequency of 70 Hz and at intensities ranging from 5 to 100 G, over four days. Various PEMF intensities significantly increased the expression of anagen-related molecules, including collagen IV, laminin, ALP, and versican. In particular, an intensity of 10 G is most potent for promoting the proliferation of hDPC and expression of anagen-related molecules. Moreover, 10 G ELF-EMF significantly increased ß-catenin and Wnt3α expression and GSK-3ß/ERK/Akt phosphorylation. Our results confirmed that ELF-EMFs enhance hDPC activation and proliferation via the GSK-3ß/ERK/Akt signaling pathway, suggesting a potential treatment strategy for alopecia.


Assuntos
Campos Eletromagnéticos , Regulação da Expressão Gênica/efeitos da radiação , Glicogênio Sintase Quinase 3 beta/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos da radiação , Biomarcadores , Proliferação de Células , Células Cultivadas , Derme/citologia , MAP Quinases Reguladas por Sinal Extracelular , Folículo Piloso/citologia , Folículo Piloso/metabolismo , Humanos , Fosforilação , Via de Sinalização Wnt/efeitos da radiação
13.
Int J Mol Med ; 45(2): 556-568, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31894311

RESUMO

Alopecia is a common and distressing condition, and developing new therapeutic agents to prevent hair loss is important. Human umbilical cord blood­derived mesenchymal stem cells (hUCB­MSCs) have been studied intensively in regenerative medicine. However, the therapeutic potential of these cells against hair loss and hair organ damage remains unclear, and the effects of hUCB­MSC transplantation on hair loss require evaluation. The current study aimed to investigate the effects of hUCB­MSCs on hair regression in vivo and restoration of anagen conduction on hair growth in vitro. The effects of hUCB­MSCs were explored in mouse catagen induction models using a topical treatment of 0.1% dexamethasone to induce hair regression. Dexamethasone was also used to simulate a stress environment in vitro. The results demonstrated that hUCB­MSCs significantly prevented hair regression induced by dexamethasone topical stimulation in vivo. Additionally, hUCB­MSCs significantly increased the proliferation of human dermal papilla cells (hDPCs) and HaCaT cells, which are key constituent cells of the hair follicle. Stimulation of vascular endothelial growth factor secretion and decreased expression of DKK­1 by hUCB­MSCs were also observed in hDPCs. Restoration of cell viability by hUCB­MSCs suggested that these cells exerted a protective effect on glucocorticoid stress­associated hair loss. In addition, anti­apoptotic effects and regulation of the autophagic flux recovery were observed in HaCaT cells. The results of the present study indicated that hUCB­MSCs may have the capacity to protect hair follicular dermal papilla cells and keratinocytes, thus preventing hair loss. Additionally, the protective effects of hUCB­MSCs may be resistant to dysregulation of autophagy under harmful stress.


Assuntos
Anti-Inflamatórios/efeitos adversos , Dexametasona/efeitos adversos , Folículo Piloso/citologia , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Células Cultivadas , Feminino , Sangue Fetal/citologia , Cabelo/citologia , Cabelo/efeitos dos fármacos , Cabelo/crescimento & desenvolvimento , Cabelo/ultraestrutura , Folículo Piloso/efeitos dos fármacos , Folículo Piloso/ultraestrutura , Humanos , Camundongos Endogâmicos C57BL
14.
Genome ; 63(3): 179-187, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31917611

RESUMO

Hair follicle (HF) growth and cycling is a complex biological process that occurs in most mammals. As HF growth and cycling directly impacts rabbit wool yield, it is important to better understand the potential regulation pattern of HF development. Our previous study demonstrated that HTATIP2 may participate in regulating rabbit HF cycles, but the molecular mechanism of HTATIP2 remained unclear. In this study, the coding sequence of the HTATIP2 gene in Angora rabbit was cloned. The length of the coding region sequence was 840 bp, which could code 279 amino acids, and exhibited high homology in different mammals. Bioinformatics analyses indicated that the HTATIP2 protein is stable, hydrophilic, located around the cytoplasm, and has a putative signal peptide. Moreover, we verified that HTATIP2 is highly expressed during catagen and telogen of the HF cycle. The overexpression vector was constructed and siRNAs were designed. Overexpression and knockdown of HTATIP2 appeared to regulate JAK-STAT pathway genes, such as BCL2, CCND1, c-Myc, and STAT2. It is therefore likely that HTATIP2 promotes cell apoptosis and inhibits cell proliferation. Our results indicate that HTATIP2 is highly expressed during catagen and telogen and may play an important role in JAK-STAT signaling. This study provides a theoretical foundation for investigating HTATIP2 in biological processes.


Assuntos
Acetiltransferases/genética , Acetiltransferases/metabolismo , Clonagem Molecular/métodos , Folículo Piloso/citologia , Animais , Apoptose , Linhagem Celular , Proliferação de Células , Folículo Piloso/metabolismo , Coelhos , Análise de Sequência de DNA/veterinária , Transdução de Sinais , Regulação para Cima ,
15.
Differentiation ; 111: 70-78, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31715508

RESUMO

Fibroblast growth factor (FGF2) is reported to affect the proliferation, differentiation, and survival abilities of stem cells. In this study, we hypothesize that FGF2 might promote the differentiation of hair follicle stem cell (HFSCs) into endothelial cells (ECs), in a manner dependent on STAT5 activation. We first treated human HFSCs with recombinant human FGF2 to determine the involvement of FGF2 in the differentiation of HFSCs. Then the expression of EC-specific markers including von Willebrand factor (vWF), VE-cadherin, CD31, FLT-1, KDR and Tie2 was evaluated using immunofluorescence and flow cytometry, while the expression of HFSC-specific markers such as K15, K19, Lgr5, Sox9 and Lhx2 was determined by flow cytometry. Next, in vitro tube formation was performed to confirm the function of FGF2, and low-density lipoprotein (LDL) uptake by ECs and HFSCs was studied by Dil-acetylated LDL assay. In addition, we transduced FGF2-treated HFSCs with constitutive-active or dominant-negative STAT5A adenovirus vectors. FGF2 up-regulated the expression of EC-specific markers, and promoted the differentiation of HFSCs into ECs, tube formation and LDL uptake. The phosphorylated STAT5 was translocated into the nucleus of HFSCs after FGF2 treatment, but this translocation was blocked by the dominant-negative STAT5A mutant. FGF2 increased the differentiation potential through the activation of STAT5 in vivo. Taken together, we find that FGF2 promotes the differentiation of HFSCs into ECs via activated STAT5, which gives a new perspective on the role of FGF2 in the development of ischemic vascular disease.


Assuntos
Diferenciação Celular , Células Endoteliais/citologia , Fator 2 de Crescimento de Fibroblastos/metabolismo , Folículo Piloso/citologia , Fator de Transcrição STAT5/metabolismo , Células-Tronco/citologia , Animais , Células Cultivadas , Células Endoteliais/metabolismo , Fator 2 de Crescimento de Fibroblastos/genética , Folículo Piloso/metabolismo , Humanos , Fator de Transcrição STAT5/genética , Células-Tronco/metabolismo
16.
Clin Exp Dermatol ; 45(4): 417-425, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31580512

RESUMO

BACKGROUND: Identification of human basal cell carcinoma (BCC) cancer stem cells and cellular hierarchy inherently implies the presence of differentiation. By conventional histological analysis, BCC demonstrates tumour nodules that appear relatively homogeneous. AIM: As BCCs arise from hair follicle (HF) keratinocytes, we sought to define the pattern of HF differentiation. METHODS: BCC, squamous cell carcinoma (SCC) and normal skin tissues were analysed using a microarray chip. The expression of individual keratins, regulatory pathways and proliferative states were analysed using reverse transcription-PCR and immunofluorescence microscopy. RESULTS: Microarray analysis of BCC, SCC and normal hair-bearing skin revealed that BCCs express a wide range of HF genes, including HF- specific keratins. BCC demonstrated outer (KRT5, KRT514, KRT516, KRT517 and KRT519) and inner (KRT25, KRT27, KRT28, KRT32, KRT35, KRT71, KRT75 and KRT85) root sheath differentiation, but not hair shaft differentiation. As in the HF, differentiation-specific keratins in BCC keratinocytes correlated with a reduced proliferative index and regulatory pathway activation despite the oncogenic drive towards tumour growth. Our findings show the close correlation between HF and BCC keratinocyte differentiation. CONCLUSION: This work has defined the differentiation pattern within BCCs, enabling development of targeted therapies that promote differentiation and result in BCC cancer stem cell exhaustion.


Assuntos
Carcinoma Basocelular/metabolismo , Folículo Piloso/metabolismo , Queratinas Específicas do Cabelo/metabolismo , Neoplasias Cutâneas/metabolismo , Carcinoma Basocelular/patologia , Diferenciação Celular , Folículo Piloso/citologia , Humanos , Análise em Microsséries , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias Cutâneas/patologia
17.
J Invest Dermatol ; 140(4): 764-773.e4, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31676413

RESUMO

The epidermis and its appendage, the hair follicle, represent an elegant developmental system in which cells are replenished with regularity because of controlled proliferation, lineage specification, and terminal differentiation. Although transcriptome data exists for human epidermal and dermal cells, the hair follicle remains poorly characterized. Through single-cell resolution profiling of the epidermis and anagen hair follicle, we characterized the anatomical, transcriptional, functional, and pathological profiles of distinct epidermal, hair follicle, and hair follicle-associated cell subpopulations including melanocytes, endothelial cells, and immune cells. We additionally traced the differentiation trajectory of interfollicular and matrix cell progenitors and explored the association of specific cell subpopulations to known molecular signatures of common skin conditions. These data simultaneously corroborate prior murine and human studies while offering new insights into epidermal and hair follicle differentiation and pathogenesis.


Assuntos
Células Endoteliais/citologia , Folículo Piloso/citologia , Pele/citologia , Células-Tronco/citologia , Animais , Diferenciação Celular , Humanos
18.
Iran Biomed J ; 24(2): 99-109, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31677605

RESUMO

Background: The recent improvements in wound healing have led to new strategies in regenerative medicine. Burn wound healing is an important issue in skin regeneration and has multiple indications for stem cell therapy. Hair follicle stem cells (HFSCs) are a highly promising source of stem cells for healing use, as these cells are accessible, active and pluripotent adult stem cells. Methods: HFSCs of the rat whisker were isolated, cultured, and labeled with DiI. Flow cytometry method was used to detect special markers of HFSCs. Deep partial-thickness burn wound was created, and labeled HFSCs were injected around the wound bed. Wound closure was recorded via digital photographs. The inflicted rats were sacrificed at 3, 7, or 14 days post burn and used for subsequent histological and tensiometry analysis. Results: Our results indicated that HFSCs were positive for Nestin and CD34 markers, but negative for Kr15. Morphological and histological photographs revealed that wound closure rate was accelerated in stem cell-treated group compared with other groups. In addition, faster re-epithelialization and collagen deposition were observed. The immunohistochemical analysis suggested that CD31 expression and vascular density enhanced in the stem cell-treated group. Further, tissue tensile strength increased in HFSCs-treated rats in comparison to the control group. Conclusion: The present study demonstrates that HFSC could accelerate burn wound healing as well as tensile strength in rats.


Assuntos
Queimaduras/terapia , Terapia Baseada em Transplante de Células e Tecidos/métodos , Folículo Piloso/citologia , Células-Tronco Pluripotentes/transplante , Cicatrização/fisiologia , Animais , Antígenos CD34/metabolismo , Colágeno/metabolismo , Masculino , Nestina/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Ratos , Ratos Wistar , Regeneração/fisiologia , Pele/patologia , Lesões dos Tecidos Moles/terapia , Resistência à Tração
19.
Clin Exp Dermatol ; 45(3): 309-317, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31556145

RESUMO

BACKGROUND: Consistent with cancer stem cell driven pattern of growth, human basal cell carcinomas (BCCs) demonstrate differentiation along hair follicle (HF) lineages. AIM: To define the pattern of differentiation and therapeutic targets that promote BCC differentiation and therefore BCC cancer stem cell exhaustion. METHODS: An alkaline phosphatase substrate kit was used to determine dermal papilla cells within the BCC stroma. Autonomous HF cycle-dependent gene expression was identified by analysis of the human homologues of a murine gene set (total 2289 genes) that is differentially expressed in hair cycle phases. The findings were validated by quantitative real-time PCR and immunofluorescence, as well as in vitro transforming growth factor (TGF)-ß2 stimulation of BCC cancer stem cell colonies. RESULTS: As in the HF, keratin expression in the inner root sheath and matrix in BCC correlated with proliferative index and was tightly regulated, despite the absence of dermal papilla cells. Cross-species microarray analysis comparing human BCC and murine synchronous HF growth cycle datasets revealed 74% concordance with telogen differentiation compared with anagen (23%, P < 0.01) and catagen (49%; P < 0.01). Incomplete anagen differentiation within BCC was characterized by reduced expression of the anagen master regulator DLX3 (-5.5-fold), and increased expression of telogen-associated genes: AEBP1 (2.2-fold), DEFB8 (35.3-fold), MMP3 (106.0-fold) and MMP12 (12.9-fold). Restoration of dermal papilla signals by in vitro addition of TGF-ß2 enhanced anagen differentiation. CONCLUSION: Our findings show that BCC cells differentiate along HF lineages and may be susceptible to exogenous HF cycle modulators.


Assuntos
Carcinoma Basocelular/patologia , Diferenciação Celular/fisiologia , Folículo Piloso/citologia , Neoplasias Cutâneas/patologia , Animais , Carcinoma Basocelular/fisiopatologia , Transformação Celular Neoplásica , Imunofluorescência , Expressão Gênica , Folículo Piloso/crescimento & desenvolvimento , Folículo Piloso/metabolismo , Humanos , Queratinas/metabolismo , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias Cutâneas/fisiopatologia
20.
J Invest Dermatol ; 140(3): 544-555.e9, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31494092

RESUMO

The dermis harbors distinct mesenchymal stem cell (MSC) populations, which play equally important roles as epidermal stem cells in skin homeostasis and regeneration. However, to reliably identify and directly isolate the in vivo counterpart of these cells is still challenging. Using the epidermal stem cell marker CD49f, we defined a CD49fhigh distinct mesenchymal subpopulation in the dermis. In vitro and in vivo differentiation assays, and transcriptome analysis demonstrated that CD49fhigh cells possess neural crest-like cell characteristics. Our results showed that the formation of hair follicle-like budding structure and the expressions of key genes regulating hair follicle development were induced when hair follicle epithelial cells were co-cultured with CD49fhigh cells. We also found that CD49fhigh cells activated Notch signaling in co-cultured hair follicle epithelial cells, whereas the inhibition of Notch signaling resulted in epidermal cyst-like spheres and loss of maintenance of hair follicle epithelial cell characteristics. Furthermore, we identified Itga7 and CD49f as an efficient biomarker panel for direct selection of CD49fhigh skin MSCs. Our results lead to a deeper understanding of heterogeneity and the function of MSCs in the skin and may facilitate potential translational applications of these cells.


Assuntos
Células Epiteliais/fisiologia , Folículo Piloso/citologia , Integrina alfa6/metabolismo , Células-Tronco Mesenquimais/metabolismo , Animais , Antígenos CD/metabolismo , Biomarcadores/metabolismo , Diferenciação Celular , Células Cultivadas , Técnicas de Cocultura , Feminino , Perfilação da Expressão Gênica , Cadeias alfa de Integrinas/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Modelos Animais , Cultura Primária de Células , Receptores Notch/metabolismo , Transdução de Sinais/fisiologia
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