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Obara K, Baba K, Hamada Y, Hoffman RM. The Potential of Hair-Follicle-Associated Pluripotent (HAP) Stem Cells for Regenerative Medicine. Methods Mol Biol 2024. [PMID: 39702862 DOI: 10.1007/7651_2024_583] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2024]
Abstract
Nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells from mouse and human have been shown to differentiate into neurons, glia, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes in vitro. HAP stem cells have promoted the recovery of peripheral nerve and spinal cord injuries in mouse models by differentiating into glial fibrillary acidic protein (GFAP)-positive Schwann cells. HAP stem cells enclosed on polyvinylidene fluoride membranes (PFM) were transplanted into the severed thoracic spinal cord of nude mice. After implantation, HAP stem cells differentiated into neurons and glial cells, which effected complete reattachment of the thoracic spinal cord.HAP stem cells were implanted into the injured brain of C57BL/6J or nude mice with induced intracerebral hemorrhage (ICH). After implantation, HAP stem cells differentiated into neurons, astrocytes, oligodendrocytes, and microglia in the ICH site, and demonstrated a significant functional improvement in mice. HAP-cell-sheets implanted on wounds in diabetic db/db mice effected wound healing. The levels of inflammation in the wound was suppressed by HAP-cell-sheet implantation. These results suggest autologous HAP stem cells can be used to heal refractory diabetic ulcers. HAP stem cells can differentiate into mature beating atrial and ventricular cardiomyocytes when cultured with specific supplements and have the potential for heart regeneration. HAP stem cells are readily obtained from scalp hair follicles, they do not develop teratomas and do not lose differentiation ability when cryopreserved. These results suggest that HAP stem cells have the potential as be a better source for regenerative medicine compared to induced pluripotent stem cells (iPS) or embryonic stem (ES) cells.
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Affiliation(s)
- Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Kyoko Baba
- Department of Plastic and Aesthetic Surgery, Kitasato University School of Medicine, Sagamihara, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Robert M Hoffman
- AntiCancer, Inc., San Diego, CA, USA.
- Department of Surgery, University of California San Diego, San Diego, CA, USA.
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Wang J, Wu X, Zhang L, Wang Q, Sun X, Ji D, Li Y. miR-133a-3p and miR-145-5p co-promote goat hair follicle stem cell differentiation by regulating NANOG and SOX9 expression. Anim Biosci 2024; 37:609-621. [PMID: 37946416 PMCID: PMC10915213 DOI: 10.5713/ab.23.0348] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2023] [Revised: 10/03/2023] [Accepted: 10/20/2023] [Indexed: 11/12/2023] Open
Abstract
OBJECTIVE Hair follicle stem cells (HFSCs) differentiation is a critical physiological progress in skin hair follicle (HF) formation. Goat HFSCs differentiation is one of the essential processes of superior-quality brush hair (SQBH) synthesis. However, knowledge regarding the functions and roles of miR-133a-3p and miR-145-5p in differentiated goat HFSCs is limited. METHODS To examine the significance of chi-miR-133a-3p and chi-miR-145-5p in differentiated HFSCs, overexpression and knockdown experiments of miR-133a-3p and miR-145-5p (Mimics and Inhibitors) separately or combined were performed. NANOG, SOX9, and stem cell differentiated markers (β-catenin, C-myc, Keratin 6 [KRT6]) expression levels were detected and analyzed by using real-time quantitative polymerase chain reaction, western blotting, and immunofluorescence assays in differentiated goat HFSCs. RESULTS miR-133a-3p and miR-145-5p inhibit NANOG (a gene recognized in keeping and maintaining the totipotency of embryonic stem cells) expression and promote SOX9 (an important stem cell transcription factor) expression in differentiated stem cells. Functional studies showed that miR-133a-3p and miR-145-5p individually or together overexpression can facilitate goat HFSCs differentiation, whereas suppressing miR-133a-3p and miR-145-5p or both inhibiting can inhibit goat HFSCs differentiation. CONCLUSION These findings could more completely explain the modulatory function of miR-133a-3p and miR-145-5p in goat HFSCs growth, which also provide more understandings for further investigating goat hair follicle development.
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Affiliation(s)
- Jian Wang
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
- International Joint Research Laboratory in Universities of Jiangsu Province of China for Domestic Animal Germplasm Resources and Genetic Improvement, Yangzhou University, Yangzhou 225009,
China
| | - Xi Wu
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
| | - Liuming Zhang
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
| | - Qiang Wang
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
| | - Xiaomei Sun
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
| | - Dejun Ji
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
| | - Yongjun Li
- Key Laboratory of Animal Genetics & Molecular Breeding of Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009,
China
- International Joint Research Laboratory in Universities of Jiangsu Province of China for Domestic Animal Germplasm Resources and Genetic Improvement, Yangzhou University, Yangzhou 225009,
China
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Li X, Ma Y, Li G, Jin G, Xu L, Li Y, Wei P, Zhang L. Leprosy: treatment, prevention, immune response and gene function. Front Immunol 2024; 15:1298749. [PMID: 38440733 PMCID: PMC10909994 DOI: 10.3389/fimmu.2024.1298749] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2023] [Accepted: 02/05/2024] [Indexed: 03/06/2024] Open
Abstract
Since the leprosy cases have fallen dramatically, the incidence of leprosy has remained stable over the past years, indicating that multidrug therapy seems unable to eradicate leprosy. More seriously, the emergence of rifampicin-resistant strains also affects the effectiveness of treatment. Immunoprophylaxis was mainly carried out through vaccination with the BCG but also included vaccines such as LepVax and MiP. Meanwhile, it is well known that the infection and pathogenesis largely depend on the host's genetic background and immunity, with the onset of the disease being genetically regulated. The immune process heavily influences the clinical course of the disease. However, the impact of immune processes and genetic regulation of leprosy on pathogenesis and immunological levels is largely unknown. Therefore, we summarize the latest research progress in leprosy treatment, prevention, immunity and gene function. The comprehensive research in these areas will help elucidate the pathogenesis of leprosy and provide a basis for developing leprosy elimination strategies.
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Affiliation(s)
- Xiang Li
- Department of Epidemiology and Health Statistics, School of Public Health, Southeast University, Nanjing, China
| | - Yun Ma
- Chronic Infectious Disease Control Section, Nantong Center for Disease Control and Prevention, Nantong, China
| | - Guoli Li
- Department of Chronic Infectious Disease Control and Prevention, Jiangsu Provincial Center for Disease Control and Prevention, Nanjing, China
| | - Guangjie Jin
- Department of Chronic Infectious Disease Control and Prevention, Jiangsu Provincial Center for Disease Control and Prevention, Nanjing, China
| | - Li Xu
- Department of Epidemiology and Health Statistics, School of Public Health, Southeast University, Nanjing, China
| | - Yunhui Li
- Department of Epidemiology and Health Statistics, School of Public Health, Southeast University, Nanjing, China
| | - Pingmin Wei
- Department of Epidemiology and Health Statistics, School of Public Health, Southeast University, Nanjing, China
| | - Lianhua Zhang
- Department of Epidemiology and Health Statistics, School of Public Health, Southeast University, Nanjing, China
- Department of Chronic Infectious Disease Control and Prevention, Jiangsu Provincial Center for Disease Control and Prevention, Nanjing, China
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Takaoka N, Yamane M, Hasegawa A, Obara K, Shirai K, Aki R, Hatakeyama H, Hamada Y, Arakawa N, Tanaka M, Hoffman RM, Amoh Y. Rat hair-follicle-associated pluripotent (HAP) stem cells can differentiate into atrial or ventricular cardiomyocytes in culture controlled by specific supplementation. PLoS One 2024; 19:e0297443. [PMID: 38277391 PMCID: PMC10817212 DOI: 10.1371/journal.pone.0297443] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2023] [Accepted: 01/04/2024] [Indexed: 01/28/2024] Open
Abstract
There has been only limited success to differentiate adult stem cells into cardiomyocyte subtypes. In the present study, we have successfully induced beating atrial and ventricular cardiomyocytes from rat hair-follicle-associated pluripotent (HAP) stem cells, which are adult stem cells located in the bulge area. HAP stem cells differentiated into atrial cardiomyocytes in culture with the combination of isoproterenol, activin A, bone morphogenetic protein 4 (BMP4), basic fibroblast growth factor (bFGF), and cyclosporine A (CSA). HAP stem cells differentiated into ventricular cardiomyocytes in culture with the combination of activin A, BMP4, bFGF, inhibitor of Wnt production-4 (IWP4), and vascular endothelial growth factor (VEGF). Differentiated atrial cardiomyocytes were specifically stained for anti-myosin light chain 2a (MLC2a) antibody. Ventricular cardiomyocytes were specially stained for anti-myosin light chain 2v (MLC2v) antibody. Quantitative Polymerase Chain Reaction (qPCR) showed significant expression of MLC2a in atrial cardiomyocytes and MLC2v in ventricular cardiomyocytes. Both differentiated atrial and ventricular cardiomyocytes showed characteristic waveforms in Ca2+ imaging. Differentiated atrial and ventricular cardiomyocytes formed long myocardial fibers and beat as a functional syncytium, having a structure similar to adult cardiomyocytes. The present results demonstrated that it is possible to induce cardiomyocyte subtypes, atrial and ventricular cardiomyocytes, from HAP stem cells.
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Affiliation(s)
- Nanako Takaoka
- Department of Dermatology, Kitasato University Graduate School of Medical Sciences, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Michiko Yamane
- Department of Dermatology, Kitasato University Graduate School of Medical Sciences, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Ayami Hasegawa
- Department of Dermatology, Kitasato University Graduate School of Medical Sciences, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Hiroyasu Hatakeyama
- Department of Physiology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Manabu Tanaka
- Bio-Imaging Center, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
| | - Robert M. Hoffman
- AntiCancer, Inc., San Diego, CA, United States of America
- Department of Surgery, University of California, San Diego, CA, United States of America
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Kitasato University School of Medicine, Minami Ward, Sagamihara, Japan
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Human-derived hair follicle stem cells and hydrogen sulfide on focal cerebral ischemia model: A comparative evaluation of radiologic, neurobehavioral and immunohistochemical results. Brain Res 2023; 1799:148170. [PMID: 36410427 DOI: 10.1016/j.brainres.2022.148170] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2022] [Revised: 11/10/2022] [Accepted: 11/15/2022] [Indexed: 11/19/2022]
Abstract
The present study investigated the effects of intracerebral human-derived hair follicle stem cells (HFBSCs), whether alone or in combination with hydrogen sulfide (H2S) in a rat model of focal cerebral ischemia. The rats were randomly assigned into 4 groups (n = 10): Control (phosphate buffered saline (PBS)), Group A (at 24 h post-middle cerebral artery occlusion(MCAo), stereotaxic intracerebral, 1,0 × 106, total 10 μL HFBSCs), Group B (3-14 d post-MCAo, intraperitoneal (i.p.), 25 μM/kg/day H2S), Group AB (HFBSCs + H2S). Cranial magnetic resonance images were recorded on postoperative 1st and 28th days. Three dimensional analysis was performed to calculate the infarct volumes. Rotarod and cylinder tests were performed after MCAo and finally all rats were euthanized by cardiac perfusion at 28 days after MCAo for immunohistochemical analysis. The reduction in infarct volumes of rats receiving HFBSC was significant. The cranial infarct volume on the postoperative 28th day was significantly higher in the group in which H2S was administered alone compared to the HFBSC alone group. All animals showed steadily improved spontaneous locomotor activity from day 7 post-MCAo on rotarod test, from day 1 on cylinder test, but showed no significant differences at all times. In all groups, the grading scores of CD34, CD5, CD11b and GFAP immunohistochemical markers did not differ significantly. In conclusion, intracerebral HFBSC treatment after 24 h of ischemic stroke may be an effective way to reduce the cranial infarct volume, whereas H2S treatment alone or in combination with HFBSC may not be sufficient for ischemic brain injury.
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Pandamooz S, Jurek B, Dianatpour M, Haerteis S, Limm K, Oefner PJ, Dargahi L, Borhani-Haghighi A, Miyan JA, Salehi MS. The beneficial effects of chick embryo extract preconditioning on hair follicle stem cells: A promising strategy to generate Schwann cells. Cell Prolif 2023:e13397. [PMID: 36631409 DOI: 10.1111/cpr.13397] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2022] [Revised: 12/06/2022] [Accepted: 12/28/2022] [Indexed: 01/13/2023] Open
Abstract
The beneficial effects of hair follicle stem cells in different animal models of nervous system conditions have been extensively studied. While chick embryo extract (CEE) has been used as a growth medium supplement for these stem cells, this is the first study to show the effect of CEE on them. The rat hair follicle stem cells were isolated and supplemented with 10% fetal bovine serum plus 10% CEE. The migration rate, proliferative capacity and multipotency were evaluated along with morphometric alteration and differentiation direction. The proteome analysis of CEE content identified effective factors of CEE that probably regulate fate and function of stem cells. The CEE enhances the migration rate of stem cells from explanted bulges as well as their proliferation, likely due to activation of AP-1 and translationally controlled tumour protein (TCTP) by thioredoxin found in CEE. The increased length of outgrowth may be the result of cyclic AMP response element binding protein (CREB) phosphorylation triggered by active CamKII contained in CEE. Further, CEE supplementation upregulates the expression of vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor. The elevated expression of target genes and proteins may be due to CREB, AP-1 and c-Myc activation in these stem cells. Given the increased transcript levels of neurotrophins, VEGF, and the expression of PDGFR-α, S100B, MBP and SOX-10 protein, it is possible that CEE promotes the fate of these stem cells towards Schwann cells.
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Affiliation(s)
- Sareh Pandamooz
- Stem Cells Technology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Benjamin Jurek
- Research Group Neurobiology of Stress Resilience, Max Planck Institute of Psychiatry, Munich, Germany.,Institute of Molecular and Cellular Anatomy, University of Regensburg, Regensburg, Germany
| | - Mehdi Dianatpour
- Stem Cells Technology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Silke Haerteis
- Institute of Molecular and Cellular Anatomy, University of Regensburg, Regensburg, Germany
| | - Katharina Limm
- Institute of Functional Genomics, University of Regensburg, Regensburg, Germany
| | - Peter J Oefner
- Institute of Functional Genomics, University of Regensburg, Regensburg, Germany
| | - Leila Dargahi
- Neuroscience Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | | | - Jaleel A Miyan
- Faculty of Biology, Medicine & Health, Division of Neuroscience & Experimental Psychology, The University of Manchester, Manchester, UK
| | - Mohammad Saied Salehi
- Clinical Neurology Research Center, Shiraz University of Medical Sciences, Shiraz, Iran
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Obara K, Shirai K, Hamada Y, Arakawa N, Hasegawa A, Takaoka N, Aki R, Hoffman RM, Amoh Y. Direct implantation of hair-follicle-associated pluripotent (HAP) stem cells repairs intracerebral hemorrhage and reduces neuroinflammation in mouse model. PLoS One 2023; 18:e0280304. [PMID: 36638123 PMCID: PMC9838830 DOI: 10.1371/journal.pone.0280304] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2022] [Accepted: 12/27/2022] [Indexed: 01/14/2023] Open
Abstract
Intracerebral hemorrhage (ICH) is a leading cause of mortality with ineffective treatment. Hair-follicle-associated pluripotent (HAP) stem cells can differentiate into neurons, glial cells and many other types of cells. HAP stem cells have been shown to repair peripheral-nerve and spinal-cord injury in mouse models. In the present study, HAP stem cells from C57BL/6J mice were implanted into the injured brain of C57BL/6J or nude mice with induced ICH. After allo transplantation, HAP stem cells differentiated to neurons, astrocytes, oligodendrocytes, and microglia in the ICH site of nude mice. After autologous transplantation in C57BL/6J mice, HAP stem cells suppressed astrocyte and microglia infiltration in the injured brain. The mRNA expression levels of IL-10 and TGF-β1, measured by quantitative Real-Time RT-PCR, in the brain of C57BL/6J mice with ICH was increased by HAP-stem-cell implantation compared to the non-implanted mice. Quantitative sensorimotor function analysis, with modified limb-placing test and the cylinder test, demonstrated a significant functional improvement in the HAP-stem-cell-implanted C57BL/6J mice, compared to non-implanted mice. HAP stem cells have critical advantages over induced pluripotent stem cells, embryonic stem cells as they do not develop tumors, are autologous, and do not require genetic manipulation. The present study demonstrates future clinical potential of HAP-stem-cell repair of ICH, currently a recalcitrant disease.
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Affiliation(s)
- Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Ayami Hasegawa
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Nanako Takaoka
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Robert M. Hoffman
- AntiCancer, Inc., San Diego, California, United States of America
- Department of Surgery, University of California San Diego, San Diego, California, United States of America
- * E-mail: (YA); (RMH)
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
- * E-mail: (YA); (RMH)
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Hair-follicle-associated pluripotent (HAP) stem cells differentiate into mature beating cardiomyocyte sheets on flexible substrates in vitro. Med Mol Morphol 2022; 55:248-257. [PMID: 35536435 DOI: 10.1007/s00795-022-00322-z] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2022] [Accepted: 04/18/2022] [Indexed: 10/18/2022]
Abstract
Cardiomyocytes have been differentiated from various stem cells such as human embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC), but it is difficult to produce mature cardiomyocytes. We showed rat hair-follicle-associated pluripotent (HAP) stem cells have pluripotency and produced mature beating cardiomyocyte sheets differentiated from rat HAP stem cells. The upper parts of rat vibrissa hair follicles were cultured in 10% FBS DMEM and stained with antibodies of the ectoderm, mesoderm, endoderm system to show the differentiation of multiple cell types. Moreover, HAP stem cells were cultured under three different conditions to decide the most suitable culture conditions for making beating cardiomyocyte sheets. The beating cardiomyocyte sheets were shown to be mature by staining sarcomere structures. Isoproterenol alone and the combination of isoproterenol, activin A, bone morphogenetic protein 4 (BMP4) and basic fibroblast growth factor (bFGF) effectively induced beating long-fiber cardiomyocytes, which formed beating sheets, only in the presence of all four agents. Flexible substrates were essential for the differentiation of sheets of mature beating cardiomyocytes for HAP stem cells. The features of the cardiomyocytes differentiated from HAP stem cells demonstrate they have clinical potential for heart regeneration.
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Obara K, Shirai K, Hamada Y, Arakawa N, Yamane M, Takaoka N, Aki R, Hoffman RM, Amoh Y. Chronic spinal cord injury functionally repaired by direct implantation of encapsulated hair-follicle-associated pluripotent (HAP) stem cells in a mouse model: Potential for clinical regenerative medicine. PLoS One 2022; 17:e0262755. [PMID: 35085322 PMCID: PMC8794105 DOI: 10.1371/journal.pone.0262755] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2021] [Accepted: 01/04/2022] [Indexed: 11/30/2022] Open
Abstract
Chronic spinal cord injury (SCI) is a highly debilitating and recalcitrant disease with limited treatment options. Although various stem cell types have shown some clinical efficacy for injury repair they have not for SCI. Hair-follicle-associated pluripotent (HAP) stem cells have been shown to differentiate into neurons, Schwan cells, beating cardiomyocytes and many other type of cells, and have effectively regenerated acute spinal cord injury in mouse models. In the present report, HAP stem cells from C57BL/6J mice, encapsulated in polyvinylidene fluoride membranes (PFM), were implanted into the severed thoracic spinal cord of C57BL/6J or athymic nude mice in the early chronic phase. After implantation, HAP stem cells differentiated to neurons, astrocytes and oligodendrocytes in the regenerated thoracic spinal cord of C57BL/6J and nude mice. Quantitative motor function analysis, with the Basso Mouse Scale for Locomotion (BMS) score, demonstrated a significant functional improvement in the HAP-stem-cell-implanted mice, compared to non-implanted mice. HAP stem cells have critical advantages over other stem cells: they do not develop teratomas; do not loose differentiation ability when cryopreserved and thus are bankable; are autologous, readily obtained from anyone; and do not require genetic manipulation. HAP stem cells therefore have greater clinical potential for SCI repair than induced pluripotent stem cells (iPSCs), neuronal stem cells (NSCs)/neural progenitor cells (NPCs) or embryonic stem cells (ESCs). The present report demonstrates future clinical potential of HAP-stem-cell repair of chronic spinal cord injury, currently a recalcitrant disease.
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Affiliation(s)
- Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Michiko Yamane
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Nanako Takaoka
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
| | - Robert M. Hoffman
- AntiCancer, Inc., San Diego, California, United States of America
- Department of Surgery, University of California San Diego, San Diego, California, United States of America
- * E-mail: (YA); (RMH)
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan
- * E-mail: (YA); (RMH)
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Peterson A, Nair L. Hair Follicle Stem Cells for Tissue Regeneration. TISSUE ENGINEERING PART B-REVIEWS 2021; 28:695-706. [PMID: 34238037 PMCID: PMC9419938 DOI: 10.1089/ten.teb.2021.0098] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
With the positive outcomes of various cell therapies currently under pre-clinical and clinical studies, there is a significant interest in novel stem cell sources with unique therapeutic properties. Studies over the past two decades or so demonstrated the feasibility to isolate multipotent/pluripotent stem cells from hair follicles. The easy accessibility, high proliferation and differentiation ability as well as lack of ethical concerns associated with this stem cell source make hair follicle stem cells (HFSCs) attractive candidate for cell therapy and tissue engineering. This review discusses the various stem cell types identified in rodent and human hair follicles and ongoing studies on the potential use of HFSCs for skin, bone, cardio-vascular, and nerve tissue engineering.
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Affiliation(s)
- Alyssa Peterson
- University of Connecticut, 7712, Storrs, Connecticut, United States;
| | - Lakshmi Nair
- University of Connecticut Health Center, 21654, Orthopaedic Surgery, Farmington, Connecticut, United States;
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Characterisation of Neurospheres-Derived Cells from Human Olfactory Epithelium. Cells 2021; 10:cells10071690. [PMID: 34359860 PMCID: PMC8307784 DOI: 10.3390/cells10071690] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2021] [Revised: 06/25/2021] [Accepted: 06/30/2021] [Indexed: 11/17/2022] Open
Abstract
A major problem in psychiatric research is a deficit of relevant cell material of neuronal origin, especially in large quantities from living individuals. One of the promising options is cells from the olfactory neuroepithelium, which contains neuronal progenitors that ensure the regeneration of olfactory receptors. These cells are easy to obtain with nasal biopsies and it is possible to grow and cultivate them in vitro. In this work, we used RNAseq expression profiling and immunofluorescence microscopy to characterise neurospheres-derived cells (NDC), that simply and reliably grow from neurospheres (NS) obtained from nasal biopsies. We utilized differential expression analysis to explore the molecular changes that occur during transition from NS to NDC. We found that processes associated with neuronal and vascular cells are downregulated in NDC. A comparison with public transcriptomes revealed a depletion of neuronal and glial components in NDC. We also discovered that NDC have several metabolic features specific to neuronal progenitors treated with the fungicide maneb. Thus, while NDC retain some neuronal/glial identity, additional protocol alterations are needed to use NDC for mass sample collection in psychiatric research.
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Yamane M, Takaoka N, Obara K, Shirai K, Aki R, Hamada Y, Arakawa N, Hoffman RM, Amoh Y. Hair-Follicle-Associated Pluripotent (HAP) Stem Cells Can Extensively Differentiate to Tyrosine-Hydroxylase-Expressing Dopamine-Secreting Neurons. Cells 2021; 10:864. [PMID: 33920157 PMCID: PMC8069047 DOI: 10.3390/cells10040864] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2021] [Revised: 04/08/2021] [Accepted: 04/08/2021] [Indexed: 12/27/2022] Open
Abstract
Hair-follicle-associated pluripotent (HAP) stem cells are located in the bulge area of hair follicles from mice and humans and have been shown to differentiate to neurons, glia, keratinocytes, smooth muscle cells, melanocytes and beating 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 this process. HAP stem cells can be banked by cryopreservation and preserve their ability to differentiate. In the present study, we demonstrated that mouse HAP stem cells cultured in neural-induction medium can extensively differentiate to dopaminergic neurons, which express tyrosine hydroxylase and secrete dopamine. These results indicate that the dopaminergic neurons differentiated from HAP stem cells may be useful in the future to improve the symptoms of Parkinson's disease in the clinic.
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Affiliation(s)
- Michiko Yamane
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Nanako Takaoka
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
| | - Robert M. Hoffman
- AntiCancer, Inc., 7917 Ostrow Street, San Diego, CA 92111, USA
- Department of Surgery, University of California, San Diego, CA 92037-7220, USA
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara 252-0374, Japan; (M.Y.); (N.T.); (K.O.); (K.S.); (R.A.); (Y.H.); (N.A.)
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Hopf A, Schaefer DJ, Kalbermatten DF, Guzman R, Madduri S. Schwann Cell-Like Cells: Origin and Usability for Repair and Regeneration of the Peripheral and Central Nervous System. Cells 2020; 9:E1990. [PMID: 32872454 PMCID: PMC7565191 DOI: 10.3390/cells9091990] [Citation(s) in RCA: 38] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2020] [Revised: 08/06/2020] [Accepted: 08/22/2020] [Indexed: 12/14/2022] Open
Abstract
Functional recovery after neurotmesis, a complete transection of the nerve fiber, is often poor and requires a surgical procedure. Especially for longer gaps (>3 mm), end-to-end suturing of the proximal to the distal part is not possible, thus requiring nerve graft implantation. Artificial nerve grafts, i.e., hollow fibers, hydrogels, chitosan, collagen conduits, and decellularized scaffolds hold promise provided that these structures are populated with Schwann cells (SC) that are widely accepted to promote peripheral and spinal cord regeneration. However, these cells must be collected from the healthy peripheral nerves, resulting in significant time delay for treatment and undesired morbidities for the donors. Therefore, there is a clear need to explore the viable source of cells with a regenerative potential similar to SC. For this, we analyzed the literature for the generation of Schwann cell-like cells (SCLC) from stem cells of different origins (i.e., mesenchymal stem cells, pluripotent stem cells, and genetically programmed somatic cells) and compared their biological performance to promote axonal regeneration. Thus, the present review accounts for current developments in the field of SCLC differentiation, their applications in peripheral and central nervous system injury, and provides insights for future strategies.
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Affiliation(s)
- Alois Hopf
- Department of Biomedical Engineering, University of Basel, Gewerbestrasse 14, 4123 Allschwil, Switzerland; (A.H.); (D.F.K.)
- Department of Biomedicine, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland; (D.J.S.); (R.G.)
| | - Dirk J. Schaefer
- Department of Biomedicine, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland; (D.J.S.); (R.G.)
- Department of Plastic, Reconstructive, Aesthetic and Hand Surgery, University Hospital Basel, University of Basel, Spitalstrasse 21, 4031 Basel, Switzerland
| | - Daniel F. Kalbermatten
- Department of Biomedical Engineering, University of Basel, Gewerbestrasse 14, 4123 Allschwil, Switzerland; (A.H.); (D.F.K.)
- Department of Plastic, Reconstructive, Aesthetic and Hand Surgery, University Hospital Basel, University of Basel, Spitalstrasse 21, 4031 Basel, Switzerland
| | - Raphael Guzman
- Department of Biomedicine, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland; (D.J.S.); (R.G.)
- Department of Neurosurgery, University Hospital Basel, Spitalstrasse 21, 4031 Basel, Switzerland
| | - Srinivas Madduri
- Department of Biomedical Engineering, University of Basel, Gewerbestrasse 14, 4123 Allschwil, Switzerland; (A.H.); (D.F.K.)
- Department of Biomedicine, University Hospital Basel, Hebelstrasse 20, 4031 Basel, Switzerland; (D.J.S.); (R.G.)
- Department of Plastic, Reconstructive, Aesthetic and Hand Surgery, University Hospital Basel, University of Basel, Spitalstrasse 21, 4031 Basel, Switzerland
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Obara K, Tohgi N, Shirai K, Mii S, Hamada Y, Arakawa N, Aki R, Singh SR, Hoffman RM, Amoh Y. Hair-Follicle-Associated Pluripotent (HAP) Stem Cells Encapsulated on Polyvinylidene Fluoride Membranes (PFM) Promote Functional Recovery from Spinal Cord Injury. Stem Cell Rev Rep 2020; 15:59-66. [PMID: 30341634 DOI: 10.1007/s12015-018-9856-3] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Our previous studies showed that nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells, which reside in the bulge area of the hair follicle, could restore injured nerve and spinal cord and differentiate into cardiac muscle cells. Here we transplanted mouse green fluorescent protein (GFP)-expressing HAP stem-cell colonies enclosed on polyvinylidene fluoride membranes (PFM) into the severed thoracic spinal cord of nude mice. After seven weeks of implantation, we found the differentiation of HAP stem cells into neurons and glial cells. Our results also showed that PFM-captured GFP-expressing HAP stem-cell colonies assisted complete reattachment of the thoracic spinal cord. Furthermore, our quantitative motor function analysis with the Basso Mouse Scale for Locomotion (BMS) score demonstrated a significant improvement in the implanted mice compared to non-implanted mice with a severed spinal cord. Our study also showed that it is easy to obtain HAP stem cells, they do not develop teratomas, and do not loose differentiation ability when cryopreserved. Collectively our results suggest that HAP stem cells could be a better source compared to induced pluripotent stem cells (iPS) or embryonic stem (ES) cells for regenerative medicine, specifically for spinal cord repair.
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Affiliation(s)
- Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Natsuko Tohgi
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Sumiyuki Mii
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Shree Ram Singh
- Basic Research Laboratory, National Cancer Institute, Frederick, MD, 21702, USA.
| | - Robert M Hoffman
- AntiCancer, Inc., 7917 Ostrow Street, San Diego, CA, 92111, USA. .,Department of Surgery, University of California, San Diego, CA, 92103, USA.
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan.
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Aran S, Zahri S, Asadi A, Khaksar F, Abdolmaleki A. Hair follicle stem cells differentiation into bone cells on collagen scaffold. Cell Tissue Bank 2020; 21:181-188. [PMID: 32016616 DOI: 10.1007/s10561-020-09812-9] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2019] [Accepted: 01/24/2020] [Indexed: 12/17/2022]
Abstract
The hair follicle is a dynamic structure which contains different niches for stem cells, therefore; it has been considered as valuable and rich sources of stem cells, due to easy access, multipotency and non-oncogenic properties. In the present study, the differentiation capacities of hair follicle stem cells into bone cells on the natural collagen scaffolds were investigated. The stem cells were extracted from the hair follicle bulge area of male Wistar rats' whisker and cultured until 3rd passage, then osteogenic differentiations were induced by culturing the cells in the specific osteogenic medium. After 3 weeks, the differentiation parameters, including morphological changes, levels of calcification and expression of the bone specific genes were detected. The hydrogel preparation and scaffold fabrication was carried out using the extracted collagen and was studied by scanning electron microscope. Comparison of the stem cells' growth and changes on the scaffold and non-scaffold conditions showed that, in the both situation, the cells revealed differentiation signs of osteocytes, including large and cubic morphology with a star-shaped nucleus. Staining by Alizarin-red and Von-Kossa methods showed the presence of red and black calcium mass on the scaffold. Expression of the osteopontin and alkaline phosphatase genes confirmed the differentiation. Considerable porosity in the surface of the scaffold was recorded by scanning electron microscopy, which made it convenient for cells' attachment and growth. The data showed that the bulge stem cells possess significant capacity for osteoblastic differentiation and collagen scaffolds were found to be an appropriate matrix for growth and differentiation of the cell.
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Affiliation(s)
- Saeideh Aran
- Department of Biology, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran
| | - Saber Zahri
- Department of Biology, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran.
| | - Asadollah Asadi
- Department of Biology, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran
| | - Fatemeh Khaksar
- Department of Biology, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran
| | - Arash Abdolmaleki
- Department of Engineering Sciences, Faculty of Advanced Technologies, University of Mohaghegh Ardabili, Namin, Iran
- Bio Science and Biotechnology Research Center (BBRC), Sabalan University of Advanced Technologies (SUAT), Namin, Iran
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16
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Shirai K, Obara K, Tohgi N, Yamazaki A, Aki R, Hamada Y, Arakawa N, Singh SR, Hoffman RM, Amoh Y. Expression of anti-aging type-XVII collagen (COL17A1/BP180) in hair follicle-associated pluripotent (HAP) stem cells during differentiation. Tissue Cell 2019; 59:33-38. [PMID: 31383286 DOI: 10.1016/j.tice.2019.06.001] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2019] [Accepted: 06/10/2019] [Indexed: 02/07/2023]
Abstract
Hair-follicle-associated pluripotent (HAP) stem cells reside in the upper part of the bulge area of the the hair follicle. HAP stem cells are nestin-positive and keratin 15-negative and have the capacity to differentiate into various types of cells in vitro. HAP stem cells are also involved in nerve and spinal cord regeneration in mouse models. Recently, it was shown that the DNA-damage response in non-HAP hair follicle stem cells induces proteolysis of type-XVII collagen (COL17A1/BP180), which is involved in hair-follicle stem-cell maintenance. COL17A1 proteolysis stimulated hair-follicle stem-cell aging, characterized by the loss of stemness signatures and hair-follicle miniaturization associated with androgenic alopecia. In the present study, we demonstrate that HAP stem cells co-express nestin and COL17A1 in vitro and in vivo. The expression of HAP stem cell markers (nestin and SSEA1) increased after HAP stem-cell colonies were formed, then decreased after differentiation to epidermal keratinocytes. In contrast COL17A1 increased after differentiation to epidermal keratinocytes. These results suggest that COL17A1 is important in differentiation of HAP stem cells.
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Affiliation(s)
- Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Koya Obara
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Natsuko Tohgi
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Aiko Yamazaki
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan
| | - Shree Ram Singh
- Basic Research Laboratory, National Cancer Institute, Frederic, MD, 21702, USA.
| | - Robert M Hoffman
- AntiCancer, Inc., San Diego, CA, USA; Department of Surgery, University of California San Diego, CA, USA.
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Sagamihara, Japan.
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Human Hair Follicle Associated-Pluripotent (hHAP) Stem Cells Differentiate to Cardiac Muscle Cells. Methods Mol Biol 2019. [PMID: 29992514 DOI: 10.1007/7651_2018_170] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
Human hair follicle-associated pluripotent (hHAP) stem cells were isolated from the upper parts of human hair follicles. hHAP stem cells were suspended in DMEM containing 10% fetal bovine serum (FBS) where they differentiated to cardiac muscle cells as well as neurons, glial cells, keratinocytes, and smooth muscle cells. The methods of this chapter are appropriate for use of human hair follicles to produce hHAP stem cells in sufficient quantities for future heart, nerve, and spinal cord regeneration in the clinic.
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18
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Mokabber H, Najafzadeh N, Mohammadzadeh Vardin M. miR-124 promotes neural differentiation in mouse bulge stem cells by repressing Ptbp1 and Sox9. J Cell Physiol 2019; 234:8941-8950. [PMID: 30417370 DOI: 10.1002/jcp.27563] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2018] [Accepted: 09/13/2018] [Indexed: 01/08/2023]
Abstract
Hair follicle stem cells (HFSCs) are able to differentiate into neurons and glial cells. Distinct microRNAs (miRNAs) regulate the proliferation and differentiation of HFSCs. However, the exact role of miR-124 in the neural differentiation of HFSCs has not been elucidated. HFSCs were isolated from mouse whisker follicles. miR-9, let-7b, and miR-124, Ptbp1 , and Sox9 expression levels were detected by real-time polymerase chain reaction (RT-PCR). The influence of miR-124 transfection was evaluated using immunostaining. We demonstrated that miR-124 and let-7b expression levels were significantly increased after the neural differentiation. Sox9 and Ptbp1 were identified as the target of miR-124 in the HFSCs. During neural differentiation and miR-124 mimicking, Ptbp1 and Sox9 levels were decreased. Moreover, the miR-124 overexpression increased MAP2 (58.43 ± 11.26) and NeuN (48.34 ± 11.15) proteins expression. The results demonstrated that miR-124 may promote the differentiation of HFSCs into neuronal cells by targeting Sox9 and Ptbp1.
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Affiliation(s)
- Haleh Mokabber
- Department of Anatomical Sciences and Pathology, Research Laboratory for Embryology and Stem Cells, Ardabil University of Medical Sciences, Ardabil, Iran
- Department of Biology, Fars Science and Research Branch, Islamic Azad University, Fars, Iran
| | - Nowruz Najafzadeh
- Department of Anatomical Sciences and Pathology, Research Laboratory for Embryology and Stem Cells, Ardabil University of Medical Sciences, Ardabil, Iran
| | - Mohammad Mohammadzadeh Vardin
- Department of Anatomical Sciences and Pathology, Research Laboratory for Embryology and Stem Cells, Ardabil University of Medical Sciences, Ardabil, Iran
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Nestin + progenitor cells isolated from adult human sweat gland stroma promote reepithelialisation and may stimulate angiogenesis in wounded human skin ex vivo. Arch Dermatol Res 2019; 311:325-330. [PMID: 30798352 PMCID: PMC6469855 DOI: 10.1007/s00403-019-01889-x] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/26/2018] [Revised: 12/11/2018] [Accepted: 12/17/2018] [Indexed: 12/22/2022]
Abstract
The combination of an aging population and an increasing prevalence of diseases associated with impaired-wound healing, including obesity, peripheral vascular disease and diabetes, is likely to result in a dramatic increase in the incidence and prevalence of chronic skin wounds. Indeed, systemic reviews are now not only trying to establish both the prevalence and the often under-estimated socio-economic costs of chronic skin wounds, but most importantly are addressing the impact that chronic wounds have on quality of life. Given the clear need for novel approaches to the management of chronic skin ulceration, ideally developed and tested in the human system in a manner that can be rapidly translated into clinical practice, we examined the effects of multipotent primary human nestin+ progenitor cells on human wound healing in an ex vivo model. Human sweat gland-derived nestin+ cells demonstrated the capacity to significantly promote two key wound healing parameters, i.e., both reepithelialisation and angiogenesis in experimentally wounded, organ-cultured human skin. The current data further support the use of full-thickness human skin wound-healing models ex vivo to pre-clinically test wound healing-promoting candidate agents. Whilst larger studies are required to substantiate a firm “proof-of-concept,” our preliminary studies encourage further efforts to systemically determine the potential of cell-based regenerative medicine strategies in general, and the use of skin appendage-associated human nestin+ cells in particular, as novel treatment strategies for chronic skin ulceration.
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Skin Stem Cells, Their Niche and Tissue Engineering Approach for Skin Regeneration. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2019; 1212:107-126. [PMID: 31065940 DOI: 10.1007/5584_2019_380] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
Skin is the main organ that covers the human body and acts as a protective barrier between the human body and the environment. Skin tissue as a stem cell source can be used for transplantation in therapeutic application in terms of its properties such as abundant, easy to access, high plasticity and high ability to regenerate. The immunological profile of these cells makes it a suitable resource for autologous and allogeneic applications. The lack of major histo-compatibility complex 1 is also advantageous in its use. Epidermal stem cells are the main stem cells in the skin and are suitable cells in tissue engineering studies for their important role in wound repair. In the last 30 years, many studies have been conducted to develop substitutions that mimic human skin. Stem cell-based skin substitutions have been developed to be used in clinical applications, to support the healing of acute and chronic wounds and as test systems for dermatological and pharmacological applications. In this chapter, tissue specific properties of epidermal stem cells, composition of their niche, regenerative approaches and repair of tissue degeneration have been examined.
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Hoffman RM, Amoh Y. Hair Follicle-Associated Pluripotent(HAP) Stem Cells. PROGRESS IN MOLECULAR BIOLOGY AND TRANSLATIONAL SCIENCE 2018; 160:23-28. [PMID: 30470290 DOI: 10.1016/bs.pmbts.2018.09.001] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
The hair follicle has been known, since 1990, to contain stem cells located in the bulge area. In 2003, we reported a new type of stem cell in the hair follicle that expresses the brain stem-cell marker nestin. We have termed these cells as hair-follicle-associated pluripotent (HAP) stem cells. HAP stem cells can differentiate into neuronal and glial cells, beating cardiac-muscle cells, and other cell types in culture. HAP stem cells can be used for nerve and spinal-cord repair such that locomotor activity is recovered. A major function in situ of the HAP stem cells is for growth of the hair follicle sensory nerve. HAP stem cells have critical advantages over embryonic stem cells and induced pluripotent stem (IPS) cells for regenerative medicine in that they are highly accessible, require no genetic manipulation, are nontumorigenic, and do not present ethical issues.
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Affiliation(s)
- Robert M Hoffman
- AntiCancer Inc., San Diego, CA, United States; Department of Surgery, University of California, San Diego, CA, United States.
| | - Yasuyuki Amoh
- Kitasato University School of Medicine, Sagamihara, Japan
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22
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Beating Heart Cells from Hair-Follicle-Associated Pluripotent (HAP) Stem Cells. Methods Mol Biol 2018. [PMID: 30196415 DOI: 10.1007/978-1-4939-8697-2_18] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
The neural stem-cell marker nestin is expressed in hair follicle stem cells located in the bulge area which are termed hair-follicle-associated pluripotent (HAP) stem cells. HAP stem cells can differentiate into neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. Subsequently, we demonstrated that HAP stem cells could affect nerve and spinal cord regeneration in mouse models. We subsequently demonstrated that HAP stem cells differentiated into beating cardiac muscle cells. The differentiation potential to cardiac muscle is greatest in the upper part of the mouse whisker follicle. The beat rate of the cardiac muscle cells differentiated from HAP stem cells was stimulated by isoproterenol and inhibited by propanolol. The addition of activin A, bone morphogenetic protein 4, and basic fibroblast growth factor, along with isoproternal, induced the cardiac muscle cells to form tissue sheets of beating heart muscle cells. Under hypoxic conditions, HAP stem cells differentiated into troponin-positive cardiac-muscle cells at a higher rate that under normoxic conditions. Hypoxia did not influence the differentiation to other cell types. 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.
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23
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Zheng Y, Huang C, Liu F, Lin H, Niu Y, Yang X, Zhang Z. Reactivation of denervated Schwann cells by neurons induced from bone marrow-derived mesenchymal stem cells. Brain Res Bull 2018. [DOI: 10.1016/j.brainresbull.2018.03.005] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023]
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Hair Follicle-Associated Pluripotent (HAP) Stem Cells in Gelfoam ® Histoculture for Use in Spinal Cord Repair. Methods Mol Biol 2018. [PMID: 29572802 DOI: 10.1007/978-1-4939-7745-1_15] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
The stem cell marker, nestin, is expressed in the hair follicle, both in cells in the bulge area (BA) and the dermal papilla (DP). Nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells of both the BA and DP have been previously shown to be able to form neurons, heart muscle cells, and other non-follicle cell types. The ability of the nestin-expressing HAP stem cells from the BA and DP to repair spinal cord injury was compared. Nestin-expressing HAP stem cells from both the BA and DP grew very well on Gelfoam®. The HAP stem cells attached to the Gelfoam® within 1 h. They grew along the grids of the Gelfoam® during the first 2 or 3 days. Later they spread into the Gelfoam®. After transplantation of Gelfoam® cultures of nestin-expressing BA or DP HAP stem cells into the injured spinal cord (including the Gelfoam®) nestin-expressing BA and DP cells were observed to be viable over 100 days post-surgery. Hematoxylin and eosin (H&E) staining showed connections between the transplanted cells and the host spine tissue. Immunohistochemistry showed many Tuj1-, Isl 1/2, and EN1-positive cells and nerve fibers in the transplanted area of the spinal cord after BA Gelfoam® or DP Gelfoam® cultures were transplanted to the spine. The spinal cord of mice was injured to effect hind-limb paralysis. Twenty-eight days after transplantation with BA or DP HAP stem cells on Gelfoam® to the injured area of the spine, the mice recovered normal locomotion.
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25
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Huang H, Young W, Chen L, Feng S, Zoubi ZMA, Sharma HS, Saberi H, Moviglia GA, He X, Muresanu DF, Sharma A, Otom A, Andrews RJ, Al-Zoubi A, Bryukhovetskiy AS, Chernykh ER, Domańska-Janik K, Jafar E, Johnson WE, Li Y, Li D, Luan Z, Mao G, Shetty AK, Siniscalco D, Skaper S, Sun T, Wang Y, Wiklund L, Xue Q, You SW, Zheng Z, Dimitrijevic MR, Masri WSE, Sanberg PR, Xu Q, Luan G, Chopp M, Cho KS, Zhou XF, Wu P, Liu K, Mobasheri H, Ohtori S, Tanaka H, Han F, Feng Y, Zhang S, Lu Y, Zhang Z, Rao Y, Tang Z, Xi H, Wu L, Shen S, Xue M, Xiang G, Guo X, Yang X, Hao Y, Hu Y, Li J, AO Q, Wang B, Zhang Z, Lu M, Li T. Clinical Cell Therapy Guidelines for Neurorestoration (IANR/CANR 2017). Cell Transplant 2018; 27:310-324. [PMID: 29637817 PMCID: PMC5898693 DOI: 10.1177/0963689717746999] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2017] [Revised: 07/22/2017] [Accepted: 11/13/2017] [Indexed: 12/11/2022] Open
Abstract
Cell therapy has been shown to be a key clinical therapeutic option for central nervous system diseases or damage. Standardization of clinical cell therapy procedures is an important task for professional associations devoted to cell therapy. The Chinese Branch of the International Association of Neurorestoratology (IANR) completed the first set of guidelines governing the clinical application of neurorestoration in 2011. The IANR and the Chinese Association of Neurorestoratology (CANR) collaborated to propose the current version "Clinical Cell Therapy Guidelines for Neurorestoration (IANR/CANR 2017)". The IANR council board members and CANR committee members approved this proposal on September 1, 2016, and recommend it to clinical practitioners of cellular therapy. These guidelines include items of cell type nomenclature, cell quality control, minimal suggested cell doses, patient-informed consent, indications for undergoing cell therapy, contraindications for undergoing cell therapy, documentation of procedure and therapy, safety evaluation, efficacy evaluation, policy of repeated treatments, do not charge patients for unproven therapies, basic principles of cell therapy, and publishing responsibility.
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Affiliation(s)
- Hongyun Huang
- Institute of Neurorestoratology, General Hospital of Armed Police Forces, Beijing, People’s Republic of China
| | - Wise Young
- W. M. Keck Center for Collaborative Neuroscience, Rutgers, State University of New Jersey, Piscataway, NJ, USA
| | - Lin Chen
- Department of Neurosurgery, Tsinghua University Yuquan Hospital, Beijing, People’s Republic of China
| | - Shiqing Feng
- Department of Orthopaedics, Tianjin Medical University General Hospital, Tianjin, People’s Republic of China
| | - Ziad M. Al Zoubi
- Jordan Ortho and Spinal Centre, Al-Saif Medical Center, Amman, Jordan
| | - Hari Shanker Sharma
- Intensive Experimental CNS Injury and Repair, University Hospital, Uppsala University, Uppsala, Sweden
| | - Hooshang Saberi
- Department of Neurosurgery, Brain and Spinal Injury Research center, Tehran University of Medical Sciences, Tehran, Iran
| | - Gustavo A. Moviglia
- Center of Research and Engineer of Tissues and Cellular Therapy, Maimonides University, Buenos Aires, Argentina
| | - Xijing He
- Department of Orthopaedics, Second Affiliated Hospital of Xi’an Jiaotong University, Xian, People’s Republic of China
| | - Dafin F. Muresanu
- Department of Neurosciences “Iuliu Hatieganu,” University of Medicine and Pharmacy, Cluj-Napoca, Romania
| | - Alok Sharma
- Department of Neurosurgery, LTM Medical College, LTMG Hospital, Mumbai, Mumbai, India
| | - Ali Otom
- Royal Rehabilitation Center, King Hussein Medical Centre-RJRC Amman, Jordan
| | - Russell J. Andrews
- Nanotechnology & Smart Systems, NASA Ames Research Center, Silicon Valley, CA, USA
| | - Adeeb Al-Zoubi
- The University of Illinois College of Medicine in Peoria, Peoria, IL, USA
| | - Andrey S. Bryukhovetskiy
- NeuroVita Clinic of Interventional and Restorative Neurology and Therapy, Kashirskoye shosse, Moscow, Russia
| | - Elena R. Chernykh
- Lab of Cellular Immunotherapy, Institute of Fundamental and Clinical Immunology, Novosibirsk, Russia
| | | | - Emad Jafar
- Jordan Ortho and Spinal Centre, Al-Saif Medical Center, Amman, Jordan
| | - W. Eustace Johnson
- Stem Cells and Regenerative Biology, Faculty of Medicine Dentistry and Life Sciences, University of Chester, Chester, United Kingdom
| | - Ying Li
- Spinal Repair Unit, Department of Brain Repair and Rehabilitation, UCL Institute of Neurology, London, United Kingdom
| | - Daqing Li
- Spinal Repair Unit, Department of Brain Repair and Rehabilitation, UCL Institute of Neurology, London, United Kingdom
| | - Zuo Luan
- Department of Pediatrics, Navy General Hospital of PLA, Beijing, People’s Republic of China
| | - Gengsheng Mao
- Institute of Neurorestoratology, General Hospital of Armed Police Forces, Beijing, People’s Republic of China
| | - Ashok K. Shetty
- Department of Molecular and Cellular Medicine, Institute for Regenerative Medicine, Texas A&M Health Science Center College of Medicine, College Station, TX, USA
| | - Dario Siniscalco
- Department of Experimental Medicine, University of Campania “Luigi Vanvitelli,” Naples, Italy
| | - Stephen Skaper
- Department of Pharmaceutical and Pharmacological Sciences, University of Padua, Padua, Italy
| | - Tiansheng Sun
- Department of orthopedics, PLA Army General Hospital, Beijing, People’s Republic of China
| | - Yunliang Wang
- Department of Neurology, 148th Hospital, Zibo, Shandong, People’s Republic of China
| | - Lars Wiklund
- Unit of Neurology, Department of Pharmacology and Clinical Neuroscience, Umea University, Ostersund, Sweden
| | - Qun Xue
- Department of Neurology, the First Affiliated Hospital of Soochow University, Suzhou Jiangsu, People’s Republic of China
| | - Si-Wei You
- Department of Ophthalmology, Xijing Hospital, Fourth Military Medical University, Xi’an, People’s Republic of China
| | - Zuncheng Zheng
- Department of Rehabilitation Medicine, The Central Hospital of Taian, Taian, Shandong, People’s Republic of China
| | | | - W. S. El Masri
- Spinal Injuries Unit, Robert Jones and Agnes Hunt Orthopaedic Hospital, Oswestry, United Kingdom
| | - Paul R. Sanberg
- Center of Excellence for Aging & Brain Repair, Morsani College of Medicine, University of South Florida, Tampa, FL, USA
| | - Qunyuan Xu
- Institute of Neuroscience, Capital Medical University, Beijing, People’s Republic of China
| | - Guoming Luan
- Department of Neurosurgery, Beijing Sanbo Brain Hospital, Capital Medical University, Beijing, People’s Republic of China
| | - Michael Chopp
- Henry Ford Hospital, Henry Ford Health System, Neurology Research, Detroit, MI, USA
| | - Kyoung-Suok Cho
- Department of Neurosurgery, Uijongbu St. Mary’s Hospital, College of Medicine, The Catholic University of Korea, Uijongbu, South Korea
| | - Xin-Fu Zhou
- Division of Health Sciences, School of Pharmacy and Medical Sciences, Sansom Institute for Health Research, University of South Australia, Adelaide, South Australia, Australia
| | - Ping Wu
- Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX, USA
| | - Kai Liu
- Division of Life Science, The Hong Kong University of Science and Technology, Kowloon, Hong Kong
| | - Hamid Mobasheri
- Biomaterials Research Center, Institute of Biochemistry and Biophysics, University of Tehran, Tehran, Iran
| | - Seiji Ohtori
- Department of Orthopedic Surgery, Graduate School of Medicine, Chiba University, Chiba, Japan
| | - Hiroyuki Tanaka
- Department of Orthopaedic Surgery, Graduate School of Medicine, Osaka University, Osaka, Japan
| | - Fabin Han
- Centre for Stem Cells and Regenerative Medicine, Liaocheng University/Liaocheng People’s Hospital, Liaocheng, Shandong, People’s Republic of China
| | - Yaping Feng
- Department of Neurosurgery, Kunming General Hospital of Chengdu Military Command of Chinese PLA, Kunming, Yunnan, People’s Republic of China
| | - Shaocheng Zhang
- Department of Orthopedics, Changhai Hospital, The Second Military Medical University, Shanghai, People’s Republic of China
| | - Yingjie Lu
- Department of Neurosurgery, Chengde Dadu Hospital, Weichang, Hebei, People’s Republic of China
| | - Zhicheng Zhang
- Department of orthopedics, PLA Army General Hospital, Beijing, People’s Republic of China
| | - Yaojian Rao
- Department of Spinal Surgery, Luoyang Orthopedic Hospital of Henan Province, Luoyang, Henan, People’s Republic of China
| | - Zhouping Tang
- Department of Neurology, Tongji Medical College of HUST, Tongji Hospital, Wuhan, People’s Republic of China
| | - Haitao Xi
- Department of Neurology, Beijing Rehabilitation Hospital of Capital Medical University, Beijing, People’s Republic of China
| | - Liang Wu
- Center of Rehabilitation, Beijing Xiaotangshan Rehabilitation Hospital, Beijing, People’s Republic of China
| | - Shunji Shen
- Department of Rehabilitation, Weihai Municipal Hospital, Weihai, Shandong, People’s Republic of China
| | - Mengzhou Xue
- Department of Neurorehabilitation, The Second Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, People’s Republic of China
| | - Guanghong Xiang
- Brain Hospital of Hunan Province, Changsha, Hunan, People’s Republic of China
| | - Xiaoling Guo
- Department of Neurology, PLA Army 266 Hospital, Chengde, Hebei, People’s Republic of China
| | - Xiaofeng Yang
- Department of Neurosurgery, The First Affiliated Hospital of Zhejiang University Medical College, Hangzhou, Zhejiang, People’s Republic of China
| | - Yujun Hao
- Department of Neurosurgery, The First Affiliated Hospital of Xinjiang Medical University, Urumqi, Xinjiang, People’s Republic of China
| | - Yong Hu
- Department of Orthopaedic and Traumatology, The University of Hong Kong, Pokfulam, Hong Kong
| | - Jinfeng Li
- Unit of Neurology, Department of Pharmacology and Clinical Neuroscience, Umea University, Ostersund, Sweden
| | - Qiang AO
- Department of tissue engineering, China Medical University, Shenyang, Liaoning, People’s Republic of China
| | - Bin Wang
- Department of Traumatology, The Second Affiliated Hospital of Guangzhou Medical University, Haizhu District, Guangzhou, People’s Republic of China
| | - Zhiwen Zhang
- Department of Neurosurgery, First Affiliated Hospital of Chinese PLA General Hospital, Beijing, People’s Republic of China
| | - Ming Lu
- Department of Neurosurgery, Second Affiliated Hospital of Hunan Normal University (163 Hospital of PLA), Changsha, Hunan, People’s Republic of China
| | - Tong Li
- Department of Neurology, The Second Affiliated Hospital of Xinxiang Medical University, Xinxiang, Henan, People’s Republic of China
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26
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Protocols for Cryopreservation of Intact Hair Follicle That Maintain Pluripotency of Nestin-Expressing Hair-Follicle-Associated Pluripotent (HAP) Stem Cells. Methods Mol Biol 2018; 1453:173-8. [PMID: 27431257 DOI: 10.1007/978-1-4939-3786-8_18] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Hair follicles contain nestin-expressing pluripotent stem cells, the origin of which is above the bulge area, below the sebaceous gland. We have termed these cells hair-follicle-associated pluripotent (HAP) stem cells. Cryopreservation methods of the hair follicle that maintain the pluripotency of HAP stem cells are described in this chapter. Intact hair follicles from green fluorescent protein (GFP) transgenic mice were cryopreserved by slow-rate cooling in TC-Protector medium and storage in liquid nitrogen. After thawing, the upper part of the hair follicle was isolated and cultured in DMEM with fetal bovine serum (FBS). After 4 weeks culture, cells from the upper part of the hair follicles grew out. The growing cells were transferred to DMEM/F12 without FBS. After 1 week culture, the growing cells formed hair spheres, each containing approximately 1 × 10(2) HAP stem cells. The hair spheres contained cells which could differentiate to neurons, glial cells, and other cell types. The formation of hair spheres by the thawed and cultured upper part of the hair follicle produced almost as many pluripotent hair spheres as fresh follicles. The hair spheres derived from cryopreserved hair follicles were as pluripotent as hair spheres from fresh hair follicles. These results suggest that the cryopreservation of the whole hair follicle is an effective way to store HAP stem cells for personalized regenerative medicine, enabling any individual to maintain a bank of pluripotent stem cells for future clinical use.
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27
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Amoh Y, Katsuoka K, Hoffman RM. Peripheral-Nerve and Spinal-Cord Regeneration in Mice Using Hair-Follicle-Associated Pluripotent (HAP) Stem Cells. Methods Mol Biol 2018; 1453:21-32. [PMID: 27431243 DOI: 10.1007/978-1-4939-3786-8_4] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/14/2023]
Abstract
Nestin, a neural stem cell marker protein, is expressed in hair follicle cells above the bulge area. These nestin-positive hair follicle-associated-pluripotent (HAP) stem cells are negative for the keratinocyte marker K15 and can differentiate into neurons, glia, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes in vitro. HAP stem cells are positive for the stem cell marker CD34, as well as K15-negative, suggesting their relatively undifferentiated state. HAP stem cells promoted the functional recovery of injured peripheral nerves and the spinal cord. HAP stem cells differentiated into glial fibrillary acidic protein (GFAP)-positive Schwann cells when implanted in severed sciatic nerves and spinal cords in mice. These results suggest that HAP stem cells provide an important accessible, autologous source of adult stem cells for regenerative medicine, that have critical advantages over ES and iPS stem cells.
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Affiliation(s)
- Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan.,AntiCancer, Inc., 7917 Ostrow Street, San Diego, CA, 92111, USA.,Department of Surgery, University of California, San Diego, San Diego, CA, 92103, USA
| | - Kensei Katsuoka
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, 252-0374, Japan
| | - Robert M Hoffman
- AntiCancer, Inc., 7917 Ostrow Street, San Diego, CA, 92111, USA. .,Department of Surgery, University of California, San Diego, San Diego, CA, 92103, USA.
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28
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Abstract
The hair follicle undergoes lifelong cycling and growth. Previous studies have been focused on epithelial stem cells in the hair follicles. Neural crest stem cells (NCSCs) are pluripotent cells that can persist in adult tissues. We have previously demonstrated that human NCSCs can be isolated from hair follicles. Here, we present a protocol to isolate NCSCs from human hair follicles based on their specific surface-marker expression of CD271/HNK1 or CD271/CD49D (alpha4 integrin). NCSCs can be expanded in the culture as neural spheres or attached cells.
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29
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Pandamooz S, Salehi MS, Zibaii MI, Ahmadiani A, Nabiuni M, Dargahi L. Epidermal neural crest stem cell-derived glia enhance neurotrophic elements in an ex vivo model of spinal cord injury. J Cell Biochem 2018; 119:3486-3496. [PMID: 29143997 DOI: 10.1002/jcb.26520] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2017] [Accepted: 11/13/2017] [Indexed: 01/09/2023]
Abstract
Growing evidence that cell-based therapies can improve recovery outcome in spinal cord injury (SCI) models substantiates their application for treatment of human with SCI. To address the effectiveness of these stem cells, potential candidates should be evaluated in proper SCI platform that allows direct real-time monitoring. In this study, the role of epidermal neural crest stem cells (EPI-NCSCs) was elucidated in an ex vivo model of SCI, and valproic acid (VPA) was administered to ameliorate the inhospitable context of injury for grafted EPI-NCSCs. Here the contusion was induced in organotypic spinal cord slice culture at day seven in vitro using a weight drop device and one hour post injury the GFP- expressing EPI-NCSCs were grafted followed by VPA administration. The evaluation of treated slices seven days after injury revealed that grafted stem cells survived on the injured slices and expressed GFAP, whereas they did not express any detectable levels of the neural progenitor marker doublecortin (DCX), which was expressed prior to transplantation. Immunoblotting data demonstrated that the expression of GFAP, BDNF, neurotrophin-3 (NT3), and Bcl2 increased significantly in stem cell treated slices. This study illustrated that the fate of transplanted stem cells has been directed to the glial lineage in the ex vivo context of injury and EPI-NCSCs may ameliorate the SCI condition through releasing neurotrophic factors directly and/or via inducing resident spinal cord cells.
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Affiliation(s)
- Sareh Pandamooz
- Neuroscience Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.,Department of Animal Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran
| | - Mohammad S Salehi
- Neuroscience Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Mohammad I Zibaii
- Laser and Plasma Research institute, Shahid Beheshti University, Tehran, Iran
| | - Abolhassan Ahmadiani
- Neuroscience Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Mohammad Nabiuni
- Department of Animal Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran
| | - Leila Dargahi
- NeuroBiology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran
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30
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Hoffman RM, Mii S, Duong J, Amoh Y. Nerve Growth and Interaction in Gelfoam ® Histoculture: A Nervous System Organoid. Methods Mol Biol 2018; 1760:163-186. [PMID: 29572803 DOI: 10.1007/978-1-4939-7745-1_16] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/08/2023]
Abstract
Nestin-expressing hair follicle-associated pluripotent (HAP) stem cells reside mainly in the bulge area (BA) of the hair follicle but also in the dermal papilla (DP). The BA appears to be origin of HAP stem cells. Long-term Gelfoam® histoculture was established of whiskers isolated from transgenic mice, in which there is nestin-driven green fluorescent protein (ND-GFP). HAP stem cells trafficked from the BA toward the DP area and extensively grew out onto Gelfoam® forming nerve-like structures. These fibers express the neuron marker β-III tubulin-positive fibers and consisted of ND-GFP-expressing cells and extended up to 500 mm from the whisker nerve stump in Gelfoam® histoculture. The growing fibers had growth cones on their tips expressing F-actin indicating that the fibers were growing axons. HAP stem cell proliferation resulted in elongation of the follicle nerve and interaction with other nerves in 3D Gelfoam® histoculture, including the sciatic nerve, trigeminal nerve, and trigeminal nerve ganglion.
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Affiliation(s)
- Robert M Hoffman
- AntiCancer Inc., San Diego, CA, USA.
- Department of Surgery, UCSD, San Diego, CA, USA.
| | - Sumiyuki Mii
- AntiCancer Inc., San Diego, CA, USA
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagihara, Japan
| | | | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagihara, Japan
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31
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Si Y, Bai J, Wu J, Li Q, Mo Y, Fang R, Lai W. LncRNA PlncRNA‑1 regulates proliferation and differentiation of hair follicle stem cells through TGF‑β1‑mediated Wnt/β‑catenin signal pathway. Mol Med Rep 2017; 17:1191-1197. [PMID: 29115537 DOI: 10.3892/mmr.2017.7944] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2017] [Accepted: 09/27/2017] [Indexed: 11/06/2022] Open
Abstract
The present study demonstrated that hair follicle stem cells (HFSc) have multidirectional differentiation potential and participate in skin wound healing processes. Long non‑coding RNAs (lncRNAs) are defined as non‑protein coding transcripts longer than 200 nucleotides, which are important in the proliferation and differentiation of cells. The purpose of the present study was to investigate the role of PlncRNA‑1 in the proliferation and differentiation of HFSc. Results revealed that PlncRNA‑1, transforming growth factor (TGF)‑β1, Wnt and β‑catenin expression levels were significantly downregulated in HFSc. PlncRNA‑1 transfection promoted proliferation and differentiation of HFSc. TGF‑β1, Wnt and β‑catenin expression levels were upregulated in HFSc following transfection of PlncRNA‑1. Results demonstrated that TGF‑β1 inhibitor LY2109761 blocked proliferation and differentiation of HFSc promoted by PlncRNA‑1 transfection. In addition, TGF‑β1 inhibitor LY2109761 led to decreased Wnt and β‑catenin expression levels in HFSc. Furthermore, PlncRNA‑1 transfection stimulated the cell cycle of HFSc, whereas TGF‑β1 inhibitor LY2109761 inhibited the cell cycle of HFSc and decreased the acceleration of the cell cycle induced by PlncRNA‑1 transfection. In conclusion, these findings suggest that PlncRNA‑1 may promote proliferation and differentiation of HFSc through upregulation of TGF‑β1‑mediated Wnt/β‑catenin signaling pathway.
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Affiliation(s)
- Yuan Si
- Department of Dermatology, The Third Affiliated Hospital, Sun Yat‑sen University, Guangzhou, Guangdong 510180, P.R. China
| | - Jingzhu Bai
- Department of Dermatology, Guangzhou First People's Hospital, Guangzhou, Guangdong 510630, P.R. China
| | - Jiang Wu
- Department of Dermatology, Guangzhou First People's Hospital, Guangzhou, Guangdong 510630, P.R. China
| | - Qun Li
- Department of Dermatology, Guangzhou First People's Hospital, Guangzhou, Guangdong 510630, P.R. China
| | - You Mo
- Department of Dermatology, Guangzhou First People's Hospital, Guangzhou, Guangdong 510630, P.R. China
| | - Ruihua Fang
- Department of Dermatology, Guangzhou First People's Hospital, Guangzhou, Guangdong 510630, P.R. China
| | - Wei Lai
- Department of Dermatology, The Third Affiliated Hospital, Sun Yat‑sen University, Guangzhou, Guangdong 510180, P.R. China
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32
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Flores AF, Varela-Vazquez A, Mayan MD, Fonseca E. Expression of connexin 43 in the human hair follicle: emphasis on the connexin 43 protein levels in the bulge and through the keratinization process. J Cutan Pathol 2017; 45:8-15. [DOI: 10.1111/cup.13050] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2017] [Revised: 09/13/2017] [Accepted: 09/21/2017] [Indexed: 12/22/2022]
Affiliation(s)
- Angel Fernandez Flores
- Department of Cellular Pathology; Hospital El Bierzo; Ponferrada Spain
- Department of Cellular Pathology; Hospital de la Reina; Ponferrada Spain
- CellCOM-SB Group, Institute for Biomedical Research of A Coruña (INIBIC); University of A Coruña (UDC); A Coruña Spain
| | - Adrian Varela-Vazquez
- CellCOM-SB Group, Institute for Biomedical Research of A Coruña (INIBIC); University of A Coruña (UDC); A Coruña Spain
| | - Maria D. Mayan
- CellCOM-SB Group, Institute for Biomedical Research of A Coruña (INIBIC); University of A Coruña (UDC); A Coruña Spain
| | - Eduardo Fonseca
- CellCOM-SB Group, Institute for Biomedical Research of A Coruña (INIBIC); University of A Coruña (UDC); A Coruña Spain
- Department of Dermatology; University Hospital of A Coruña; A Coruña Spain
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Schomann T, Mezzanotte L, De Groot JCMJ, Rivolta MN, Hendriks SH, Frijns JHM, Huisman MA. Neuronal differentiation of hair-follicle-bulge-derived stem cells co-cultured with mouse cochlear modiolus explants. PLoS One 2017; 12:e0187183. [PMID: 29084289 PMCID: PMC5662184 DOI: 10.1371/journal.pone.0187183] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2017] [Accepted: 10/16/2017] [Indexed: 11/18/2022] Open
Abstract
Stem-cell-based repair of auditory neurons may represent an attractive therapeutic option to restore sensorineural hearing loss. Hair-follicle-bulge-derived stem cells (HFBSCs) are promising candidates for this type of therapy, because they (1) have migratory properties, enabling migration after transplantation, (2) can differentiate into sensory neurons and glial cells, and (3) can easily be harvested in relatively high numbers. However, HFBSCs have never been used for this purpose. We hypothesized that HFBSCs can be used for cell-based repair of the auditory nerve and we have examined their migration and incorporation into cochlear modiolus explants and their subsequent differentiation. Modiolus explants obtained from adult wild-type mice were cultured in the presence of EF1α-copGFP-transduced HFBSCs, constitutively expressing copepod green fluorescent protein (copGFP). Also, modiolus explants without hair cells were co-cultured with DCX-copGFP-transduced HFBSCs, which demonstrate copGFP upon doublecortin expression during neuronal differentiation. Velocity of HFBSC migration towards modiolus explants was calculated, and after two weeks, co-cultures were fixed and processed for immunohistochemical staining. EF1α-copGFP HFBSC migration velocity was fast: 80.5 ± 6.1 μm/h. After arrival in the explant, the cells formed a fascicular pattern and changed their phenotype into an ATOH1-positive neuronal cell type. DCX-copGFP HFBSCs became green-fluorescent after integration into the explants, confirming neuronal differentiation of the cells. These results show that HFBSC-derived neuronal progenitors are migratory and can integrate into cochlear modiolus explants, while adapting their phenotype depending on this micro-environment. Thus, HFBSCs show potential to be employed in cell-based therapies for auditory nerve repair.
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Affiliation(s)
- Timo Schomann
- Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands
| | - Laura Mezzanotte
- Optical Molecular Imaging Group, Department of Radiology, Erasmus Medical Center, Rotterdam, South Holland, the Netherlands
| | - John C. M. J. De Groot
- Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands
| | - Marcelo N. Rivolta
- Centre for Stem Cell Biology, Department of Biomedical Science, University of Sheffield, Sheffield, England, United Kingdom
| | - Sanne H. Hendriks
- Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands
| | - Johan H. M. Frijns
- Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands
| | - Margriet A. Huisman
- Department of Otorhinolaryngology and Head & Neck Surgery, Leiden University Medical Center, Leiden, South Holland, the Netherlands
- * E-mail:
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34
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Yamazaki A, Obara K, Tohgi N, Shirai K, Mii S, Hamada Y, Arakawa N, Aki R, Hoffman RM, Amoh Y. Implanted hair-follicle-associated pluripotent (HAP) stem cells encapsulated in polyvinylidene fluoride membrane cylinders promote effective recovery of peripheral nerve injury. Cell Cycle 2017; 16:1927-1932. [PMID: 28886268 DOI: 10.1080/15384101.2017.1363941] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/18/2022] Open
Abstract
Hair follicle-associated-pluripotent (HAP) stem cells are located in the bulge area of the hair follicle, express the stem-cell marker, nestin, and have been shown to differentiate to nerve cells, glial cells, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes. Transplanted HAP stem cells promote the recovery of peripheral nerve and spinal cord injuries and have the potential for heart regeneration as well. In the present study, we implanted mouse green fluorescent protein (GFP)-expressing HAP stem-cell spheres encapsulated in polyvinylidene fluoride (PVDF)-membrane cylinders into the severed sciatic nerve of immunocompetent and immunocompromised (nude) mice. Eight weeks after implantation, immunofluorescence staining showed that the HAP stem cells differentiated into neurons and glial cells. Fluorescence microscopy showed that the HAP stem cell hair spheres promoted rejoining of the sciatic nerve of both immunocompetent and immunodeficient mice. Hematoxylin and eosin (H&E) staining showed that the severed scatic nerves had regenerated. Quantitative walking analysis showed that the transplanted mice recovered the ability to walk normally. HAP stem cells are readily accessible from everyone, do not form tumors, and can be cryopreserved without loss of differentiation potential. These results suggest that HAP stem cells may have greater potential than iPS or ES cells for regenerative medicine.
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Affiliation(s)
- Aiko Yamazaki
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Kohya Obara
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Natsuko Tohgi
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Kyoumi Shirai
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Sumiyuki Mii
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Yuko Hamada
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Nobuko Arakawa
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Ryoichi Aki
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
| | - Robert M Hoffman
- b AntiCancer, Inc ., San Diego , CA , USA.,c Department of Surgery , University of California San Diego , CA , USA
| | - Yasuyuki Amoh
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward, Sagamihara , Japan
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Abstract
Various types of stem cells reside in the skin, including keratinocyte progenitor cells, melanocyte progenitor cells, skin-derived precursors (SKPs), and nestin-expressing hair follicle-associated-pluripotent (HAP) stem cells. HAP stem cells, located in the bulge area of the hair follicle, have been shown to differentiate to nerve cells, glial cells, keratinocytes, smooth muscle cells, cardiac muscle cells, and melanocytes. HAP stem cells are positive for the stem-cell marker CD34, as well as K15-negative, suggesting their relatively undifferentiated state. Therefore, HAP stem cells may be the most primitive stem cells in the skin. Moreover, HAP stem cells can regenerate the epidermis and at least parts of the hair follicle. These results suggest that HAP stem cells may be the origin of other stem cells in the skin. Transplanted HAP stem cells promote the recovery of peripheral-nerve and spinal-cord injuries and have the potential for heart regeneration as well. HAP stem cells are readily accessible from everyone, do not form tumors, and can be cryopreserved without loss of differentiation potential. These results suggest that HAP stem cells may have greater potential than iPS or ES cells for regenerative medicine.
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Affiliation(s)
- Yasuyuki Amoh
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Robert M Hoffman
- b AntiCancer, Inc. , San Diego , CA , USA.,c Department of Surgery , University of California San Diego , San Diego , CA , USA
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Li Y, Yao D, Zhang J, Liu B, Zhang L, Feng H, Li B. The Effects of Epidermal Neural Crest Stem Cells on Local Inflammation Microenvironment in the Defected Sciatic Nerve of Rats. Front Mol Neurosci 2017; 10:133. [PMID: 28588447 PMCID: PMC5438963 DOI: 10.3389/fnmol.2017.00133] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2017] [Accepted: 04/20/2017] [Indexed: 12/21/2022] Open
Abstract
Cell-based therapy is a promising strategy for the repair of peripheral nerve injuries (PNIs). epidermal neural crest stems cells (EPI-NCSCs) are thought to be important donor cells for repairing PNI in different animal models. Following PNI, inflammatory response is important to regulate the repair process. However, the effects of EPI-NCSCs on regulation of local inflammation microenviroment have not been investigated extensively. In the present study, these effects were studied by using 10 mm defected sciatic nerve, which was bridged with 15 mm artificial nerve composed of EPI-NCSCs, extracellular matrix (ECM) and poly (lactide-co-glycolide) (PLGA). Then the expression of pro- and anti-inflammatory cytokines, polarization of macrophages, regulation of fibroblasts and shwann cells (SCs) were assessed by western blot, immunohistochemistry, immunofluorescence staining at 1, 3, 7 and 21 days after bridging. The structure and the function of the bridged nerve were determined by observation under light microscope and by examination of right lateral foot retraction time (LFRT), sciatic function index (SFI), gastrocnemius wet weight and electrophysiology at 9 weeks. After bridging with EPI-NCSCs, the expression of anti-inflammatory cytokines (IL-4 and IL-13) was increased, but decreased for pro-inflammatory cytokines (IL-6 and TNF-α) compared to the control bridging, which was consistent with increase of M2 macrophages and decrease of M1 macrophages at 7 days after transplantation. Likewise, myelin-formed SCs were significantly increased, but decreased for the activated fibroblasts in their number at 21 days. The recovery of structure and function of nerve bridged with EPI-NCSCs was significantly superior to that of DMEM. These results indicated that EPI-NCSCs could be able to regulate and provide more suitable inflammation microenvironment for the repair of defected sciatic nerve.
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Affiliation(s)
- Yue Li
- Department of Neurosurgery, Southwest Hospital/State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical UniversityChongqing, China
| | - Dongdong Yao
- Research Institute of Surgery, Daping Hospital/State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical UniversityChongqing, China.,School of Life Sciences/Key Laboratory of Freshwater Fish Reproduction and Development of Education Ministry, Southwest UniversityChongqing, China
| | - Jieyuan Zhang
- Research Institute of Surgery, Daping Hospital/State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical UniversityChongqing, China
| | - Bin Liu
- School of Life Sciences/Key Laboratory of Freshwater Fish Reproduction and Development of Education Ministry, Southwest UniversityChongqing, China
| | - Lu Zhang
- Children's Hospital of Chongqing Medical University/Ministry of Education Key Laboratory of Child Development and Disorders, Chongqing Medical UniversityChongqing, China
| | - Hua Feng
- Department of Neurosurgery, Southwest Hospital/State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical UniversityChongqing, China
| | - Bingcang Li
- Research Institute of Surgery, Daping Hospital/State Key Laboratory of Trauma, Burns and Combined Injury, Third Military Medical UniversityChongqing, China
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Joulai Veijouyeh S, Mashayekhi F, Yari A, Heidari F, Sajedi N, Moghani Ghoroghi F, Nobakht M. In vitro induction effect of 1,25(OH) 2D 3 on differentiation of hair follicle stem cell into keratinocyte. Biomed J 2017; 40:31-38. [PMID: 28411880 PMCID: PMC6138590 DOI: 10.1016/j.bj.2016.08.007] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2015] [Accepted: 08/02/2016] [Indexed: 12/13/2022] Open
Abstract
Background Stem cells are characterized by self-renewal and differentiation capabilities. The bulge hair follicle stem cells (HFSCs) are able to convert to epithelial components. The active metabolite of vitamin D, 1,25(OH)2D3, plays important roles in this differentiation process. In the present study has found that 1,25(OH)2D3 induces the HFSCs differentiation into keratinocyte. Methods HFSCs are isolated from rat whiskers and cultivated in DMEM medium. To isolate bulge stem cell population, flow cytometry and immunocytochemistry using K15, CD34 and nestin biomarkers were performed. In order to accelerate the HFSCs differentiation into keratinocyte, HFSCs were treated with 10−12 M, 1,25(OH)2D3 every 48 h for a week. Results Immunocytochemistry results showed that bulge stem cells are nestin and CD34 positive but K15 negative before differentiation. Subsequently flow cytometry results, showed that the expression of nestin, CD34 and K15 were 70.96%, 93.03% and 6.88% respectively. After differentiation, the immunocytochemical and flow cytometry results indicated that differentiated cells have positive reaction to K15 with 68.94% expression level. Conclusion It was concluded that 10−12 M, 1,25(OH)2D3 could induce the HFSCs differentiation into keratinocytes.
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Affiliation(s)
- Sanaz Joulai Veijouyeh
- Department of Anatomy, School of Medicine, Iran University of Medical Science, Tehran, Iran; Department of Biology, University Campus 2, University of Guilan, Rasht, Iran
| | - Farhad Mashayekhi
- Department of Biology, Faculty of Sciences, University of Guilan, Rasht, Iran
| | - Abazar Yari
- Department of Anatomy, School of Medicine, Alborz University of Medical Science, Karaj, Iran
| | - Fatemeh Heidari
- Department of Anatomy, School of Medicine, Qom University of Medical Science, Qom, Iran
| | - Nayereh Sajedi
- Department of Anatomy, School of Medicine, Isfahan University of Medical Science, Isfahan, Iran
| | | | - Maliheh Nobakht
- Department of Anatomy, School of Medicine, Iran University of Medical Science, Tehran, Iran; Anti-Microbial Resistance Research Center, Iran University of Medical Science, Tehran, Iran; Physiology Research Center, Iran University of Medical Science, Tehran, Iran.
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Quan R, Du W, Zheng X, Xu S, Li Q, Ji X, Wu X, Shao R, Yang D. VEGF165 induces differentiation of hair follicle stem cells into endothelial cells and plays a role in in vivo angiogenesis. J Cell Mol Med 2017; 21:1593-1604. [PMID: 28244687 PMCID: PMC5542910 DOI: 10.1111/jcmm.13089] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2016] [Accepted: 12/13/2016] [Indexed: 01/20/2023] Open
Abstract
Within the vascular endothelial growth factor (VEGF) family of five subtypes, VEGF165 secreted by endothelial cells has been identified to be the most active and widely distributed factor that plays a vital role in courses of angiogenesis, vascularization and mesenchymal cell differentiation. Hair follicle stem cells (HFSCs) can be harvested from the bulge region of the outer root sheath of the hair follicle and are adult stem cells that have multi-directional differentiation potential. Although the research on differentiation of stem cells (such as fat stem cells and bone marrow mesenchymal stem cells) to the endothelial cells has been extensive, but the various mechanisms and functional forms are unclear. In particular, study on HFSCs' directional differentiation into vascular endothelial cells using VEGF165 has not been reported. In this study, VEGF165 was used as induction factor to induce the differentiation from HFSCs into vascular endothelial cells, and the results showed that Notch signalling pathway might affect the differentiation efficiency of vascular endothelial cells. In addition, the in vivo transplantation experiment provided that HFSCs could promote angiogenesis, and the main function is to accelerate host-derived neovascularization. Therefore, HFSCs could be considered as an ideal cell source for vascular tissue engineering and cell transplantation in the treatment of ischaemic diseases.
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Affiliation(s)
- Renfu Quan
- Research Institute of Orthopedics, The Affiliated JiangNan Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Weibin Du
- Research Institute of Orthopedics, The Affiliated JiangNan Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Xuan Zheng
- Research Institute of Orthopedics, The Affiliated JiangNan Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Shichao Xu
- Research Institute of Orthopedics, The Affiliated JiangNan Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Qiang Li
- Research Institute of Orthopedics, The Affiliated JiangNan Hospital of Zhejiang Chinese Medical University, Hangzhou, China
| | - Xing Ji
- Department of Pharmacology, School of Medical, Zhejiang University, Hangzhou, China
| | - Ximei Wu
- Department of Pharmacology, School of Medical, Zhejiang University, Hangzhou, China
| | - Rongxue Shao
- Research Institute of Orthopedics, Zhejiang Chinese Medical University, Hangzhou, Zhejiang Province, China
| | - Disheng Yang
- Research Institute of Orthopedics, The Second Affiliated Hospital, School of Medical, Zhejiang University, Hangzhou, China
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Cao W, Li L, Kajiura S, Amoh Y, Tan Y, Liu F, Hoffman RM. Aging hair follicles rejuvenated by transplantation to a young subcutaneous environment. Cell Cycle 2017; 15:1093-8. [PMID: 26940664 DOI: 10.1080/15384101.2016.1156269] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/20/2023] Open
Abstract
We demonstrate in the present study that young host mice rejuvenate aged hair follicles after transplantation. Young mice promote the hair shaft growth of transplanted old hair follicles, as well as young follicles, in contrast to old host mice, which did not support hair-shaft growth from transplanted old or young follicles. Nestin-expressing hair follicle-associated pluripotent (HAP) stem cells of transplanted old and young hair follicles remained active in young host nude mice. In contrast, the nestin-expressing HAP stem cells in young and old hair follicles transplanted to old nude mice were not as active as in young nude host mice. The present study shows that transplanted old hair follicles were rejuvenated by young host mice, suggesting that aging may be reversible.
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Affiliation(s)
- Wenluo Cao
- a AntiCancer Inc. , San Diego , CA , USA.,b Department of Surgery , University of California , San Diego , CA , USA.,c Department of Anatomy , Second Military Medical University , Shanghai , China
| | - Lingna Li
- a AntiCancer Inc. , San Diego , CA , USA
| | - Satoshi Kajiura
- a AntiCancer Inc. , San Diego , CA , USA.,b Department of Surgery , University of California , San Diego , CA , USA.,d Department of Dermatology , Kitasato University , Sagimahara , Japan
| | - Yasuyuki Amoh
- d Department of Dermatology , Kitasato University , Sagimahara , Japan
| | - Yuying Tan
- a AntiCancer Inc. , San Diego , CA , USA
| | - Fang Liu
- c Department of Anatomy , Second Military Medical University , Shanghai , China
| | - Robert M Hoffman
- a AntiCancer Inc. , San Diego , CA , USA.,b Department of Surgery , University of California , San Diego , CA , USA
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Lee SH, Jin WP, Seo NR, Pang KM, Kim B, Kim SM, Lee JH. Recombinant human fibroblast growth factor-2 promotes nerve regeneration and functional recovery after mental nerve crush injury. Neural Regen Res 2017; 12:629-636. [PMID: 28553345 PMCID: PMC5436363 DOI: 10.4103/1673-5374.205104] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023] Open
Abstract
Several studies have shown that fibroblast growth factor-2 (FGF2) can directly affect axon regeneration after peripheral nerve damage. In this study, we performed sensory tests and histological analyses to study the effect of recombinant human FGF-2 (rhFGF2) treatment on damaged mental nerves. The mental nerves of 6-week-old male Sprague-Dawley rats were crush-injured for 1 minute and then treated with 10 or 50 μg/mL rhFGF2 or PBS in crush injury area with a mini Osmotic pump. Sensory test using von Frey filaments at 1 week revealed the presence of sensory degeneration based on decreased gap score and increased difference score. However, at 2 weeks, the gap score and difference score were significantly rebounded in the mental nerve crush group treated with 10 μg/mL rhFGF2. Interestingly, treatment with 10 μg/mL rhFGF had a more obviously positive effect on the gap score than treatment with 50 μg/mL rhFGF2. In addition, retrograde neuronal tracing with Dil revealed a significant increase in nerve regeneration in the trigeminal ganglion at 2 and 4 weeks in the rhFGF2 groups (10 μg/mL and 50 μg/mL) than in the PBS group. The 10 μg/mL rhFGF2 group also showed an obviously robust regeneration in axon density in the mental nerve at 4 weeks. Our results demonstrate that 10 μg/mL rhFGF induces mental nerve regeneration and sensory recovery after mental nerve crush injury.
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Affiliation(s)
- Sung Ho Lee
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea
| | - Wei-Peng Jin
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea
| | - Na Ri Seo
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea
| | - Kang-Mi Pang
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea
| | - Bongju Kim
- Dental Life Science Research Institute, Clinical Translational Research Center for Dental Science, Seoul National University Dental Hospital, Seoul, Korea
| | - Soung-Min Kim
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea
| | - Jong-Ho Lee
- Dental Research Institute, Department of Oral and Maxillofacial Surgery, Seoul National University School of Dentistry, Seoul, Korea.,Dental Research Institute, Seoul National University, Seoul, Korea
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Shen Q, Yu W, Fang Y, Yao M, Yang P. Beta-catenin can induce hair follicle stem cell differentiation into transit-amplifying cells through c-myc activation. Tissue Cell 2016; 49:28-34. [PMID: 28049551 DOI: 10.1016/j.tice.2016.12.005] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2016] [Revised: 12/12/2016] [Accepted: 12/17/2016] [Indexed: 12/24/2022]
Abstract
Hair follicle stem cells play important roles in maintaining homeostasis and skin tissue self-renewal. Transit-amplifying cells represent the transition of cells from hair follicle stem cells into differentiated epidermal cells. Thus far, the signaling pathway and the molecular biological mechanism that regulate the proliferation and differentiation of hair follicle stem cells remain unclear. In this paper, we studied the relationship between β-catenin and c-myc during the process of the differentiation of hair follicle stem cells into transit-amplifying cells. Based on our results, the expression of β-catenin can activate the nuclear gene c-myc and regulate the expression of transit-amplifying cell markers K15, K19, a6-integrin and β1-integrin, indicating that β-catenin is involved in the transformation process from hair follicle stem cells to transit-amplifying cells and suggesting that β-catenin plays an important biological role in the induction of this differentiation process.
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Affiliation(s)
- Qiong Shen
- Department of Ultrasound Diagnosis, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 201999, China
| | - Weirong Yu
- Department of Burns and Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 201999, China
| | - Yong Fang
- Department of Burns and Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 201999, China; Institute of Traumatic Medicine, School of Medicine, Shanghai Jiao Tong University, Shanghai 201999, China
| | - Min Yao
- Department of Burns and Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 201999, China; Institute of Traumatic Medicine, School of Medicine, Shanghai Jiao Tong University, Shanghai 201999, China; Wellman Center for Photomedicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA
| | - Penggao Yang
- Department of Burns and Plastic Surgery, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 201999, China.
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Yamazaki A, Yashiro M, Mii S, Aki R, Hamada Y, Arakawa N, Kawahara K, Hoffman RM, Amoh Y. Isoproterenol directs hair follicle-associated pluripotent (HAP) stem cells to differentiate in vitro to cardiac muscle cells which can be induced to form beating heart-muscle tissue sheets. Cell Cycle 2016; 15:760-5. [PMID: 27104748 DOI: 10.1080/15384101.2016.1146837] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/22/2022] Open
Abstract
Nestin-expressing hair-follicle-associated pluripotent (HAP) stem cells are located in the bulge area of the follicle. Previous studies have shown that HAP stem cells can differentiate to neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. HAP stem cells effected nerve and spinal cord regeneration in mouse models. Recently, we demonstrated that HAP stem cells differentiated to beating cardiac muscle cells. The differentiation potential to cardiac muscle cells was greatest in the upper part of the follicle. The beat rate of the cardiac muscle cells was stimulated by isoproterenol. In the present study, we observed that isoproterenol directs HAP stem cells to differentiate to cardiac muscle cells in large numbers in culture compared to HAP stem cells not supplemented with isoproterenol. The addition of activin A, bone morphogenetic protein 4, and basic fibroblast growth factor, along with isoproternal, induced the cardiac muscle cells to form tissue sheets of beating heart muscle cells. These results demonstrate that HAP stem cells have great potential to form beating cardiac muscle cells in tissue sheets.
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Affiliation(s)
- Aiko Yamazaki
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Masateru Yashiro
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Sumiyuki Mii
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Ryoichi Aki
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Yuko Hamada
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Nobuko Arakawa
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Katsumasa Kawahara
- b Department of Physiology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan.,c Department of Cellular & Molecular Physiology , Kitasato Univ Grad Sch Med Sci , Minami Ward , Sagamihara , Japan
| | - Robert M Hoffman
- d AntiCancer, Inc. , San Diego , CA , USA.,e Department of Surgery , University of California San Diego , San Diego , CA USA
| | - Yasuyuki Amoh
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
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Tohgi N, Obara K, Yashiro M, Hamada Y, Arakawa N, Mii S, Aki R, Hoffman RM, Amoh Y. Human hair-follicle associated pluripotent (hHAP) stem cells differentiate to cardiac-muscle cells. Cell Cycle 2016; 16:95-99. [PMID: 27880068 DOI: 10.1080/15384101.2016.1253642] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
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.
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Affiliation(s)
- Natsuko Tohgi
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Koya Obara
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Masateru Yashiro
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Yuko Hamada
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Nobuko Arakawa
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Sumiyuki Mii
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Ryoichi Aki
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
| | - Robert M Hoffman
- b AntiCancer, Inc. , San Diego , CA , USA.,c Department of Surgery , University of California San Diego , CA , USA
| | - Yasuyuki Amoh
- a Department of Dermatology , Kitasato University School of Medicine , Minami Ward , Sagamihara , Japan
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Shirai K, Hamada Y, Arakawa N, Yamazaki A, Tohgi N, Aki R, Mii S, Hoffman RM, Amoh Y. Hypoxia Enhances Differentiation of Hair Follicle-Associated-Pluripotent (HAP) Stem Cells to Cardiac-Muscle Cells. J Cell Biochem 2016; 118:554-558. [PMID: 27627796 DOI: 10.1002/jcb.25734] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2016] [Accepted: 09/12/2016] [Indexed: 01/19/2023]
Abstract
We have previously demonstrated that the neural stem-cell marker nestin is expressed in hair-follicle stem cells located in the bulge area which are termed hair-follicle-associated pluripotent (HAP) stem cells. HAP stem cells from mouse and human could form spheres in culture, termed hair spheres, which are keratin 15-negative and nestin-positive and could differentiate to neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. Subsequently, we demonstrated that nestin-expressing stem cells could effect nerve and spinal cord regeneration in mouse models. Recently, we demonstrated that HAP stem cells differentiated to beating cardiac muscle cells. We recently observed that isoproterenol directs HAP stem cells to differentiate to cardiac-muscle cells in large numbers in culture compared to HAP stem cells not supplemented with isoproterenol. The addition of activin A, bone morphogenetic protein 4, and basic fibroblast growth factor, along with isoproternal, induced the cardiac muscle cells to form tissue sheets of beating heart muscle cells. In the present study, we report that, under hypoxic conditions, HAP stem cells differentiated to troponin-positive cardiac-muscle cells at a higher rate that under normoxic conditions. Hypoxia did not influence the differentiation to other cell types. For future use of HAP stem cells for cardiac muscle regeneration, hypoxia should enhance the rate of differentiation thereby providing patients more opportunities to use their own HAP stem cells which are easily accessible, for this purpose. J. Cell. Biochem. 118: 554-558, 2017. © 2016 Wiley Periodicals, Inc.
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Affiliation(s)
- Kyoumi Shirai
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan.,Department of Dermatology, National Hospital Organization Yokohama Medical Center, Totsuka Ward, Yokohama, 245-8575, Japan
| | - Yuko Hamada
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Nobuko Arakawa
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Aiko Yamazaki
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Natsuko Tohgi
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Ryoichi Aki
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Sumiyuki Mii
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
| | - Robert M Hoffman
- AntiCancer, Inc., 7917 Ostrow Street, San Diego, California, 92111.,Department of Surgery, University of California, San Diego, California, 92103
| | - Yasuyuki Amoh
- Department of Dermatology, Kitasato University School of Medicine, Minami Ward, Sagamihara, Kanagawa, 252-0374, Japan
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Bai T, Liu F, Zou F, Zhao G, Jiang Y, Liu L, Shi J, Hao D, Zhang Q, Zheng T, Zhang Y, Liu M, Li S, Qi L, Liu JY. Epidermal Growth Factor Induces Proliferation of Hair Follicle-Derived Mesenchymal Stem Cells Through Epidermal Growth Factor Receptor-Mediated Activation of ERK and AKT Signaling Pathways Associated with Upregulation of Cyclin D1 and Downregulation of p16. Stem Cells Dev 2016; 26:113-122. [PMID: 27702388 DOI: 10.1089/scd.2016.0234] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
The maintenance of highly proliferative capacity and full differentiation potential is a necessary step in the initiation of stem cell-based regenerative medicine. Our recent study showed that epidermal growth factor (EGF) significantly enhanced hair follicle-derived mesenchymal stem cell (HF-MSC) proliferation while maintaining the multilineage differentiation potentials. However, the underlying mechanism remains unclear. Herein, we investigated the role of EGF in HF-MSC proliferation. HF-MSCs were isolated and cultured with or without EGF. Immunofluorescence staining, flow cytometry, cytochemistry, and western blotting were used to assess proliferation, cell signaling pathways related to the EGF receptor (EGFR), and cell cycle progression. HF-MSCs exhibited surface markers of mesenchymal stem cells and displayed trilineage differentiation potentials toward adipocytes, chondrocytes, and osteoblasts. EGF significantly increased HF-MSC proliferation as well as EGFR, ERK1/2, and AKT phosphorylation (p-EGFR, p-ERK1/2, and p-AKT) in a time- and dose-dependent manner, but not STAT3 phosphorylation. EGFR inhibitor (AG1478), PI3K-AKT inhibitor (LY294002), ERK inhibitor (U0126), and STAT3 inhibitor (STA-21) significantly blocked EGF-induced HF-MSC proliferation. Moreover, AG1478, LY294002, and U0126 significantly decreased p-EGFR, p-AKT, and p-ERK1/2 expression. EGF shifted HF-MSCs at the G1 phase to the S and G2 phase. Concomitantly, cyclinD1, phosphorylated Rb, and E2F1expression increased, while that of p16 decreased. In conclusion, EGF induces HF-MSC proliferation through the EGFR/ERK and AKT pathways, but not through STAT-3. The G1/S transition was stimulated by upregulation of cyclinD1 and inhibition of p16 expression.
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Affiliation(s)
- Tingting Bai
- 1 Department of Pathobiology, Key Laboratory of Ministry of Education, College of Basic Medicine, Jilin University , Changchun, China .,2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Feilin Liu
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China .,3 Department of Ophthalmology, Second Hospital of Jilin University , Changchun, China
| | - Fei Zou
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China .,4 Department of Pediatrics, First Hospital of Jilin University , Changchun, China
| | - Guifang Zhao
- 5 Department of Pathology, Jilin Medical College , Jilin, China
| | - Yixu Jiang
- 1 Department of Pathobiology, Key Laboratory of Ministry of Education, College of Basic Medicine, Jilin University , Changchun, China .,2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Li Liu
- 1 Department of Pathobiology, Key Laboratory of Ministry of Education, College of Basic Medicine, Jilin University , Changchun, China .,2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Jiahong Shi
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Deshun Hao
- 1 Department of Pathobiology, Key Laboratory of Ministry of Education, College of Basic Medicine, Jilin University , Changchun, China .,2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Qi Zhang
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Tong Zheng
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Yingyao Zhang
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Mingsheng Liu
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
| | - Shilun Li
- 6 Department of Oncology, First People's Hospital of Lishu County , Lishu County, China
| | - Liangchen Qi
- 7 Department of Thoracic Surgery, China-Japan Union Hospital of Jilin University , Changchun, China
| | - Jin Yu Liu
- 2 Department of Toxicology, School of Public Health, Jilin University , Changchun, China
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Uchugonova A, Cao W, Hoffman RM, Koenig K. Comparison of label-free and GFP multiphoton imaging of hair follicle-associated pluripotent (HAP) stem cells in mouse whiskers. Cell Cycle 2016; 14:3430-3. [PMID: 26397024 DOI: 10.1080/15384101.2015.1090064] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/02/2023] Open
Abstract
Hair-follicle-associated pluripotent (HAP) stem cells can differentiate into many cell types, including neurons and heart muscle cells, and have been shown to repair peripheral nerves and the spinal cord in mice. HAP stem cells can be obtained from each individual patient for regenerative medicine which overcomes problems with immune rejection. Previously, we have demonstrated that genetically-encoded protein markers such as GFP in transgenic mice can be used to visualize HAP stem cells in vivo by multiphoton tomography. Detection and visualization of stem cells in vivo without exogenous labels such as GFP would be important for human application. In the present report, we demonstrate label-free visualization of hair follicle stem cells in mouse whiskers by multiphoton tomography due to the intrinsic fluorophores such as NAD(P)H/flavins. We compared multiphoton tomography of GFP-labeled HAP stem cells and unlabeled stem cells in isolated mouse whiskers. We show that observation of HAP stem cells by label-free multiphoton tomography is comparable to detection using GFP-labeled stem cells. The results described here have important implications for detection and isolation of human HAP stem cells for regenerative medicine.
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Affiliation(s)
- Aisada Uchugonova
- a Department of Biophotonics and Laser Technology ; Saarland University ; Saarbruecken , Germany.,b AntiCancer Inc. ; San Diego , CA USA.,c Department of Surgery ; University of California San Diego ; San Diego , CA USA
| | - Wenluo Cao
- b AntiCancer Inc. ; San Diego , CA USA.,c Department of Surgery ; University of California San Diego ; San Diego , CA USA
| | - Robert M Hoffman
- b AntiCancer Inc. ; San Diego , CA USA.,c Department of Surgery ; University of California San Diego ; San Diego , CA USA
| | - Karsten Koenig
- a Department of Biophotonics and Laser Technology ; Saarland University ; Saarbruecken , Germany.,d JenLab GmbH ; Jena , Germany
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Discovery of HAP Stem Cells. Methods Mol Biol 2016. [PMID: 27431242 DOI: 10.1007/978-1-4939-3786-8_3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
Cells expressing the stem cell marker, nestin, were selectively labeled in transgenic mice by placing green fluorescent protein (GFP) under the control of the nestin promoter in transgenic mice. In these transgenic mice, neural and other stem cells brightly expressed GFP. The mice were termed nestin-driven GFP (ND-GFP) mice. During early anagen or growth phase of the hair follicle, ND-GFP appeared in the permanent upper hair follicle immediately below the sebaceous glands in the follicle bulge. The relatively small, oval-shaped, nestin-expressing cells in the bulge area surrounded the hair shaft and were interconnected by short dendrites. The location of the nestin-expressing cells in the hair follicle varied with the hair cycle. During telogen or resting phase and in early anagen, the GFP-positive cells are mainly in the bulge area. However, in mid- and late-anagen, the GFP-expressing cells were located in the upper outer-root sheath as well as in the bulge area. The expression of the unique protein, nestin, in both neural stem cells and hair follicle stem cells, which suggested their relationship. The ND-GFP hair follicle stem cells were later termed hair-follicle-associated pluripotent (HAP) stem cells.
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Protocols for Gelfoam(®) Histoculture of Hair-Shaft-Producing Mouse Whisker Follicles Containing Nestin-GFP-Expressing Hair-Follicle-Associated Pluripotent (HAP) Stem Cells for Long Time Periods. Methods Mol Biol 2016. [PMID: 27431254 DOI: 10.1007/978-1-4939-3786-8_15] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
Gelfoam(®)-histocultured whisker follicles from nestin-driven-green fluorescent protein (ND-GFP) mice produced growing pigmented and unpigmented hair shafts. Hair-shaft length increased rapidly by day 4 and continued growing until at least day 12 after which the hair-shaft length was constant. By day 63 in histoculture, the number of ND-GFP hair follicle-associated pluripotent (HAP) stem cells increased significantly and the follicles were intact. Three-dimensional Gelfoam(®) histoculture of hair follicles can provide a very long-term period for evaluating novel agents to promote hair growth.
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Protocols for Ectopic Hair Growth from Transplanted Whisker Follicles on the Spinal Cord of Mice. Methods Mol Biol 2016. [PMID: 27431253 DOI: 10.1007/978-1-4939-3786-8_14] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
Isolated whisker follicles from nestin-driven green fluorescent protein (ND-GFP) mice, containing hair-associated pluripotent (HAP) stem cells, were histocultured in three dimensions on Gelfoam(®) for 3 weeks for subsequent transplantation to the spinal cord in order to heal an induced injury with the HAP stem cells. The hair shafts were removed from Gelfoam(®)-histocultured whisker follicles, and the remaining parts of the whisker follicles, containing GFP-nestin-expressing (HAP) stem cells, were transplanted into the injured spinal cord of nude mice, along with the Gelfoam(®). After 90 days, the mice were sacrificed and the spinal cord injuries were observed to have healed. ND-GFP expression was intense at the healed area of the spinal cord, as observed by fluorescence microscopy, demonstrating that the HAP stem cells were involved in healing the spinal cord. The transplanted whisker follicles produced remarkably long hair shafts in the spinal cord over 90 days and curved and enclosed the spinal cord. This result changes our concept of hair growth, demonstrating it is not limited to the skin and that hair growth appears related to HAP stem cells as both increased in tandem on the spinal cord.
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Yamazaki A, Hamada Y, Arakawa N, Yashiro M, Mii S, Aki R, Kawahara K, Hoffman RM, Amoh Y. Early-age-dependent selective decrease of differentiation potential of hair-follicle-associated pluripotent (HAP) stem cells to beating cardiac-muscle cells. Cell Cycle 2016; 15:2619-2625. [PMID: 27428074 DOI: 10.1080/15384101.2016.1208870] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/21/2022] Open
Abstract
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.
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Affiliation(s)
- Aiko Yamazaki
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Yuko Hamada
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Nobuko Arakawa
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Masateru Yashiro
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Sumiyuki Mii
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Ryoichi Aki
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
| | - Katsumasa Kawahara
- b Department of Physiology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan.,c Department of Cellular & Molecular Physiology , Kitasato University Graduate School of Medical Sciences , Minami Ward, Sagamihara , Japan
| | - Robert M Hoffman
- d AntiCancer, Inc. , San Diego , CA , USA.,e Department of Surgery , University of California San Diego , San Diego , CA , USA
| | - Yasuyuki Amoh
- a Department of Dermatology , Kitasato University School of Medicine, Minami Ward , Sagamihara , Japan
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