Your browser doesn't support javascript.
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 50
Filtrar
Mais filtros










Base de dados
Tipo de estudo
Intervalo de ano de publicação
1.
Sci Rep ; 10(1): 446, 2020 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-31949236

RESUMO

Adrenal cortex autotransplantation with ACTH stimulation may be an alternative therapy for patients with bilateral adrenalectomy to avoid adrenal crisis, but its underlying mechanism has not been elucidated. Previously, we detected Dhh upregulation in rat adrenocortical autografts after transplantation. Here, we investigated potential regulators such as Gata4, Gata6, Sry and Sox9 which affect Dhh transcription in adrenocortical autografts with or without ACTH stimulation. In ACTH-stimulated autografts, Gata4 and Gata6 were downregulated compared to control autografts. This response was linked to rDhh repression. A reporter assay using the upstream region of rDhh and a GATA binding motif revealed that rDhh promoters were significantly upregulated by co-transfection with Gata4 or Gata6 or both. Sry and Sox9 expression in autografts with or without ACTH stimulation were verified by PCR and RNAscope analyses. The ovarian differentiation factors Foxl2 and Rspo1 were also upregulated in the autografts. Gata4 and Gata6 were found to be significant factors in the regulation of rDhh expression and could be associated with adrenocortical autograft maintenance. Gonadal primordia with bipotential testicular and ovarian functions may also be present in these autografts.

2.
Hypertension ; 73(6): 1283-1290, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31006333

RESUMO

Peripheral 18-oxocortisol (18oxoF) level could contribute to the detection of aldosterone-producing adenoma (APA) in patients with primary aldosteronism. However, peripheral 18oxoF varies among such patients, which is a big drawback concerning its clinical application. We studied 48 cases of APA, 35 harboring KCNJ5 mutation, to clarify the significance of clinical and pathological parameters about peripheral 18oxoF. Peripheral 18oxoF concentration ranged widely from 0.50 to 183.13 ng/dL and correlated positively with intratumoral areas stained positively for steroidogenic enzymes ( P<0.0001). The peripheral 18oxoF level also correlated significantly with that of circulating aldosterone ( P<0.0001) but not with that of cortisol, a precursor of 18oxoF. However, a significant correlation was detected between peripheral 18oxoF and intratumoral glucocorticoids ( P<0.05). In addition, peripheral 18oxoF correlated positively with the number of hybrid cells double positive for 11ß-hydroxylase and aldosterone synthase ( P<0.0001). Comparing between the cases with and those without KCNJ5 mutation, the KCNJ5-mutated group demonstrated a significantly higher concentration of peripheral 18oxoF (28.4±5.6 versus 3.0±0.9 ng/dL, P<0.0001) and a larger intratumoral environment including the hybrid cells ( P<0.001), possibly representing a deviation from normal aldosterone biosynthesis. After multivariate analysis, KCNJ5 mutation status turned out to be the most associated factor involved in 18oxoF synthesis in APA ( P<0.0001). Results of our present study first revealed that enhanced 18oxoF synthesis in APA could come from a functional deviation of aldosterone biosynthesis from the normal zona glomerulosa and the utility of peripheral 18oxoF measurement could be influenced by the prevalence of KCNJ5 mutation in an APA.


Assuntos
Neoplasias do Córtex Suprarrenal/genética , Adenoma Adrenocortical/genética , Aldosterona/metabolismo , DNA de Neoplasias/genética , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/genética , Hidrocortisona/análogos & derivados , Mutação/genética , Neoplasias do Córtex Suprarrenal/metabolismo , Adenoma Adrenocortical/metabolismo , Análise Mutacional de DNA , Feminino , Canais de Potássio Corretores do Fluxo de Internalização Acoplados a Proteínas G/metabolismo , Humanos , Hidrocortisona/biossíntese , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
3.
Glia ; 67(5): 950-966, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30637802

RESUMO

Direct conversion is considered a promising approach to obtain tissue-specific cells for cell therapies; however, this strategy depends on exogenous gene expression that may cause undesired adverse effects such as tumorigenesis. By optimizing the Schwann cell induction system, which was originally developed for trans-differentiation of bone marrow mesenchymal stem cells into Schwann cells, we established a system to directly convert adult human skin fibroblasts into cells comparable to authentic human Schwann cells without gene introduction. Serial treatments with beta-mercaptoethanol, retinoic acid, and finally a cocktail of basic fibroblast growth factor, forskolin, platelet-derived growth factor-AA, and heregulin-ß1 (EGF domain) converted fibroblasts into cells expressing authentic Schwann cell markers at an efficiency of approximately 75%. Genome-wide gene expression analysis suggested the conversion of fibroblasts into the Schwann cell-lineage. Transplantation of induced Schwann cells into severed peripheral nerve of rats facilitated axonal regeneration and robust functional recovery in sciatic function index comparable to those of authentic human Schwann cells. The contributions of induced Schwann cells to myelination of regenerated axons and re-formation of neuromuscular junctions were also demonstrated. Our data clearly demonstrated that cells comparable to functional Schwann cells feasible for the treatment of neural disease can be induced from adult human skin fibroblasts without gene introduction. This direct conversion system will be beneficial for clinical applications to peripheral and central nervous system injuries and demyelinating diseases.


Assuntos
Diferenciação Celular/fisiologia , Fibroblastos/fisiologia , Traumatismos dos Nervos Periféricos/cirurgia , Recuperação de Função Fisiológica/fisiologia , Células de Schwann/fisiologia , Células de Schwann/transplante , Animais , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Modelos Animais de Doenças , Fibroblastos/efeitos dos fármacos , Proteínas de Fluorescência Verde/metabolismo , Humanos , Locomoção/fisiologia , Masculino , Microscopia Eletrônica , Proteína P0 da Mielina/metabolismo , Traumatismos dos Nervos Periféricos/fisiopatologia , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Fatores de Transcrição SOXE/metabolismo , Células de Schwann/ultraestrutura , Soro/fisiologia , Pele/citologia , Fatores de Tempo , Tretinoína/farmacologia
4.
Regen Ther ; 8: 15-19, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30271861

RESUMO

The 16th Congress of the Japanese Society for Regenerative Medicine was held from March 7-9, 2017, at Sendai International Center (Sendai city). The theme of this congress was "the renaissance of regenerative medicine" and it was an opportunity for information exchange between industry-leading researchers, doctors/dentists, and industry professionals. The objectives of the congress were to provide a place to promote and develop research in regenerative medicine. Numerous topics were covered in the 1 presidential lecture, 1 congress chair's lecture, 3 special lectures, the special symposia (2 sessions, 10 topics), symposia (41 sessions, 227 topics), evening symposia (3 sessions, 12 topics), joint symposium with another society (1 session, 4 topics), and presentations covering regular presentations including distinct presentations (oral presentations, 2 sessions, 8 topics), oral presentations (65 sessions, 383 topics), and poster presentations (44 sessions, 339 topics). There were co-organized seminars including 31 sessions for co-organized luncheon seminars, 2 sessions for co-organized evening seminars, and an up-to-date technology introduction corner, which hosted 153 organizations. Additionally, 4 special seminars and 3 hands-on training programs were hosted as part of the hands-on learning program for high school students during the congress. There were 3527 participants, and the event was a great success.

5.
Dev Growth Differ ; 60(6): 326-340, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29984494

RESUMO

Proliferation of ependymal cells of the adult spinal cord (SCEp cells) in the intact condition has been considered as a quite rare event. To visualize proliferating/proliferated SCEp cells, we used the intensive 5-bromo-2'-deoxyuridine (BrdU) administration method to find that about two cells in the ependymal layer incorporated BrdU in a 10-µm-thick section. Because these two cells were not considered to undergo further proliferation, we analyzed the positioning and motility of two neighboring BrdU-incorporated proliferated cells and elucidated the tendency of the movement of SCEp cells to the outer side inside the ependymal layer. Even if it was rare, one of the proliferated cells in the ependymal layer differentiated into astrocytes. Gene introduction of Notch intracellular domain (NICD), a constitutively active form of Notch1, into SCEp cells demonstrated both increase in proliferation and induction of differentiation into astrocytes. Overexpression of Sox2 promoted proliferation in SCEp cells. The reaction of gene introduction of NICD and Sox2 indicates the similarity of intracellular signaling between SCEp cells and neural stem cells. Also, considering the fact that SCEp cells express these two factors in the intact condition, Notch and Sox2 are important for the cell fate control of SCEp cells in the intact condition.


Assuntos
Astrócitos/metabolismo , Diferenciação Celular , Proliferação de Células , Epêndima/metabolismo , Células-Tronco Neurais/metabolismo , Transdução de Sinais/fisiologia , Animais , Antígenos de Diferenciação/biossíntese , Antígenos de Diferenciação/fisiologia , Astrócitos/citologia , Epêndima/citologia , Regulação da Expressão Gênica , Masculino , Células-Tronco Neurais/citologia , Ratos , Ratos Wistar , Fatores de Transcrição SOXB1
6.
Cell Transplant ; 27(2): 285-298, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29637816

RESUMO

Multilineage-differentiating stress-enduring (Muse) cells are endogenous nontumorigenic stem cells collectable as stage-specific embryonic antigen 3 (SSEA-3) + from various organs including the bone marrow and are pluripotent-like. The potential of human bone marrow-derived Muse cells to commit to cardiac lineage cells was evaluated. We found that (1) initial treatment of Muse cells with 5'-azacytidine in suspension culture successfully accelerated demethylation of cardiac marker Nkx2.5 promoter; (2) then transferring the cells onto adherent culture and treatment with early cardiac differentiation factors including wingless-int (Wnt)-3a, bone morphogenetic proteins (BMP)-2/4, and transforming growth factor (TGF) ß1; and (3) further treatment with late cardiac differentiation cytokines including cardiotrophin-1 converted Muse cells into cardiomyocyte-like cells that expressed α-actinin and troponin-I with a striation-like pattern. MLC2a expression in the final step suggested differentiation of the cells into an atrial subtype. MLC2v, a marker for a mature ventricular subtype, was expressed when cells were treated with Dickkopf-related protein 1 (DKK-1) and Noggin, inhibitors of Wnt3a and BMP-4, respectively, between steps (2) and (3). None of the steps included exogenous gene transfection, making induced cells feasible for future clinical application.


Assuntos
Miócitos Cardíacos/metabolismo , Células-Tronco Pluripotentes/metabolismo , Antígenos Glicosídicos Associados a Tumores/metabolismo , Proteína Morfogenética Óssea 2/metabolismo , Proteína Morfogenética Óssea 4/metabolismo , Diferenciação Celular/fisiologia , Células Cultivadas , Proteína Homeobox Nkx-2.5/metabolismo , Humanos , Miócitos Cardíacos/citologia , Células-Tronco Pluripotentes/citologia , Antígenos Embrionários Estágio-Específicos/metabolismo , Fatores de Crescimento Transformadores/metabolismo
7.
Brain Res ; 1675: 20-27, 2017 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-28870825

RESUMO

Ependymal cells have been considered one of prime targets for gene therapy in the central nervous system as they can secrete proteins directly into the cerebrospinal fluid. In this study, we have explored the probability of permanent exogenous gene expression using a combined adenovirus/transposon system. To this end, we created three adenoviruses; adenovirus #1 containing a CAG promoter-driven enhanced green fluorescent protein tagged with a palmitoylation site (palEGFP), whose DNA sequence was flanked by two different Tol2 ends, #2 containing a human FoxJ1 promoter-driven T2TP transposase, and #3 containing an EF-1 alpha promoter-driven T2TP transposase. We injected these adenoviruses into the lateral ventricles of adult rats to assess the duration of transgene expression, by which adenoviruses selectively infected to ependymal cells because they express the specific receptor. In animals injected with only adenovirus #1, we found palEGFP-expressing ependymal cells 1week after injection, but these cells had disappeared by 2weeks. In animals that received adenoviruses #1 and #2 in combination, despite detecting many palEGFP-expressing ependymal cells within the initial 2weeks, transgene expression in ependymal cells was almost disappeared 1month after injection. In contrast, many palEGFP-expressing astrocytes, oligodendrocytes, and neurons were found near the sites injected with adenoviruses #1 and #3, even 1month after injection. There was no prominent infiltration of immunological cells during the observation period. These findings indicate that an adenovirus-mediated transposon gene transfer system can lead to prolonged, but not permanent, expression of exogenous genes in ependymal cells of adult rats.


Assuntos
Adenoviridae/genética , Adenoviridae/metabolismo , Elementos de DNA Transponíveis/fisiologia , Epêndima/metabolismo , Técnicas de Transferência de Genes , Transgenes/fisiologia , Animais , Epêndima/citologia , Expressão Gênica , Masculino , Ratos , Ratos Wistar
8.
J Am Soc Nephrol ; 28(10): 2946-2960, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28674043

RESUMO

Multilineage-differentiating stress-enduring (Muse) cells are nontumorigenic endogenous pluripotent-like stem cells that can be collected from various organs. Intravenously administered Muse cells have been shown to spontaneously migrate to damaged tissue and replenish lost cells, but the effect in FSGS is unknown. We systemically administered human bone marrow-derived Muse cells without concurrent administration of immunosuppressants to severe combined immune-deficient (SCID) and BALB/c mouse models with adriamycin-induced FSGS (FSGS-SCID and FSGS-BALB/c, respectively). In FSGS-SCID mice, human Muse cells preferentially integrated into the damaged glomeruli and spontaneously differentiated into cells expressing markers of podocytes (podocin; 31%), mesangial cells (megsin; 13%), and endothelial cells (CD31; 41%) without fusing to the host cells; attenuated glomerular sclerosis and interstitial fibrosis; and induced the recovery of creatinine clearance at 7 weeks. Human Muse cells induced similar effects in FSGS-BALB/c mice at 5 weeks, despite xenotransplant without concurrent immunosuppressant administration, and led to improvement in urine protein, creatinine clearance, and plasma creatinine levels more impressive than that in the FSGS-SCID mice at 5 weeks. However, functional recovery in FSGS-BALB/c mice was impaired at 7 weeks due to immunorejection, suggesting the importance of Muse cell survival as glomerular cells in the FSGS kidney for tissue repair and functional recovery. In conclusion, Muse cells are unique reparative stem cells that preferentially home to damaged glomeruli and spontaneously differentiate into glomerular cells after systemic administration. Introduction of genes to induce differentiation is not required before Muse cell administration; thus, Muse cells may be a feasible therapeutic strategy in FSGS.


Assuntos
Glomerulosclerose Segmentar e Focal/terapia , Transplante de Células-Tronco , Animais , Diferenciação Celular , Movimento Celular , Doxorrubicina , Glomerulosclerose Segmentar e Focal/induzido quimicamente , Humanos , Testes de Função Renal , Camundongos Endogâmicos BALB C , Camundongos SCID , Regeneração
9.
Histochem Cell Biol ; 146(1): 45-57, 2016 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-26921198

RESUMO

We previously demonstrated that NG2-positive oligodendrocyte precursor cells (OPCs) do not express DM-20 mRNA and identified a distinct DM-20 mRNA-positive cell population expressing glutathione-S-transferase pi isoform (GST-pi) in the nucleus (GST-pi(Nuc)) of the adult rat spinal cord. As GST-pi intranuclear localization correlates with progenitor cell properties, we examined the differentiation status of this cell population under the intensive 5-bromo-2'-deoxyuridine (BrdU) administration method, consisting of intraperitoneal BrdU injections every 2 h for 48 h. We observed that a certain population of proliferating/proliferated cells expressed DM-20 mRNA, and sometimes two proliferating/proliferated cells were observed still attached to each other. We performed triple staining for BrdU, DM-20 mRNA, and NG2 and found pairs of neighboring BrdU-positive cells, which were considered to originate from the same progenitor cells and where both cells expressed DM-20 mRNA. Triple staining for BrdU, DM-20 mRNA, and GST-pi detected proliferating/proliferated cells exhibiting the GST-pi(Nuc)/DM-20 mRNA-positive expression pattern. These findings suggested the presence of a GST-pi(Nuc)/DM-20 mRNA-positive oligodendrocyte-lineage progenitor cell population in the adult rat spinal cord. However, we did not find any pair of neighboring BrdU-positive cells with this expression pattern. These observations collectively support the idea that GST-pi(Nuc)/DM-20 mRNA-expressing cells are the progeny of NG2-positive OPCs rather than a novel type of oligodendrocyte-lineage progenitor cells and that DM-20 mRNA expression is dynamically regulated during differentiation of OPCs into oligodendrocytes.


Assuntos
Diferenciação Celular , Glutationa S-Transferase pi/metabolismo , Proteína Proteolipídica de Mielina/genética , Oligodendroglia/citologia , Oligodendroglia/metabolismo , RNA Mensageiro/biossíntese , Medula Espinal/citologia , Animais , Masculino , Oligodendroglia/enzimologia , Ratos , Ratos Wistar , Medula Espinal/metabolismo
10.
Histochem Cell Biol ; 145(2): 147-61, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26563642

RESUMO

Proteolipid protein (PLP) is the major component of myelin; its gene encodes two major splicing variants: PLP and DM-20. Compared with PLP, DM-20 lacks the amino acids encoded by exon IIIb. The expression of PLP/DM-20 in cells outside the oligodendrocyte-lineage is unclear. To address this issue, we analyzed the detailed expression pattern of PLP/DM-20 mRNA in the adult rat spinal cord by in situ hybridization (ISH) with a cRNA probe complementary to DM-20 mRNA, which has been used to detect both PLP and DM-20 both mRNA. ISH did not label the cells expressing NeuN nor glial fibrillary acidic protein but detected those expressing Olig2, indicating that PLP/DM-20 mRNA are expressed only in oligodendrocyte-lineage cells. This cell population was expected to contain NG2-expressing oligodendrocyte precursor cells (OPCs), because some exhibited the expression of glutathione S-transferase pi isoform in the nucleus. A recent publication showed that OPCs express PLP but not DM-20 mRNA. However, no OPCs were detected. We performed ISH with a cRNA probe that specifically recognizes PLP mRNA to successfully detect some OPCs. Additionally, OPCs were detected by ISH with a cRNA probe complementary to DM-20 mRNA that was digested via alkaline hydrolysis prior to ISH. These findings collectively demonstrate that PLP and DM-20 mRNA expression is restricted to oligodendrocyte-lineage cells, and imply that the undigested cRNA probe complementary to the full-length DM-20 mRNA sequence only recognizes DM-20 mRNA and not the PLP counterpart when applied to ISH without denaturation/digestion methods.


Assuntos
Linhagem da Célula , Proteína Proteolipídica de Mielina/genética , Oligodendroglia/metabolismo , RNA Mensageiro/análise , Medula Espinal/citologia , Animais , Linhagem da Célula/genética , Hibridização In Situ , Masculino , RNA Mensageiro/genética , Ratos , Ratos Wistar
11.
Mol Cell Endocrinol ; 422: 57-63, 2016 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-26597777

RESUMO

INTRODUCTION: Cytochrome P450 11B2 (CYP11B2) plays a pivotal role in aldosterone synthesis, while cytochrome P450 11B1 (CYP11B1) and cytochrome P450 17A1 (CYP17) are involved in cortisol synthesis in normal human adrenal glands. However, their detailed distribution in aldosterone-producing adenoma (APA) remains incompletely settled. MATERIALS AND METHODS: We examined the status of CYP11B1/CYP11B2 and CYP11B2/CYP17A1 expressions in 27 APA (double staining) cases and 21 APA (triple staining) cases by using immunofluorescence staining and semi-quantitative evaluation. RESULTS: Tumor cells co-expressing CYP11B1/B2 (hybrid cell type A), CYP11B2/17 (hybrid cell type B), CYP11B1/17 (hybrid cell type C), and CYP11B1/B2/17 (triple-positive cell) were identified. The area and cell number of these cells were relatively small, but the size of individual hybrid cells were different between three hybrid cell types (A/B/C) and triple-positive cells. CONCLUSION: The presence of hybrid cells indicated the marked intratumoral heterogeneity of steroidogenesis in APAs, particularly in those producing glucocorticoids and mineralocorticoids.


Assuntos
Adenoma/metabolismo , Aldosterona/metabolismo , Citocromo P-450 CYP11B2/metabolismo , Esteroide 11-beta-Hidroxilase/metabolismo , Esteroide 17-alfa-Hidroxilase/metabolismo , Adenoma/genética , Corticosteroides/biossíntese , Tamanho Celular , Citocromo P-450 CYP11B2/genética , Imunofluorescência , Humanos , Células Híbridas/metabolismo , Mineralocorticoides/biossíntese , Esteroide 11-beta-Hidroxilase/genética , Esteroide 17-alfa-Hidroxilase/genética
12.
Nat Protoc ; 8(7): 1391-415, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23787896

RESUMO

Multilineage-differentiating stress-enduring (Muse) cells are distinct stem cells in mesenchymal cell populations with the capacity to self-renew, to differentiate into cells representative of all three germ layers from a single cell, and to repair damaged tissues by spontaneous differentiation into tissue-specific cells without forming teratomas. We describe step-by-step procedures for isolating and evaluating these cells. Muse cells are also a practical cell source for human induced pluripotent stem (iPS) cells with markedly high generation efficiency. They can be collected as cells that are double positive for stage-specific embryonic antigen-3 (SSEA-3) and CD105 from commercially available mesenchymal cells, such as adult human bone marrow stromal cells and dermal fibroblasts, or from fresh adult human bone marrow samples. Under both spontaneous and induced differentiation conditions, they show triploblastic differentiation. It takes 4-6 h to collect and 2 weeks to confirm the differentiation and self-renewal capacity of Muse cells.


Assuntos
Técnicas de Cultura de Células/métodos , Diferenciação Celular , Linhagem da Célula/fisiologia , Células-Tronco/citologia , Antígenos CD/metabolismo , Antígenos Glicosídicos Associados a Tumores/metabolismo , Células da Medula Óssea/citologia , Células Cultivadas , Endoglina , Citometria de Fluxo/métodos , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Mesoderma/citologia , Receptores de Superfície Celular/metabolismo , Antígenos Embrionários Estágio-Específicos/metabolismo
13.
J Clin Invest ; 123(1): 272-84, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-23202734

RESUMO

A cell-based therapy for the replacement of dopaminergic neurons has been a long-term goal in Parkinson's disease research. Here, we show that autologous engraftment of A9 dopaminergic neuron-like cells induced from mesenchymal stem cells (MSCs) leads to long-term survival of the cells and restoration of motor function in hemiparkinsonian macaques. Differentiated MSCs expressed markers of A9 dopaminergic neurons and released dopamine after depolarization in vitro. The differentiated autologous cells were engrafted in the affected portion of the striatum. Animals that received transplants showed modest and gradual improvements in motor behaviors. Positron emission tomography (PET) using [11C]-CFT, a ligand for the dopamine transporter (DAT), revealed a dramatic increase in DAT expression, with a subsequent exponential decline over a period of 7 months. Kinetic analysis of the PET findings revealed that DAT expression remained above baseline levels for over 7 months. Immunohistochemical evaluations at 9 months consistently demonstrated the existence of cells positive for DAT and other A9 dopaminergic neuron markers in the engrafted striatum. These data suggest that transplantation of differentiated autologous MSCs may represent a safe and effective cell therapy for Parkinson's disease.


Assuntos
Terapia Baseada em Transplante de Células e Tecidos/métodos , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/transplante , Células-Tronco Mesenquimais/metabolismo , Transtornos Parkinsonianos/terapia , Animais , Antígenos de Diferenciação/biossíntese , Corpo Estriado/diagnóstico por imagem , Corpo Estriado/metabolismo , Neurônios Dopaminérgicos/citologia , Regulação da Expressão Gênica , Integrina alfa6beta4/biossíntese , Macaca fascicularis , Masculino , Células-Tronco Mesenquimais/citologia , Transtornos Parkinsonianos/diagnóstico por imagem , Transtornos Parkinsonianos/metabolismo , Tomografia por Emissão de Pósitrons , Radiografia , Transplante Autólogo
14.
Cell Transplant ; 22(9): 1613-25, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23127893

RESUMO

After severe spinal cord injury, spontaneous functional recovery is limited. Numerous studies have demonstrated cell transplantation as a reliable therapeutic approach. However, it remains unknown whether grafted neuronal cells could replace lost neurons and reconstruct neuronal networks in the injured spinal cord. To address this issue, we transplanted bone marrow stromal cell-derived neural progenitor cells (BM-NPCs) in a rat model of complete spinal cord transection 9 days after the injury. BM-NPCs were induced from bone marrow stromal cells (BMSCs) by gene transfer of the Notch-1 intracellular domain followed by culturing in the neurosphere method. As reported previously, BM-NPCs differentiated into neuronal cells in a highly selective manner in vitro. We assessed hind limb movements of the animals weekly for 7 weeks to monitor functional recovery after local injection of BM-NPCs to the transected site. To test the sensory recovery, we performed functional magnetic resonance imaging (fMRI) using electrical stimulation of the hind limbs. In the injured spinal cord, transplanted BM-NPCs were confirmed to express neuronal markers 7 weeks following the transplantation. Grafted cells successfully extended neurites beyond the transected portion of the spinal cord. Adjacent localization of synaptophysin and PSD-95 in the transplanted cells suggested synaptic formations. These results indicated survival and successful differentiation of BM-NPCs in the severely injured spinal cord. Importantly, rats that received BM-NPCs demonstrated significant motor recovery when compared to the vehicle injection group. Volumes of the fMRI signals in somatosensory cortex were larger in the BM-NPC-grafted animals. However, neuronal activity was diverse and not confined to the original hind limb territory in the somatosensory cortex. Therefore, reconstruction of neuronal networks was not clearly confirmed. Our results indicated BM-NPCs as an effective method to deliver neuronal lineage cells in a severely injured spinal cord. However, reestablishment of neuronal networks in completed transected spinal cord was still a challenging task.


Assuntos
Transplante de Medula Óssea/métodos , Neurônios/transplante , Traumatismos da Medula Espinal/cirurgia , Células Estromais/transplante , Animais , Diferenciação Celular/fisiologia , Modelos Animais de Doenças , Feminino , Imagem por Ressonância Magnética , Regeneração Nervosa/fisiologia , Neurônios/citologia , Distribuição Aleatória , Ratos , Ratos Wistar , Recuperação de Função Fisiológica , Traumatismos da Medula Espinal/patologia , Transplante de Células-Tronco/métodos , Células Estromais/citologia
15.
Cytometry A ; 83(1): 18-26, 2013 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22693162

RESUMO

Induced pluripotent stem (iPS) cells have attracted a great deal of attention, although the mechanism by which they are generated is still not fully understood. Currently, two theories, the stochastic and elite models, have been proposed. Some reports provide theoretical support for the stochastic model. Other reports, however, support the elite model. For example, some human fibroblasts, such as Multilineage-differentiating stress enduring (Muse) cells, are reported to be pluripotent and a primary source of iPS cells. Thus, the mechanism of iPS cell generation continues to be debated. In this review, we discuss the properties of the original cell source, such as the components of the original populations and the potential of each population to become iPS cells, and further discuss the implications of the two theories for iPS cell research.


Assuntos
Diferenciação Celular , Modelos Estatísticos , Células-Tronco Pluripotentes/citologia , Proliferação de Células , Fibroblastos/citologia , Humanos , Células-Tronco Mesenquimais/citologia
16.
PLoS One ; 7(12): e48677, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23272044

RESUMO

Induced pluripotent stem (iPS) cells can be generated from somatic cells by the forced expression of four factors, Oct3/4, Sox2, Klf4, and c-Myc. While a great variety of colonies grow during induction, only a few of them develop into iPS cells. Researchers currently use visual observation to identify iPS cells and select colonies resembling embryonic stem (ES) cells, and there are no established objective criteria. Therefore, we exhaustively analyzed the morphology and gene expression of all the colonies generated from human fibroblasts after transfection with four retroviral vectors encoding individual factors (192 and 203 colonies in two experiments) and with a single polycistronic retroviral vector encoding all four factors (199 and 192 colonies in two experiments). Here we demonstrate that the morphologic features of emerged colonies can be categorized based on six parameters, and all generated colonies that could be passaged were classified into seven subtypes in colonies transfected with four retroviral vectors and six subtypes with a single polycistronic retroviral vector, both including iPS cell colonies. The essential qualifications for iPS cells were: cells with a single nucleolus; nucleus to nucleolus (N/Nls) ratio ∼2.19: cell size ∼43.5 µm(2): a nucleus to cytoplasm (N/C) ratio ∼0.87: cell density in a colony ∼5900 cells/mm(2): and number of cell layer single. Most importantly, gene expression analysis revealed for the first time that endogenous Sox2 and Cdx2 were expressed specifically in iPS cells, whereas Oct3/4 and Nanog, popularly used markers for identifying iPS cells, are expressed in colonies other than iPS cells, suggesting that Sox2 and Cdx2 are reliable markers for identifying iPS cells. Our findings indicate that morphologic parameters and the expression of endogenous Sox2 and Cdx2 can be used to accurately identify iPS cells.


Assuntos
Técnicas de Cultura de Células , Regulação da Expressão Gênica , Células-Tronco Pluripotentes Induzidas/citologia , Animais , Fator de Transcrição CDX2 , Nucléolo Celular/metabolismo , Núcleo Celular/metabolismo , Citoplasma/metabolismo , Células-Tronco Embrionárias/citologia , Fibroblastos/citologia , Fibroblastos/metabolismo , Perfilação da Expressão Gênica , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Reação em Cadeia da Polimerase/métodos , RNA/metabolismo , Retroviridae/genética , Fatores de Transcrição SOXB1/metabolismo
17.
Parkinsons Dis ; 2012: 873706, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22530164

RESUMO

Cell transplantation is a strategy with great potential for the treatment of Parkinson's disease, and many types of stem cells, including neural stem cells and embryonic stem cells, are considered candidates for transplantation therapy. Mesenchymal stem cells are a great therapeutic cell source because they are easy accessible and can be expanded from patients or donor mesenchymal tissues without posing serious ethical and technical problems. They have trophic effects for protecting damaged tissues as well as differentiation ability to generate a broad spectrum of cells, including dopamine neurons, which contribute to the replenishment of lost cells in Parkinson's disease. This paper focuses mainly on the potential of mesenchymal stem cells as a therapeutic cell source and discusses their potential clinical application in Parkinson's disease.

18.
Cell Mol Life Sci ; 69(22): 3739-50, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22527723

RESUMO

Induced pluripotent stem (iPS) cells have attracted a great deal attention as a new pluripotent stem cell type that can be generated from somatic cells, such as fibroblasts, by introducing the transcription factors Oct3/4, Sox2, Klf4, and c-Myc. The mechanism of generation, however, is not fully understood. Two mechanistic theories have been proposed; the stochastic model purports that every cell type has the potential to be reprogrammed to become an iPS cell and the elite model proposes that iPS cell generation occurs only from a subset of cells. Some reports have provided theoretical support for the stochastic model, but a recent publication demonstrated findings that support the elite model, and thus the mechanism of iPS cell generation remains under debate. To enhance our understanding of iPS cells, it is necessary to clarify the properties of the original cell source, i.e., the components of the original populations and the potential of each population to become iPS cells. In this review, we discuss the two theories and their implications in iPS cell research.


Assuntos
Células-Tronco Adultas/citologia , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/fisiologia , Animais , Diferenciação Celular , Linhagem Celular , Fibroblastos/metabolismo , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Camundongos , Modelos Biológicos , Fator 3 de Transcrição de Octâmero/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição SOXB1/metabolismo
19.
Anat Sci Int ; 87(1): 24-44, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22237924

RESUMO

Mesenchymal cell populations, referred to as mesenchymal stem cells or multipotent stromal cells (MSCs), which include bone marrow stromal cells (BMSCs), umbilical cord stromal cells and adipose stromal cells (ASCs), participate in tissue repair when transplanted into damaged or degenerating tissues. The trophic support and immunomodulation provided by MSCs can protect against tissue damage, and the differentiation potential of these cells may help to replace lost cells. MSCs are easily accessible and can be expanded on a large scale. In addition, BMSCs and ASCs can be harvested from the patient himself. Thus, MSCs are considered promising candidates for cell therapy. In this review, I will discuss recently discovered high-efficiency induction systems for deriving Schwann cells and neurons from MSCs. Other features of MSCs that are important for tissue repair include the self-renewing property of stem cells and their potential for differentiation. Thus, I will also discuss the stemness of MSCs and describe the discovery of a certain stem cell type among adult MSCs that can self-renew and differentiate into cells of all three germ layers. Furthermore, I will explore the prospects of using this cell population for cell therapy.


Assuntos
Diferenciação Celular/fisiologia , Transplante de Células-Tronco Mesenquimais/métodos , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Neurais/fisiologia , Neurônios/fisiologia , Células de Schwann/fisiologia , Animais , Indução Embrionária/fisiologia , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Neurais/citologia , Neurônios/citologia , Células de Schwann/citologia
20.
Methods Mol Biol ; 826: 89-102, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22167642

RESUMO

We have found a novel type of pluripotent stem cells, Multilineage-differentiating stress enduring (Muse) cells that can be isolated from mesenchymal cell populations. Muse cells are characterized by stress tolerance, expression of pluripotency markers, self-renewal, and the ability to differentiate into endodermal-, mesodermal-, and ectodermal-lineage cells from a single cell, demonstrating that they are pluripotent stem cells. They can be isolated as cells positive for stage-specific embryonic antigen-3, a human pluripotent stem cell marker. Here, we introduce the isolation method for Muse cells and the effect of transplantation of these cells on chronic liver diseases.


Assuntos
Diferenciação Celular/fisiologia , Separação Celular/métodos , Doença Hepática Terminal/terapia , Células-Tronco Mesenquimais/citologia , Células-Tronco Pluripotentes/citologia , Transplante de Células-Tronco/métodos , Adulto , Antígenos Glicosídicos Associados a Tumores/metabolismo , Citometria de Fluxo/métodos , Células HEK293 , Humanos , Imuno-Histoquímica , Lentivirus , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Antígenos Embrionários Estágio-Específicos/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA