RESUMO
Comparison of mesenchymal stromal cells of embryonic and adult rat spleen showed that splenic cells from 20-day rat fetuses exhibit the capacity for clonal growth, express surface antigens CD73, CD90, and CD106, and have weak osteogenic and adipogenic potencies, while splenic cells from adult animals are characterized by lower cloning efficiency, rapid decrease of proliferative activity during passaging, the absence of CD73 and CD90 expression, and are incapable of osteogenesis The observed changes are probably related to extinction of myelopoiesis in the spleen during the postnatal ontogeny.
Assuntos
Diferenciação Celular , Proliferação de Células , Células-Tronco Mesenquimais/fisiologia , Baço/citologia , Animais , Antígenos CD/metabolismo , Forma Celular , Células Cultivadas , Feminino , Feto/citologia , Osteogênese , Fenótipo , Gravidez , Ratos WistarRESUMO
Fetal liver is known as a source of multipotent mesenchymal stromal cells. These cells are routinely isolated by adhesion to plastic, but thus prepared culture is contaminated by other cells. For instance, primary cell culture of from rat fetal liver, apart from fibroblasts with phenotypic characteristics of mesenchymal stromal cells, contained skeletal muscle precursors, myofibroblasts, and epitheliocytes expressing cytokeratin-19 (the latter was also detected in some fibroblast-like cells probably undergoing epithelio-mesenchymal transition). During passaging, fibroblasts become practically the only cell type in the culture.
Assuntos
Feto/citologia , Fígado/citologia , Animais , Células Cultivadas , Feminino , Fibroblastos/citologia , Miofibroblastos/citologia , Gravidez , RatosRESUMO
Hematopoietic differentiation and formation of hepatic tissue both take place in mammalian liver during its prenatal development. Hematopoietic and hepatic stem cells self-renew, proliferate and differentiate within specific microenvironment that is organized by stromal elements. Stroma of developing liver consists of different cell populations such as mesenchymal stromal cells, Ito cells, portal fibroblasts and myofibroblasts, vascular endothelial and smooth muscle cells, cells undergoing epithelial-to-mesenchymal transition. In this review, their phenotypical and functional properties, possible derivation and role in the regulation of hematopoiesis and hepatogenesis are discussed.
Assuntos
Hematopoese/fisiologia , Fígado , Animais , Diferenciação Celular , Proliferação de Células , Células Endoteliais/citologia , Células Endoteliais/fisiologia , Transição Epitelial-Mesenquimal , Feto , Fibroblastos/citologia , Fibroblastos/fisiologia , Células Estreladas do Fígado/citologia , Células Estreladas do Fígado/fisiologia , Humanos , Fígado/citologia , Fígado/crescimento & desenvolvimento , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Camundongos , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/fisiologia , Miofibroblastos/citologia , Miofibroblastos/fisiologiaRESUMO
Fetal liver stroma consists of different cell populations that are studied insufficiently. We have found skeletal muscle precursors that express MyoD in liver of 17 day and 20 day rat fetuses. Spontaneous myotube formation was observed in primary cultures of liver cells of 15, 17 and 20 day fetuses. Estimating antigenic profiles of these myogenic elements by immunocytochemistry and PCR methods unambiguously indicated their skeletal muscle nature. Comparative study of major myogenic gene expression demonstrated dependence of the myogenic potencies of liver cells on both the stage of fetal development and the duration of cell cultivation. It was shown that fetal liver MSCs were capable of myotube formation in the induction medium with 5-azacitidin. The results of our study, thereby, indicate that 15-20 day prenatal rat liver contains pre-existing skeletal muscle precursors expressing MyoD and, probably, inducible muscle precursors.
Assuntos
Desmina/metabolismo , Fígado/embriologia , Células-Tronco Mesenquimais/citologia , Desenvolvimento Muscular , Fibras Musculares Esqueléticas/citologia , Proteína MyoD/metabolismo , Miogenina/metabolismo , Animais , Azacitidina/farmacologia , Diferenciação Celular , Desmina/genética , Feminino , Desenvolvimento Fetal/fisiologia , Feto , Expressão Gênica , Imuno-Histoquímica , Fígado/citologia , Fígado/metabolismo , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Desenvolvimento Muscular/efeitos dos fármacos , Fibras Musculares Esqueléticas/metabolismo , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Proteína MyoD/genética , Miogenina/genética , Gravidez , Cultura Primária de Células/métodos , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Ratos , Ratos Wistar , Fatores de TempoRESUMO
Fetal liver during period of its hematopoietic activity contains mesenchymal stromal cells (MSC) that are known to play a major role in establishing hematopoietic microenvironment. These cells are capable of clonal growing and multilineage differentiation, but only limited data exist about changes in their properties during prenatal development. We compared cloning efficiency of MSC from liver of 14, 16 and 20 day rat fetuses and evaluated their potentials to in vitro osteo- and adipogenesis and in vivo chondrogenesis after whole organ ectopic transplantation. Content of clonogenic MSC in suspension of liver cells was maximal in 16 day fetuses and to a lesser extent in 20 day ones. MSC derived from 16 day fetuses demonstrated maximal potential to estimated lineages. Osteogenic potential of MSC from 14 day fetuses was comparable to whereas their adipogenic and chondrogenic abilities were inferior to that from 16 day fetuses. Cells from 20 day fetuses had only weak adipogenic potency and failed to differentiate into osteogenic of chondrogenic pathways. The results indicate that both number and differentiation potential of MSC in developing rat liver correlate with dynamics of hematopoiesis in this organ. Detected changes may be ascribed to the decline of hematopoiesis in liver and acquisition its definitive functions.
Assuntos
Adipogenia , Condrogênese , Desenvolvimento Fetal/fisiologia , Transplante de Fígado/métodos , Fígado/embriologia , Células-Tronco Mesenquimais/citologia , Osteogênese , Animais , Diferenciação Celular , Linhagem da Célula , Coristoma , Feminino , Feto , Hematopoese/fisiologia , Imuno-Histoquímica , Rim , Fígado/citologia , Células-Tronco Mesenquimais/metabolismo , Gravidez , Cultura Primária de Células/métodos , Ratos , Ratos Wistar , Fatores de TempoRESUMO
The action of three growth factors (EGF, bFGF, and PDGF) on mesenchymal stromal cell (MSC) subpopulations from mature bone marrow (BM) and rat embryo liver (EL) was investigated. These cells are plastic-adhesive and have different rates of adhesion (AC1-AC4 subpopulations). The efficiency of colony-formation, the size of colonies, and the number of early osteogenic progenitors with alkaline phosphatase activity in colonies and induced osteogenesis were analyzed. It was shown that EGF increased the number of bone marrow (BM) MSC colonies, but it had no influence on osteogenic differentiation. bFGF suppressed colony formation, but it stimulated both early and late stages of steogenesis. PDGF increased the size and the number of colonies in AC2 and AC3 subpopulations, but it stimulated only the ostegenesis terminal stage. The distinction between MSC subpopulations from two organs were found: MSC from EL had small osteogenic capacities and low sensitivity to grow factors; MSC from BM had no such characteristics. MSC subpopulations with different adhesion properties and from different tissues had compatible sensitivity to growth factors. Thus, these cells have no parent-progeny relationship.
Assuntos
Medula Óssea/efeitos dos fármacos , Fator de Crescimento Epidérmico/farmacologia , Fator 2 de Crescimento de Fibroblastos/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Fator de Crescimento Derivado de Plaquetas/farmacologia , Células Estromais/efeitos dos fármacos , Animais , Adesão Celular/efeitos dos fármacos , Adesão Celular/fisiologia , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/fisiologia , Células Cultivadas , Ensaio de Unidades Formadoras de Colônias , Fator de Crescimento Epidérmico/fisiologia , Feminino , Fator 2 de Crescimento de Fibroblastos/fisiologia , Fígado/citologia , Fígado/embriologia , Células-Tronco Mesenquimais/citologia , Osteogênese/fisiologia , Fator de Crescimento Derivado de Plaquetas/fisiologia , Ratos , Ratos Wistar , Proteínas Recombinantes/farmacologia , Células Estromais/citologiaRESUMO
Interactions with extracellular matrix including fibronectin (Fn) play an important role in regulation of cell growth and differentiation. Influence of Fn and its individual domains on adhesion and osteogenic potencies of rat mesenchymal stromal cells (MSCs) was estimated. Investigation of bone marrow or fetal liver MSCs adhesion dynamics showed that after 7 days of cultivation on Fn the number of adhered clonogenic cells derived from both sources was comparable to their number observed on plastic but their content in suspension was commonly decreased. Population of fetal liver MSCs differed from bone marrow-derived population by greater fraction of cells that adhered for the first 7 days. Bone marrow MSC cultures on Fn were characterized by reduced activity of alkaline phosphatase as compared with cultivation on plastic; furthermore, they deposed significantly smaller amount of calcium salts under cultivation in osteogenic medium. Cultivation of MSCs on Fn fragments demonstrated the primary role of its cell-binding domain in the inhibition of osteogenesis.
Assuntos
Fibronectinas/fisiologia , Fígado/fisiologia , Mesoderma/fisiologia , Osteogênese , Animais , Células da Medula Óssea/fisiologia , Adesão Celular/efeitos dos fármacos , Técnicas de Cultura de Células , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células , Células Cultivadas , Feminino , Feto/citologia , Fibronectinas/química , Fibronectinas/farmacologia , Humanos , Fígado/citologia , Mesoderma/citologia , Estrutura Terciária de Proteína , Ratos , Ratos Wistar , Células Estromais/fisiologiaRESUMO
Comparative characteristics of mesenchymal stromal cells (MSC) from fetal liver and adult bone marrow are of great interest due to similar contribution to organization of the hematopoietic microenvironments in these organs in different developmental periods. It is known that MSC play a pivotal role in organization of hematopoietic stem cells "niches". Moreover, the histogenetic affinity between MSC from both sources is not ruled out. Antigen profiling using immunocytochemistry and RT-PCR confirmed that cell populations under study corresponded to MSC criteria and had no hematopoietic, lymphoid and endothelial cells after second passage. Comparative analysis of osteogenic and adipogenic marker expression revealed distinct differentiation potentials of MSC from two hematopoietic organs: adipogenic potentials of hepatic MSC was weaker but ability to differentiate to osteogenic lineage was extremely low, if any, in comparison with marked osteo- and adipogenic potentials of adult bone marrow MSC. Similar cell phenotype but unequal differentiation potentials under identical conditions in vitro may be attributed to a distinct developmental program in pre- and postnatal histogenesis of MSC from different sources.
Assuntos
Adipogenia , Antígenos CD/biossíntese , Células da Medula Óssea/fisiologia , Fígado/citologia , Células-Tronco Mesenquimais/fisiologia , Osteogênese , Células-Tronco Adultas/fisiologia , Animais , Antígenos CD/análise , Antígenos CD/genética , Biomarcadores , Células Cultivadas , Células-Tronco Fetais/fisiologia , Feto/citologia , Imuno-Histoquímica , Reação em Cadeia da Polimerase , Análise Serial de Proteínas , RatosRESUMO
The review summarizes current concepts concerning the molecular genetic mechanisms of the osteogenic differentiation of mesenchymal stromal cells, which is controlled by a complex of signaling proteins and transcription factors. The interaction of regulatory factors involved in the most important signaling pathways at different stages of this differentiation is discussed.
Assuntos
Diferenciação Celular/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Osteogênese/fisiologia , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Animais , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/genética , Células Estromais/citologia , Células Estromais/fisiologia , Fatores de Transcrição/genéticaRESUMO
Multipotent bone marrow mesenchymal stromal cells are progenitors of various cell types capable of long-term self-maintenance. These cells are an adequate model for studying the most important problems in cell biology, such as self-maintenance of stem cells and regulation of their differentiation. Moreover, these cells are a promising resource for regenerative medicine. In this context, isolation of the earliest multipotent mesenchymal stromal cells, their in vitro maintenance in an undifferentiated state, and stimulation of their differentiation in a desired direction appear to be most important. To successfully use the multipotent mesenchymal stromal cells both in fundamental studies and in therapy, it is necessary to modify and standardize the composition of culture medium, replacing blood serum with certain growth factors. These factors have influence on the proliferation and differentiation of most cell types, including multipotent mesenchymal stromal cells. This paper is a review of available data concerning the effects of some growth factors on the multipotent mesenchymal stromal cells of the bone marrow.
Assuntos
Células da Medula Óssea/citologia , Diferenciação Celular/fisiologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Células-Tronco Mesenquimais/citologia , Animais , Antígenos de Diferenciação/metabolismo , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/fisiologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/fisiologiaRESUMO
The mesenchymal stromal cell is a multipotent precursor of osteoblasts, adipocytes, and some other cell types. In this study, a comparative analysis of cultured mesenchymal stromal cells from the rat bone marrow at the early and late stages of subculturing has been performed using molecular genetic and cytological methods. The culture has undergone 11 passages during 140 days. Upon long-term culturing, the mesenchymal stromal cells have proved to lose their potential for adipogenic differentiation but preserve the potential for osteogenesis. Morphological characters typical of osteogenic differentiation can be observed at the earlier stages of culturing (passages 1-4) but disappear at later stages (passages 9-11), despite mineralization of the extracellular matrix and the expression of osteogenic differentiation markers. A comparative analysis of the proliferation potential of stromal cells has shown that differences in the period of cell population doubling at the early and later stages of culturing are insignificant. An almost complete arrest of cell growth has been observed in the middle of the culture period (passages 5 and 6).
Assuntos
Células da Medula Óssea/citologia , Células-Tronco Mesenquimais/citologia , Células-Tronco Multipotentes/citologia , Adipócitos/citologia , Adipócitos/fisiologia , Animais , Antígenos de Diferenciação/biossíntese , Células da Medula Óssea/fisiologia , Diferenciação Celular/fisiologia , Proliferação de Células , Células Cultivadas , Células-Tronco Mesenquimais/fisiologia , Células-Tronco Multipotentes/fisiologia , Osteoblastos/citologia , Osteoblastos/fisiologia , Ratos , Ratos Wistar , Células Estromais/citologia , Células Estromais/fisiologia , Fatores de TempoRESUMO
Mesenchymal stem cells present in the bone marrow and some other organs are primitive pluripotent precursors of osseous, cartilaginous, adipose, and other mesenchymal tissues. The recently revealed capacity of these cells for differentiation into nonmesenchymal derivatives is of considerable theoretical and practical interest. However, many aspects of the biology of these cells remain obscure despite active research. This review considers possible sources and methods for the isolation of mesenchymal stem cells, their potential for proliferation and differentiation in different directions, and outlooks of their therapeutic application. A model of parent-progeny relationships of stromal cells is proposed, and the problems of regulation of proliferation and differentiation of mesenchymal precursors as well as their role in the maintenance of regeneration and tissue functioning are discussed.
Assuntos
Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Animais , Antígenos/análise , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Proliferação de Células , Perfilação da Expressão Gênica , Humanos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/química , FenótipoRESUMO
The sensitivity of stromal stem cells (CFU-f) from rat bone marrow and fetal liver to the cytotoxic effect of 5-fluorouracil (5-FU) was compared in vivo and in vitro. Cells from both tissues demonstrated a similar resistance to 5-FU in vitro; however, stromal stem cells from fetal liver proved notably more sensitive to 5-FU compared to marrow CFU-f in vivo. Cells forming colonies of different size were identified in stem cell populations from both tissues. Cells giving rise to small colonies had a higher resistance to 5-FU both in vivo and in vitro.
Assuntos
Antimetabólitos/farmacologia , Medula Óssea/metabolismo , Resistência a Medicamentos/fisiologia , Fluoruracila/farmacologia , Fígado/metabolismo , Células-Tronco/metabolismo , Animais , Células Cultivadas , Ensaio de Unidades Formadoras de Colônias/métodos , Resistência a Medicamentos/efeitos dos fármacos , Feminino , Feto/citologia , Feto/metabolismo , Fígado/citologia , Ratos , Ratos Wistar , Células-Tronco/citologia , Células Estromais/citologia , Células Estromais/metabolismoRESUMO
Effect of alkylating agent dipin was studied on hematopoietic (CFU-S) and stromal (CFU-F) progenitor cells. Single administration of dipin (0.06 mg/g) to adult (CBAxC57Bl/6) F1 hybrid mice induced a long-term (2 years) oscillations in the numbers of day 7 CFU-S and day 11 CFU-S in the bone marrow and spleen. Dipin also damaged the hematopoietic stroma as indicated by decreased numbers of CFU-F which remained low for at least a year. The capacity of stromal cells to form ectopic hematopoietic foci was considerably inhibited and also remained low for 10 months. The obtained data suggest high dipin sensitivity of the earliest hematopoietic and stromal cells. The dynamics of CFU-S numbers in the hematopoietic organs supports their functioning on the basis of clonal succession (Kay, 1965).
Assuntos
Antineoplásicos Alquilantes/toxicidade , Aziridinas/toxicidade , Células da Medula Óssea/efeitos dos fármacos , Células-Tronco Hematopoéticas/efeitos dos fármacos , Animais , Camundongos , Baço/citologia , Células Estromais/efeitos dos fármacosRESUMO
Hemopoietic tissues were studied in vertebrates launched aboard the Soviet (Russian) biosatellites ("Cosmos-1129, 1514, 1667, 1887 and 2044"; "Bion-10 and 11") between 1980 and 1996. In the bone marrow of rats exposed to spaceflight conditions, a statistically significant decrease in cell number was revealed in the progenitor cell compartment accounting for the compensatory response of granulocyte-macrophage (CFU-gm) and erythrocyte lineages (BFU-e and CFU-e) and in the compartment of multipotent hemopoietic stem cells (CFU-s), which is responsible for the permanent renewal of hemopoietic tissue. The number of stromal fibroblastic progenitors (CFC-f) in the bone marrow of these rats was also reduced. Apparently, changes in the hemopoietic stroma damage the hemopoietic microenvironment and, hence, may be responsible for changes observed in the hemopoietic tissue proper. Attempts were made to develop methods for analyzing morphologically indiscernible clonogenic hemopoietic cells of newts, and studies on the effects of spaceflight factors on these cells were performed. The results showed that the numbers of clonogenic cells in newts of the flight group newts were significantly lower than in control newts. The data obtained are used as the basis for formulating the problems to be studied, drawing up a program for further research on the effects of spaceflight factors on stem and other clonogenic hemopoietic cells, and developing new experimental models for analyzing stem cells, the state of the hemopoietic stroma, etc.
Assuntos
Células da Medula Óssea/citologia , Fatores Estimuladores de Colônias/metabolismo , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Voo Espacial , Ausência de Peso , Animais , Células da Medula Óssea/metabolismo , Células da Medula Óssea/fisiologia , Ensaio de Unidades Formadoras de Colônias , Feminino , Granulócitos , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Hematopoéticas/fisiologia , Fígado/citologia , Masculino , Pleurodeles/sangue , Pleurodeles/fisiologia , Gravidez , Ratos , Ratos Wistar , Baço/citologia , Células EstromaisRESUMO
Effects of ten day long exposure to gamma-irradiation at low doses (mean dose rate of 1.5-2.0 m Gy/day, total dose of 15 m Gy) on hemopoietic (CFU-S) and stromal (CFU-F) progenitor cells from murine bone marrow were examined. The CFU-F content measured as in vitro fibroblastic colony number showed 1.5-4.5-fold increase. Additionally, the size of ectopic marrow transplants evaluated by counting myelokariocytes and CFU-S numbers also increased. No significant changes of CFU-S proliferation rate were found.
Assuntos
Células da Medula Óssea/efeitos da radiação , Animais , Células da Medula Óssea/citologia , Células Cultivadas , Ensaio de Unidades Formadoras de Colônias , Feminino , Fibroblastos/efeitos da radiação , Raios gama , Células-Tronco Hematopoéticas/efeitos da radiação , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Doses de Radiação , Células Estromais/efeitos da radiação , Fatores de TempoRESUMO
A simple and reliable technique has been developed for transplantation of the whole mouse spleen on the kidney. This technique ensures sufficient structural and functional recovery (transfer) of the original splenic microenvironments with a survival rate of 100%. The method allows counting the antibody-producing cells, as well as the spleen colony-forming units on the ectopic splenic territories. It may be used for studying hemopoietic and lymphoid microenvironments or for other special purposes.
Assuntos
Baço/transplante , Animais , Células Produtoras de Anticorpos/imunologia , Ensaio de Unidades Formadoras de Colônias , Sobrevivência de Enxerto , Rim/cirurgia , Masculino , Métodos , Camundongos , Baço/imunologiaRESUMO
The growth of macroscopic hemopoietic colonies was observed during postradiation regeneration in the spleen of dwarf hamsters (as well as of mice). The erythroid, granulocyte and megakaryocyte colonies were morphologically identified. The ratio between the erythroid and granulocyte colonies amounts to approximately 11. The formation of macroscopic spleen colonies and their morphology can be used for the functional characterization of hemopoietic microenvironment in the dwarf hamster.
Assuntos
Cricetinae/fisiologia , Células-Tronco Hematopoéticas/citologia , Baço/citologia , Animais , Feminino , Células-Tronco Hematopoéticas/classificação , Células-Tronco Hematopoéticas/efeitos da radiação , Masculino , Regeneração/efeitos da radiação , Baço/efeitos da radiaçãoRESUMO
It was shown for the first time that when the liver of 14 day-old mouse fetuses was transplanted under the renal capsule or in the subcutaneous connective tissue of adult recipients, successive de novo formation of hyaline cartilage, bone, and hemopoietic foci took place. We propose that the liver mesenchyme, which preserves wider differentiation potencies in fetuses, is the cellular source of different types of mechanocytes: cartilaginous, bone, and reticular cells.
Assuntos
Condrogênese , Transplante de Tecido Fetal , Transplante de Fígado , Osteogênese , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBARESUMO
The growth of bone marrow transplants under the renal capsule is stimulated in the mice irradiated at a dose of 700 r. The size of transplants increases, primarily, at the expense of hemopoietic tissue. The osteogenic potencies of stromal elements remain at a high enough level as well. Some features which characterize the decrease of viability of hemopoietic cells may be due to incomplete restoration of the system of sinusoidal capillaries.