Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 83
Filtrar
1.
J Orthop Res ; 38(2): 258-268, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31429977

RESUMO

Liver kinase B1 (LKB1), a serine/threonine protein, is a key regulator in stem cell function and energy metabolism. Herein, we describe the role of LKB1 in modulating the differentiation of synovium-derived stem cells (SDSCs) toward chondrogenic, adipogenic, and osteogenic lineages. Human fetal SDSCs were transduced with CRISPR associated protein 9 (Cas9)-single-guide RNA vectors to knockout or lentiviral vectors to overexpress the LKB1 gene. Analyses including ICE (Inference of CRISPR Edits) data from Sanger sequencing and quantitative polymerase chain reaction (qPCR) as well as Western blot demonstrated successful knockout (KO) or overexpression (OE) of LKB1 in human fetal SDSCs without any detectable side effects in morphology, proliferation rate, and cell cycle. LKB1 KO increased CD146 expression; interestingly, LKB1 OE increased SSEA4 level. The qPCR data showed that LKB1 KO upregulated the levels of SOX2 and NANOG while LKB1 OE lowered the expression of POU5F1 and KLF4. Furthermore, LKB1 KO enhanced, and LKB1 OE inhibited, chondrogenic and adipogenic differentiation potential. However, perhaps due to the inherent inability to achieve osteogenesis, LKB1 did not obviously affect osteogenic differentiation. These data demonstrate that LKB1 plays a significant role in determining human SDSCs' adipogenic and chondrogenic differentiation, which might provide an approach for fine-tuning the direction of stem cell differentiation in tissue engineering and regeneration. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:258-268, 2020.


Assuntos
Diferenciação Celular , Células-Tronco Fetais/fisiologia , Proteínas Serina-Treonina Quinases/metabolismo , Sequência de Bases , Antígeno CD146/metabolismo , Células Cultivadas , Vetores Genéticos , Humanos , Lentivirus , Proteínas Serina-Treonina Quinases/genética , Antígenos Embrionários Estágio-Específicos/metabolismo , Membrana Sinovial/citologia
2.
Mol Cell ; 76(2): 320-328, 2019 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-31563431

RESUMO

Germline cells are the beginning of new individuals in multicellular animals, including humans. Our understanding of these cell types is limited by the difficulty of analyzing the precious and heterogeneous germline tissue samples. The rapid development of single-cell sequencing technologies provides a chance for comprehensive profiling of the omics dynamics of human germline development. In this review, we discuss progress in analyzing the development of human germline cells, including preimplantation and implantation embryos, fetal germ cells (FGCs), and adult spermatogenesis by single-cell transcriptome and epigenome sequencing technologies.


Assuntos
Células-Tronco Fetais/fisiologia , Regulação da Expressão Gênica no Desenvolvimento , Óvulo/fisiologia , Análise de Sequência de DNA , Análise de Célula Única/métodos , Espermatozoides/fisiologia , Blastocisto/fisiologia , Montagem e Desmontagem da Cromatina , Metilação de DNA , Desenvolvimento Embrionário/genética , Epigênese Genética , Feminino , Genótipo , Humanos , Masculino , Fenótipo , Espermatogênese/genética
3.
Curr Neurovasc Res ; 16(3): 187-193, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31258084

RESUMO

BACKGROUND: Retinal degeneration and related eye disorders have limited treatment interventions. Since stem cell therapy has shown promising results, ciliary epithelium (CE) derived stem cells could be a better choice given the fact that cells from eye niche can better integrate with the degenerating retina, rewiring the synaptic damage. OBJECTIVE: To test the effect of human fetal pigmented ciliary epithelium-derived neurospheres in the mouse model of laser-induced retinal degeneration. METHODS: C57 male mice were subjected to retinal injury by Laser photocoagulation. Human fetal pigmented ciliary epithelium was obtained from post-aborted human eyeballs and cultured with epidermal growth factor (rhEGF) and fibroblast growth factor (rhFGF). The six day neurospheres were isolated, dissociated and transplanted into the subretinal space of the laser injured mice at the closest proximity to Laser shots. Mice were analyzed for functional vision through electroretinogram (ERG) and sacrificed at 1 week and 12 week time points. Retinal, Neurotropic, Apoptotic and proliferation markers were analysed using real-time polymerase chain reaction (PCR). RESULTS: The CE neurospheres showed an increase in the expression of candidate genes analyzed in the study at 1 week time point, which sustained for longer time point of 12 weeks. CONCLUSION: We showed the efficacy of human CE cells in the regeneration of retinal degeneration in murine model for the first time. CE cells need to be explored comprehensively both in disease and degeneration.


Assuntos
Células-Tronco Fetais/fisiologia , Lasers/efeitos adversos , Regeneração Nervosa/fisiologia , Degeneração Retiniana/terapia , Epitélio Pigmentado da Retina/fisiologia , Transplante de Células-Tronco/métodos , Animais , Células Cultivadas , Cílios/fisiologia , Cílios/transplante , Células Epiteliais/fisiologia , Células Epiteliais/transplante , Células-Tronco Fetais/química , Células-Tronco Fetais/transplante , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Degeneração Retiniana/etiologia , Degeneração Retiniana/patologia , Epitélio Pigmentado da Retina/transplante
4.
J Comp Neurol ; 527(18): 3023-3033, 2019 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-31173371

RESUMO

Human-induced pluripotent stem cells (hiPSCs) have facilitated studies on organ development and differentiation into specific lineages in in vitro systems. Although numerous studies have focused on cellular differentiation into neural lineage using hPSCs, most studies have initially evaluated embryoid body (EB) formation, eventually yielding terminally differentiated neurons with limited proliferation potential. This study aimed to establish human primitive neural stem cells (pNSCs) from exogene-free hiPSCs without EB formation. To derive pNSCs, we optimized N2B27 neural differentiation medium through supplementation of two inhibitors, CHIR99021 (GSK-3 inhibitor) and PD0325901 (MEK inhibitor), and growth factors including basic fibroblast growth factor (bFGF) and human leukemia inhibitory factor (hLIF). Consequently, pNSCs were efficiently derived and cultured over a long term. pNSCs displayed differentiation potential into neurons, astrocytes, and oligodendrocytes. These early NSC types potentially promote the clinical application of hiPSCs to cure human neurological disorders.


Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Fetais/fisiologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Células-Tronco Neurais/fisiologia , Linhagem da Célula/fisiologia , Células Cultivadas , Humanos
5.
Eur J Dermatol ; 29(6): 585-595, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31903948

RESUMO

The skin is the first protective barrier of our body. Wound healing is therefore an essential mechanism. However, this phenomenon can be impaired when wounds are too large or chronic, for example, in diabetes. Interestingly, adult skin heals with scars, whereas foetuses present scarless regeneration. The objective of this review is to highlight the difference in healing pathways between foetal and adult skin and to present the recent therapeutic strategies envisaged using foetal properties in the clinic. The main features that distinguish foetal wound healing from adult wound healing are less tissue inflammation, faster reepithelialisation, and less contraction of the neodermis, allowing foetal tissues to regenerate. Recently, new therapies in regenerative medicine have been introduced using these foetal properties. For the first time, our team has developed CICAFAST, an innovative dressing composed of foetal keratinocytes and fibroblasts, which has been tested on a skin graft donor site in a clinical Phase 1/2 trial.


Assuntos
Cicatriz/fisiopatologia , Feto/fisiologia , Pele/fisiopatologia , Transplante de Tecidos/métodos , Cicatrização/fisiologia , Ferimentos e Lesões/cirurgia , Adulto , Procedimentos Cirúrgicos Dermatológicos , Feminino , Células-Tronco Fetais/fisiologia , Transplante de Tecido Fetal , Fibroblastos/fisiologia , Humanos , Inflamação/fisiopatologia , Queratinócitos/fisiologia , Gravidez , Reepitelização/fisiologia , Fenômenos Fisiológicos da Pele , Transplante de Pele
6.
Curr Stem Cell Res Ther ; 13(7): 608-617, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30027853

RESUMO

Nerve injury is a large problem that produces much pain in patients. Injury to the nervous system causes serious consequences and affects a person's quality of life. The development of tissue engineering has created a brighter future for nerve regeneration, and research has not stopped since the discovery of stem cells. Stem cells are a type of pluripotent cell that exhibits the capacity of selfdifferentiation and proliferation. Many studies have demonstrated the ability of stem cells to differentiate into other types of cells, including neurons, after induction with trophic factors in vivo and in vitro. Scientists have isolated a variety of stem cells from different organs and tissues in the human body and demonstrated that these cells were efficacious in regenerative medicine. The use of these cells provides a non-surgical method for the treatment of neurological diseases, such as nerve defects. However, many problems must be resolved before using these cells in the clinical field. The microenvironment and delivery methods of cells also affect the regeneration process. The present article comprehensively summarizes the progress of stem cells in the field of nerve regeneration in the recent decades.


Assuntos
Regeneração Nervosa , Células-Tronco Neurais/citologia , Neurônios/citologia , Traumatismos dos Nervos Periféricos/terapia , Medicina Regenerativa/métodos , Engenharia Tecidual/métodos , Animais , Diferenciação Celular , Sistema Nervoso Central/citologia , Sistema Nervoso Central/lesões , Polpa Dentária/citologia , Polpa Dentária/fisiologia , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/fisiologia , Células-Tronco Fetais/citologia , Células-Tronco Fetais/fisiologia , Folículo Piloso/citologia , Folículo Piloso/fisiologia , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/fisiologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Neurais/fisiologia , Neurônios/fisiologia , Sistema Nervoso Periférico/citologia , Sistema Nervoso Periférico/lesões
7.
Stem Cell Rev Rep ; 14(5): 632-641, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29948753

RESUMO

Chimerism occurs naturaly throughout gestation and can also occur as a consequence of transfusion and transplantation therapy. It consists of the acquisition and long-term persistence of a genetically distinct population of allogenic cells inside another organism. Previous reports have suggested that feto-maternal microchimerism could exert a beneficial effect on the treatment of hematological and solid tumors in patients treated by PBSCT. In this review we report the mechanism of transplacental fetal stem cell trafficking during pregnancy and the effect of their long-term persistence on autoimmunity, GVHD, PBSCT, cancer and stem cell treatment.


Assuntos
Doenças Autoimunes/fisiopatologia , Quimerismo , Células-Tronco Fetais/fisiologia , Feminino , Células-Tronco Fetais/patologia , Células-Tronco Fetais/transplante , Feto/citologia , Feto/fisiologia , Humanos , Placenta/citologia , Placenta/fisiologia , Gravidez
8.
Exp Cell Res ; 370(1): 1-12, 2018 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-29883712

RESUMO

Understanding mechanisms in lineage differentiation is critical for organ development, pathophysiology and oncogenesis. To determine whether microRNAs (miRNA) may serve as drivers or adjuncts in hepatic differentiation, we studied human embryonic stem cell-derived hepatocytes and primary hepatocytes representing fetal or adult stages. Model systems were used for hepatic lineage advancement or regression under culture conditions with molecular assays. Profiles of miRNA in primary fetal and adult hepatocytes shared similarities and distinctions from pluripotent stem cells or stem cell-derived early fetal-like hepatocytes. During phenotypic regression in fetal or adult hepatocytes, miRNA profiles oscillated to regain stemness-associated features that had not been extinguished in stem cell-derived fetal-like hepatocytes. These oscillations in stemness-associated features were not altered in fetal-like hepatocytes by inhibitory mimics for dominantly-expressed miRNA, such as hsa-miR-99b, -100, -214 and -221/222. The stem cell-derived fetal-like hepatocytes were permissive for miRNA characterizing mature hepatocytes, including mimics for hsa-miR-122, -126, -192, -194 and -26b, although transfections of the latter did not advance hepatic differentiation. Examination of genome-wide mRNA expression profiles in stem cell-derived or primary fetal hepatocytes indicated targets of highly abundant miRNA regulated general processes, e.g., cell survival, growth and proliferation, functional maintenance, etc., without directing cell differentiation. Among upstream regulators of gene networks in stem cell-derived hepatocytes included HNF4A, SNAI1, and others, which affect transcriptional circuits directing lineage development or maintenance. Therefore, miRNA expression oscillated in response to microenvironmental conditions, whereas lineage-specific transcriptional regulators, such as HNF4A, were necessary for directing hepatic differentiation. This knowledge will be helpful for understanding the contribution of stem cells in pathophysiological states and oncogenesis, as well as for applications of stem cell-derived hepatocytes.


Assuntos
Diferenciação Celular/fisiologia , Células-Tronco Fetais/fisiologia , Hepatócitos/fisiologia , Fígado/fisiologia , MicroRNAs/genética , Células-Tronco Pluripotentes/fisiologia , Transcrição Genética/genética , Linhagem da Célula/genética , Células Cultivadas , Feto/fisiologia , Regulação da Expressão Gênica/genética , Células-Tronco Embrionárias Humanas/fisiologia , Humanos
9.
J Med Ethics ; 44(9): 606-610, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29491041

RESUMO

Organoids are three-dimensional biological structures grown in vitro from different kinds of stem cells that self-organise mimicking real organs with organ-specific cell types. Recently, researchers have managed to produce human organoids which have structural and functional properties very similar to those of different organs, such as the retina, the intestines, the kidneys, the pancreas, the liver and the inner ear. Organoids are considered a great resource for biomedical research, as they allow for a detailed study of the development and pathologies of human cells; they also make it possible to test new molecules on human tissue. Furthermore, organoids have helped research take a step forward in the field of personalised medicine and transplants. However, some ethical issues have arisen concerning the origin of the cells that are used to produce organoids (ie, human embryos) and their properties. In particular, there are new, relevant and so-far overlooked ethical questions concerning cerebral organoids. Scientists have created so-called mini-brains as developed as a few-months-old fetus, albeit smaller and with many structural and functional differences. However, cerebral organoids exhibit neural connections and electrical activity, raising the question whether they are or (which is more likely) will one day be somewhat sentient. In principle, this can be measured with some techniques that are already available (the Perturbational Complexity Index, a metric that is directly inspired by the main postulate of the Integrated Information Theory of consciousness), which are used for brain-injured non-communicating patients. If brain organoids were to show a glimpse of sensibility, an ethical discussion on their use in clinical research and practice would be necessary.


Assuntos
Encéfalo/fisiologia , Ética em Pesquisa , Organoides/fisiologia , Células-Tronco/fisiologia , Estado de Consciência/fisiologia , Células-Tronco Fetais/fisiologia , Humanos
10.
J Pediatr Surg ; 53(6): 1134-1136, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29580785

RESUMO

PURPOSE: We sought to examine donor mesenchymal stem cell (MSC) kinetics after transamniotic stem cell therapy (TRASCET) in experimental spina bifida. METHODS: Pregnant Sprague-Dawley dams exposed to retinoic acid for the induction of fetal neural tube defects received volume-matched intra-amniotic injections on gestational day 17 (E17; term=E22): either amniotic fluid MSCs (afMSCs) labeled with a luciferase reporter gene (n=78), or luciferase protein alone (n=66). Samples from twelve organ systems from each surviving fetus with spina bifida (total n=60) were screened via microplate luminometry at term. RESULTS: Donor afMSCs were identified exclusively in the placenta, umbilical cord, spleen, bone marrow, hip bones, defect, and brain. Luminometry was negative in control fetuses receiving luciferase alone (p<0.001). Signal intensity in relative light units (RLUs) was moderately correlated between the defect and the hip bones (rho=0.38, p=0.048), and between the placenta and the brain (rho=0.40, p=0.038). CONCLUSIONS: Amniotic mesenchymal stem cells engraft to specific sites after concentrated intra-amniotic injection in the setting of spina bifida. A hematogenous route encompassing the bone marrow as well as distant central nervous system homing are fundamental constituents of cell trafficking. These findings must be considered during eventual patient selection for transamniotic stem cell therapy in the prenatal management of spina bifida.


Assuntos
Movimento Celular , Células-Tronco Fetais/transplante , Terapias Fetais/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Disrafismo Espinal/terapia , Líquido Amniótico/citologia , Animais , Feminino , Células-Tronco Fetais/fisiologia , Células-Tronco Mesenquimais/fisiologia , Gravidez , Ratos Endogâmicos Lew , Ratos Sprague-Dawley , Disrafismo Espinal/embriologia
11.
J Neuropathol Exp Neurol ; 77(4): 325-343, 2018 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-29420729

RESUMO

When spinal roots are torn off from the spinal cord, both the peripheral and central nervous system get damaged. As the motoneurons lose their axons, they start to die rapidly, whereas target muscles atrophy due to the denervation. In this kind of complicated injury, different processes need to be targeted in the search for the best treatment strategy. In this study, we tested glial cell-derived neurotrophic factor (GDNF) treatment and fetal lumbar cell transplantation for their effectiveness to prevent motoneuron death and muscle atrophy after the spinal root avulsion and delayed reimplantation. Application of exogenous GDNF to injured spinal cord greatly prevented the motoneuron death and enhanced the regeneration and axonal sprouting, whereas no effect was seen on the functional recovery. In contrast, cell transplantation into the distal nerve did not affect the host motoneurons but instead mitigated the muscle atrophy. The combination of GDNF and cell graft reunited the positive effects resulting in better functional recovery and could therefore be considered as a promising strategy for nerve and spinal cord injuries that involve the avulsion of spinal roots.


Assuntos
Células-Tronco Fetais/transplante , Fator Neurotrófico Derivado de Linhagem de Célula Glial/uso terapêutico , Neurônios Motores/fisiologia , Recuperação de Função Fisiológica/efeitos dos fármacos , Traumatismos da Medula Espinal/tratamento farmacológico , Traumatismos da Medula Espinal/cirurgia , Animais , Sobrevivência Celular , Colina O-Acetiltransferase/metabolismo , Embrião de Mamíferos , Feminino , Células-Tronco Fetais/fisiologia , Asseio Animal/fisiologia , Proteínas de Homeodomínio/metabolismo , Músculo Esquelético/metabolismo , Músculo Esquelético/patologia , Bainha de Mielina/metabolismo , Regeneração Nervosa , Proteínas de Neurofilamentos/metabolismo , Ratos , Ratos Sprague-Dawley , Recuperação de Função Fisiológica/fisiologia , Reimplante , Medula Espinal/citologia , Tubulina (Proteína)/metabolismo
12.
Int Braz J Urol ; 44(3): 608-616, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29211403

RESUMO

PURPOSE: To identify the fetal stem cell (FSC) response to maternal renal injury with emphasis on renal integrity improvement and Y chromosome detection in damaged maternal kidney. MATERIALS AND METHODS: Eight non-green fluorescent protein (GFP) transgenic Sprague- Dawley rats were mated with GFP-positive transgenic male rats. Renal damage was induced on the right kidney at gestational day 11. The same procedure was performed in eight non-pregnant rats as control group. Three months after delivery, right nephrectomy was performed in order to evaluate the injured kidney. The fresh perfused kidneys were stained with anti-GFP antibody. Polymerase chain reaction (PCR) assay was also performed for the Y chromosome detection. Cell culture was performed to detect the GFP-positive cells. Technetium-99m-DMSA renal scan and single-photon emission computed tomography (SPECT) were performed after renal damage induction and 3 months later to evaluate the improvement of renal integrity. RESULTS: The presence of FSCs was confirmed by immune histochemical staining as well as immunofluorescent imaging of the damaged part. Gradient PCR of female rat purified DNA demonstrated the presence of Y-chromosome in the damaged maternal kidney. Moreover, the culture of kidney cells showed GPF- positive cells by immunofluorescence microscopy. The acute renal scar was repaired and the integrity of damaged kidney reached to near normal levels in experimental group as shown in DMSA scan. However, no significant improvement was observed in control group. CONCLUSION: FSC seems to be the main mechanism in repairing of the maternal renal injury during pregnancy as indicated by Y chromosome and GFP-positive cells in the sub-cultured medium.


Assuntos
Quimerismo , Células-Tronco Fetais/fisiologia , Nefropatias/fisiopatologia , Troca Materno-Fetal/fisiologia , Cicatrização/fisiologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Feminino , Imunofluorescência , Imuno-Histoquímica , Nefropatias/diagnóstico por imagem , Nefropatias/patologia , Masculino , Reação em Cadeia da Polimerase , Gravidez , Compostos Radiofarmacêuticos , Ratos Sprague-Dawley , Ácido Dimercaptossuccínico Tecnécio Tc 99m , Fatores de Tempo , Tomografia Computadorizada de Emissão de Fóton Único , Cromossomo Y
13.
Exp Neurol ; 291: 20-35, 2017 05.
Artigo em Inglês | MEDLINE | ID: mdl-28131724

RESUMO

Primary human fetal cells have been used in clinical trials of cell replacement therapy for the treatment of neurodegenerative disorders such as Huntington's disease (HD). However, human fetal primary cells are scarce and difficult to work with and so a renewable source of cells is sought. Human fetal neural stem cells (hfNSCs) can be generated from human fetal tissue, but little is known about the differences between hfNSCs obtained from different developmental stages and brain areas. In the present work we characterized hfNSCs, grown as neurospheres, obtained from three developmental stages: 4-5, 6-7 and 8-9weeks post conception (wpc) and four brain areas: forebrain, cortex, whole ganglionic eminence (WGE) and cerebellum. We observed that, as fetal brain development proceeds, the number of neural precursors is diminished and post-mitotic cells are increased. In turn, primary cells obtained from older embryos are more sensitive to the dissociation process, their viability is diminished and they present lower proliferation ratios compared to younger embryos. However, independently of the developmental stage of derivation proliferation ratios were very low in all cases. Improvements in the expansion rates were achieved by mechanical, instead of enzymatic, dissociation of neurospheres but not by changes in the seeding densities. Regardless of the developmental stage, neurosphere cultures presented large variability in the viability and proliferation rates during the initial 3-4 passages, but stabilized achieving significant expansion rates at passage 5 to 6. This was true also for all brain regions except cerebellar derived cultures that did not expand. Interestingly, the brain region of hfNSC derivation influences the expansion potential, being forebrain, cortex and WGE derived cells the most expandable compared to cerebellar. Short term expansion partially compromised the regional identity of cortical but not WGE cultures. Nevertheless, both expanded cultures were multipotent and kept the ability to differentiate to region specific mature neuronal phenotypes.


Assuntos
Diferenciação Celular/fisiologia , Proliferação de Células/fisiologia , Células-Tronco Fetais/citologia , Células-Tronco Neurais/citologia , Células-Tronco Neurais/fisiologia , Análise de Variância , Encéfalo/citologia , Encéfalo/embriologia , Sobrevivência Celular , Células Cultivadas , Células-Tronco Fetais/fisiologia , Feto , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Idade Gestacional , Humanos , Antígeno Ki-67 , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , RNA Mensageiro/metabolismo
14.
Methods Mol Biol ; 1506: 101-115, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-27830548

RESUMO

Liver transplantation is the only therapeutic treatment for patients with end-stage liver diseases. However, donor organ scarcity is the major limitation, and therefore, alternative strategies are urgently needed. The ultimate goal for successful cell-based therapies is the ability of transplanted cells to efficiently engraft and reconstitute injured liver mass. To evaluate the repopulation capacity of transplanted cells, it is essential to identify their specific characteristics, as well as to study the mechanism(s) Through which transplanted donor cells replace tissue mass in hepatic microenvironments, using well-established cell transplantation models. To date, rat fetal liver stem/progenitor cells represent the most efficient cell population to reconstitute the near-normal liver and the liver microenvironment with advanced fibrosis/cirrhosis, and therefore, can be used for developing strategies in engineering potential donor cells in the future that will be useful for clinical application in hepatic cell therapy.The present protocol describes the isolation of epithelial stem/progenitor cells derived from ED14/15 fetal livers of DPPIV+ F344 or F344-Tg(EGFP) F455/Rrrc rats, the immunohistochemical staining method to detect E-cadherin-positive epithelial cells within unfractionated cell isolates, their transplantation into different DPPIV- liver microenvironments (near-normal, retrorsine-treated, and TAA-induced fibrotic/cirrhotic liver), as well as detection methods to follow the fate of transplanted cells in the recipient liver (see Fig. 1).


Assuntos
Separação Celular/métodos , Células Epiteliais/transplante , Células-Tronco Fetais/transplante , Hepatócitos/transplante , Cirrose Hepática Experimental/cirurgia , Transplante de Células-Tronco/métodos , Animais , Caderinas/metabolismo , Diferenciação Celular/efeitos dos fármacos , Células Epiteliais/fisiologia , Feminino , Células-Tronco Fetais/fisiologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/fisiologia , Humanos , Imuno-Histoquímica/métodos , Fígado/citologia , Fígado/efeitos dos fármacos , Fígado/cirurgia , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/cirurgia , Cirrose Hepática Experimental/induzido quimicamente , Masculino , Alcaloides de Pirrolizidina/farmacologia , Ratos , Ratos Endogâmicos F344 , Tioacetamida/toxicidade
15.
Stem Cell Res Ther ; 7(1): 134, 2016 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-27612565

RESUMO

BACKGROUND: Distraction osteogenesis (DO) is one of the most dramatic reconstructive techniques for inducing bone regeneration, but it involves an undesirably long period for bone consolidation. Developing innovative approaches to enhance bone consolidation is a burning need. Human fetal mesenchymal stem cells (hFMSCs) have been shown to express more primitive developmental genes than those of human adult mesenchymal stem cells (hAMSCs), which is a preferable source for cell therapy and tissue regeneration. In the present study, we investigated the immunogenicity of using the human mesenchymal stem cell (MSC) secretome on rat cells, the effects of secretome on osteogenic differentiation of rat bone marrow-derived MSCs (rBMSCs), and the potential application of hFMSC secretome in promoting bone consolidation in a rat DO model. METHODS: Secretome was collected from MSC culture and was used to treat rBMSCs. Following secretome treatment, cell proliferation, alkaline phosphatase staining, Alizarin Red S staining, and mRNA expression of osteogenic differentiation-related genes (including ALP, Runx2, OCN, OPN, and Osx) in the rBMSCs were checked, as well as mixed rat peripheral blood lymphocyte reaction. hFMSC secretome was injected locally into the regenerates from the end of lengthening every 3 days in the rat DO model, until termination. The regenerates were subject to weekly x-rays, micro-computed tomography (µCT) and mechanical testing examination. The bone quality was assessed by histology and immunohistochemistry examinations. RESULTS: Compared to the secretome from rBMSCs and hAMSCs, hFMSC secretome had the best osteogenic induction ability and low immunogenicity. hFMSC secretome with different doses showed no effect on cell viability. hFMSC secretome at the dose of 100 µg/µl could significantly increase the expression of alkaline phosphatase and all the osteogenic marker genes, as well as the amount of calcium deposits in the rBMSCs. Finally, the local application of hFMSC secretome in distraction regenerates in a rat DO model significantly improved bone consolidation according to the results of µCT, mechanical test, and histological and immunohistochemistry analysis. CONCLUSIONS: The current study demonstrated that hFMSC secretome promotes osteogenesis of rBMSCs and bone consolidation during DO. hFMSC secretome may be a new therapeutic strategy to enhance bone consolidation in patients undergoing DO treatment.


Assuntos
Regeneração Óssea/fisiologia , Osso e Ossos/metabolismo , Osso e Ossos/fisiologia , Células-Tronco Fetais/metabolismo , Feto/metabolismo , Células-Tronco Mesenquimais/metabolismo , Fosfatase Alcalina/metabolismo , Animais , Células da Medula Óssea/metabolismo , Células da Medula Óssea/fisiologia , Diferenciação Celular/fisiologia , Proliferação de Células , Terapia Baseada em Transplante de Células e Tecidos/métodos , Células Cultivadas , Células-Tronco Fetais/fisiologia , Feto/fisiologia , Humanos , Masculino , Células-Tronco Mesenquimais/fisiologia , Osteogênese por Distração/métodos , Ratos , Ratos Sprague-Dawley
16.
Kidney Int ; 90(2): 289-299, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27234568

RESUMO

The treatment of renal failure has changed little in decades. Organ transplantation and dialysis continue to represent the only therapeutic options available. However, decades of fundamental research into the response of the kidney to acute injury and the processes driving progression to chronic kidney disease are beginning to open doors to new options. Similarly, continued investigations into the cellular and molecular basis of normal kidney development, together with major advances in stem cell biology, are now delivering options in regenerative medicine not possible as recently as a decade ago. In this review, we will discuss advances in regenerative medicine as it may be applied to the kidney. This will cover cellular therapies focused on ameliorating injury and improving repair as well as advancements in the generation of new renal tissue from stem/progenitor cells.


Assuntos
Terapia Baseada em Transplante de Células e Tecidos/métodos , Nefropatias/terapia , Rim/fisiologia , Regeneração , Transplante de Células-Tronco , Animais , Reprogramação Celular , Células-Tronco Fetais/fisiologia , Humanos , Rim/citologia , Células-Tronco Mesenquimais/fisiologia , Cicatrização
17.
Rev. Ateneo Argent. Odontol ; 55(1): 69-70, 2016. ilus
Artigo em Espanhol | LILACS | ID: lil-794296

RESUMO

La utilización de células indiferenciadas embrionarias y de células diferenciadas inducidas para que se comporten como las anteriores permite dar origen adiferentes tejidos que pueden ser usados en medicina reconstructiva en reemplazo de los deteriorados...


Assuntos
Humanos , Células-Tronco Multipotentes/fisiologia , Células-Tronco Pluripotentes/fisiologia , Células-Tronco Totipotentes/fisiologia , Células-Tronco/fisiologia , Procedimentos Cirúrgicos Reconstrutivos/métodos , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Fetais/fisiologia , Engenharia Tecidual/métodos
18.
Stem Cell Res Ther ; 6: 251, 2015 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-26684339

RESUMO

BACKGROUND: Adult skeletal stem cells (SSCs) often exhibit limited in vitro expansion with undesirable phenotypic changes and loss of differentiation capacity. Foetal tissues offer an alternative cell source, providing SSCs which exhibit desirable differentiation capacity over prolonged periods, ideal for extensive in vitro and ex vivo investigation of fundamental bone biology and skeletal development. METHODS: We have examined the derivation of distinct cell populations from human foetal femora. Regionally isolated populations including epiphyseal and diaphyseal cells were carefully dissected. Expression of the SSC marker Stro-1 was also found in human foetal femora over a range of developmental stages and subsequently utilised for immuno-selection. RESULTS: Regional populations exhibited chondrogenic (epiphyseal) and osteogenic (diaphyseal) phenotypes following in vitro and ex vivo characterisation and molecular analysis, indicative of native SSC maturation during skeletal development. However, each population exhibited potential for induced multi-lineage differentiation towards bone (bone nodule formation), cartilage (proteoglycan and mucopolysaccharide deposition) and fat (lipid deposition), suggesting the presence of a shared stem cell sub-population. This shared sub-population may be comprised of Stro-1+ cells, which were later identified and immuno-selected from whole foetal femora exhibiting multi-lineage differentiation capacity in vitro and ex vivo. CONCLUSIONS: Distinct populations were isolated from human foetal femora expressing osteochondral differentiation capacity. Stro-1 immuno-selected SSCs were isolated from whole femora expressing desirable multi-lineage differentiation capacity over prolonged in vitro expansion, superior to their adult-derived counterparts, providing a valuable cell source with which to study bone biology and skeletal development.


Assuntos
Células-Tronco Fetais/citologia , Mioblastos Esqueléticos/citologia , Adipogenia , Animais , Antígenos de Superfície/metabolismo , Regeneração Óssea , Diferenciação Celular , Separação Celular , Embrião de Galinha , Condrogênese , Diáfises/citologia , Epífises/citologia , Fêmur/citologia , Fêmur/embriologia , Células-Tronco Fetais/fisiologia , Feto/citologia , Humanos , Técnicas In Vitro , Mioblastos Esqueléticos/fisiologia , Osteogênese
19.
Methods Mol Biol ; 1340: 25-40, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26445828

RESUMO

As the only cell type found in healthy adult cartilage, chondrocytes are the obvious and most direct starting point for cartilage tissue engineering. Human adult, juvenile, neonatal, and fetal chondrocytes have all been demonstrated to produce cartilage matrix components in vitro for production of engineered tissues. In this chapter, procedures are outlined for isolation of chondrocytes from human fetal and adult cartilage. Methods for expansion and cryopreservation of the cells and characterization of gene expression using quantitative polymerase chain reaction (Q-PCR) analysis are also described.


Assuntos
Células-Tronco Adultas/fisiologia , Técnicas de Cultura de Células , Diferenciação Celular , Condrócitos/fisiologia , Células-Tronco Fetais/fisiologia , Medicina Regenerativa/métodos , Engenharia Tecidual/métodos , Proliferação de Células , Separação Celular , Células Cultivadas , Condrogênese , Criopreservação , Regulação da Expressão Gênica , Humanos , Fenótipo , Reação em Cadeia da Polimerase em Tempo Real
20.
Neuroscience ; 289: 123-33, 2015 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-25595970

RESUMO

Fetal striatal transplantation has emerged as a new therapeutic strategy in Huntington's disease (HD). Hypoxia is one of the microenvironmental stress conditions to which fetal tissue is exposed as soon as it is isolated and transplanted into the diseased host brain. Mechanisms that support neuroblast survival and replenishment of damaged cells within the HD brain in the hypoxic condition have yet to be fully elucidated. This study is aimed at investigating the molecular pathways associated with the hypoxic condition in human fetal striatal neuroblasts (human striatal precursor (HSP) cells), using the hypoxia-mimetic agent cobalt chloride (CoCl2). We analyzed the effect of CoCl2 on HSP cell proliferation and on the expression of hypoxia-related proteins, such as hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF). Moreover, we evaluated fibroblast growth factor 2 (FGF2; 50ng/ml) and endothelin-1 (ET-1; 100nM) proliferative/survival effects in HSP cells in normoxic and hypoxic conditions. Dose-response experiments using increasing concentrations of CoCl2 (50-750µM) showed that the HSP cell growth was unaffected after 24h, while it increased at 48h, with the maximal effect observed at 400µM. In contrast, cell survival was impaired at 72h. Hypoxic conditions determined HIF-1α protein accumulation and increased gene and protein expression of VEGF, while FGF2 and ET-1 significantly stimulated HSP cell proliferation both in normoxic and hypoxic conditions, thus counteracting the apoptotic CoCl2 effect at 72h. The incubation with selective receptor (FGFR1, endothelin receptor A (ETA) and endothelin receptor B (ETB)) inhibitors abolished the FGF2 and ET-1 neuroprotective effect. In particular, ET-1 stimulated HSP cell survival through ETA in normoxic conditions and through ETB during hypoxia. Accordingly, ETA expression was down-regulated, while ETB expression was up-regulated by CoCl2 treatment. Overall, our results support the idea that HSP cells possess the machinery for their adaptation to hypoxic conditions and that neurotrophic factors, such as FGF2 and ET-1, may sustain neurogenesis and long-term survival through complex receptor-mediated mechanisms.


Assuntos
Hipóxia Celular/fisiologia , Corpo Estriado/fisiopatologia , Endotelina-1/metabolismo , Células-Tronco Fetais/fisiologia , Fatores de Crescimento de Fibroblastos/metabolismo , Células-Tronco Neurais/fisiologia , Hipóxia Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Fármacos do Sistema Nervoso Central/toxicidade , Cobalto/toxicidade , Relação Dose-Resposta a Droga , Fator 2 de Crescimento de Fibroblastos/metabolismo , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , RNA Mensageiro/metabolismo , Receptor de Endotelina A/metabolismo , Receptor de Endotelina B/metabolismo , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...