Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 10 de 10
Filtrar
Mais filtros










Base de dados
Intervalo de ano de publicação
1.
Leukemia ; 30(11): 2221-2231, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27150009

RESUMO

Mesenchymal stromal cells (MSCs) have been shown to reverse radiation damage to marrow stem cells. We have evaluated the capacity of MSC-derived extracellular vesicles (MSC-EVs) to mitigate radiation injury to marrow stem cells at 4 h to 7 days after irradiation. Significant restoration of marrow stem cell engraftment at 4, 24 and 168 h post irradiation by exposure to MSC-EVs was observed at 3 weeks to 9 months after transplant and further confirmed by secondary engraftment. Intravenous injection of MSC-EVs to 500cGy exposed mice led to partial recovery of peripheral blood counts and restoration of the engraftment of marrow. The murine hematopoietic cell line, FDC-P1 exposed to 500cGy, showed reversal of growth inhibition, DNA damage and apoptosis on exposure to murine or human MSC-EVs. Both murine and human MSC-EVs reverse radiation damage to murine marrow cells and stimulate normal murine marrow stem cell/progenitors to proliferate. A preparation with both exosomes and microvesicles was found to be superior to either microvesicles or exosomes alone. Biologic activity was seen in freshly isolated vesicles and in vesicles stored for up to 6 months in 10% dimethyl sulfoxide at -80 °C. These studies indicate that MSC-EVs can reverse radiation damage to bone marrow stem cells.


Assuntos
Vesículas Extracelulares/fisiologia , Células-Tronco Hematopoéticas/efeitos da radiação , Células-Tronco Mesenquimais/citologia , Animais , Células da Medula Óssea , Dano ao DNA , Vesículas Extracelulares/transplante , Sobrevivência de Enxerto , Humanos , Masculino , Camundongos , Efeitos da Radiação , Transplante de Células-Tronco , Transplante Heterólogo , Resultado do Tratamento
2.
Leukemia ; 28(4): 813-22, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23989430

RESUMO

Prevailing wisdom holds that hematopoietic stem cells (HSCs) are predominantly quiescent. Although HSC cycle status has long been the subject of scrutiny, virtually all marrow stem cell research has been based on studies of highly purified HSCs. Here we explored the cell cycle status of marrow stem cells in un-separated whole bone marrow (WBM). We show that a large number of long-term multi-lineage engraftable stem cells within WBM are in S/G2/M phase. Using bromodeoxyuridine, we show rapid transit through the cell cycle of a previously defined relatively dormant purified stem cell, the long-term HSC (LT-HSC; Lineage(-)/c-kit(+)/Sca-1(+)/Flk-2(-)). Actively cycling marrow stem cells have continually changing phenotype with cell cycle transit, likely rendering them difficult to purify to homogeneity. Indeed, as WBM contains actively cycling stem cells, and highly purified stem cells engraft predominantly while quiescent, it follows that the population of cycling marrow stem cells within WBM are lost during purification. Our studies indicate that both the discarded lineage-positive and lineage-negative marrow cells in a stem cell separation contain cycling stem cells. We propose that future work should encompass this larger population of cycling stem cells that is poorly represented in current studies solely focused on purified stem cell populations.


Assuntos
Células da Medula Óssea/citologia , Ciclo Celular , Linhagem da Célula , Células-Tronco Hematopoéticas/citologia , Animais , Citometria de Fluxo , Masculino , Camundongos , Camundongos Endogâmicos C57BL
4.
Circulation ; 104(5): 594-9, 2001 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-11479259

RESUMO

BACKGROUND: Therapeutic angiogenesis by the administration of recombinant vascular endothelial growth factor (rVEGF) is a novel strategy for the treatment of ischemic disorders. rVEGF has been delivered as a protein, by plasmid DNA, and by genetically engineered cells with different pharmacokinetic and physiological properties. In the present study, we examined a new method for delivery of rVEGF using implantable bioartificial muscle (BAM) tissues made from genetically modified primary skeletal myoblasts. Our goal was to determine whether the rVEGF delivered by this technique promoted controlled angiogenesis in nonischemic and/or ischemic adult mouse tissue. METHODS AND RESULTS: Primary adult mouse myoblasts were retrovirally transduced to secrete human or mouse rVEGF and tissue-engineered into implantable 1x10 to 15-mm BAMs containing parallel arrays of postmitotic myofibers. In vitro, they secreted 290 to 511 ng of bioactive mouse or human VEGF/BAM per day. rVEGF BAMs implanted subcutaneously into syngeneic mice caused a 30-fold increase in the number of CD31-positive capillary cells within the BAM by 1 week compared with control BAMs. Implantation of rVEGF-secreting BAMs into ischemic hindlimbs resulted in a 2- to 3-fold increase in capillary density of neighboring host muscle by 1 week and out to 4 weeks with no evidence of hemangioma formation. CONCLUSIONS: Local delivery of rVEGF from BAMs rapidly increases capillary density both within the BAM itself and in adjacent ischemic muscle tissue. Genetically engineered muscle tissue provides a method for therapeutic protein delivery in a dose-regulated fashion.


Assuntos
Fatores de Crescimento Endotelial/administração & dosagem , Linfocinas/administração & dosagem , Músculo Esquelético/metabolismo , Neovascularização Fisiológica/efeitos dos fármacos , Animais , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Células Cultivadas , Fatores de Crescimento Endotelial/genética , Fatores de Crescimento Endotelial/metabolismo , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/metabolismo , Técnicas de Transferência de Genes , Sobrevivência de Enxerto/efeitos dos fármacos , Sobrevivência de Enxerto/genética , Humanos , Imuno-Histoquímica , Isquemia , Linfocinas/genética , Linfocinas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C3H , Músculo Esquelético/irrigação sanguínea , Músculo Esquelético/transplante , Neovascularização Fisiológica/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/análise , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/metabolismo , Fatores de Tempo , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio Vascular
5.
FASEB J ; 13(9): 1031-8, 1999 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10336885

RESUMO

Space travel causes rapid and pronounced skeletal muscle wasting in humans that reduces their long-term flight capabilities. To develop effective countermeasures, the basis of this atrophy needs to be better understood. Space travel may cause muscle atrophy indirectly by altering circulating levels of factors such as growth hormone, glucocorticoids, and anabolic steroids and/or by a direct effect on the muscle fibers themselves. To determine whether skeletal muscle cells are directly affected by space travel, tissue-cultured avian skeletal muscle cells were tissue engineered into bioartificial muscles and flown in perfusion bioreactors for 9 to 10 days aboard the Space Transportation System (STS, i.e., Space Shuttle). Significant muscle fiber atrophy occurred due to a decrease in protein synthesis rates without alterations in protein degradation. Return of the muscle cells to Earth stimulated protein synthesis rates of both muscle-specific and extracellular matrix proteins relative to ground controls. These results show for the first time that skeletal muscle fibers are directly responsive to space travel and should be a target for countermeasure development.


Assuntos
Músculo Esquelético/metabolismo , Atrofia Muscular/metabolismo , Voo Espacial , Ausência de Peso/efeitos adversos , Animais , Aves , Células Cultivadas , Técnicas de Cultura/métodos , Lactatos/metabolismo , Músculo Esquelético/patologia , Atrofia Muscular/patologia , Proteínas/metabolismo
6.
Hum Gene Ther ; 10(4): 565-77, 1999 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-10094200

RESUMO

Murine skeletal muscle cells transduced with foreign genes and tissue engineered in vitro into bioartificial muscles (BAMs) are capable of long-term delivery of soluble growth factors when implanted into syngeneic mice (Vandenburgh et al., 1996b). With the goal of developing a therapeutic cell-based protein delivery system for humans, similar genetic tissue-engineering techniques were designed for human skeletal muscle stem cells. Stem cell myoblasts were isolated, cloned, and expanded in vitro from biopsied healthy adult (mean age, 42 +/- 2 years), and elderly congestive heart failure patient (mean age, 76 +/- 1 years) skeletal muscle. Total cell yield varied widely between biopsies (50 to 672 per 100 mg of tissue, N = 10), but was not significantly different between the two patient groups. Percent myoblasts per biopsy (73 +/- 6%), number of myoblast doublings prior to senescence in vitro (37 +/- 2), and myoblast doubling time (27 +/- 1 hr) were also not significantly different between the two patient groups. Fusion kinetics of the myoblasts were similar for the two groups after 20-22 doublings (74 +/- 2% myoblast fusion) when the biopsy samples had been expanded to 1 to 2 billion muscle cells, a number acceptable for human gene therapy use. The myoblasts from the two groups could be equally transduced ex vivo with replication-deficient retroviral expression vectors to secrete 0.5 to 2 microg of a foreign protein (recombinant human growth hormone, rhGH)/10(6) cells/day, and tissue engineered into human BAMs containing parallel arrays of differentiated, postmitotic myofibers. This work suggests that autologous human skeletal myoblasts from a potential patient population can be isolated, genetically modified to secrete foreign proteins, and tissue engineered into implantable living protein secretory devices for therapeutic use.


Assuntos
Órgãos Artificiais , Terapia Genética , Hormônio do Crescimento/genética , Músculo Esquelético/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Diferenciação Celular , Divisão Celular , Feminino , Hormônio do Crescimento/uso terapêutico , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Músculo Esquelético/citologia , Radioimunoensaio , Proteínas Recombinantes/genética , Proteínas Recombinantes/uso terapêutico , Transdução Genética
7.
Methods Mol Med ; 18: 217-25, 1999.
Artigo em Inglês | MEDLINE | ID: mdl-21370179

RESUMO

Skeletal muscle structure is regulated by many factors, including nutrition, hormones, electrical activity, and tension. The muscle cells are subjected to both passive and active mechanical forces at all stages of development, and these forces play important but poorly understood roles in regulating muscle organogenesis and growth. For example, during embryogenesis, the rapidly growing skeleton places large passive mechanical forces on the attached muscle tissue. These forces not only help to organize the proliferating mononucleated myoblasts into the oriented, multinucleated myofibers of a functional muscle, but also tightly couple the growth rate of muscle to that of bone. Postnatally, the actively contracting, innervated muscle fibers are subjected to different patterns of active and passive tensions that regulate longitudinal and cross-sectional myofiber growth. These mechanically induced organogenic processes have been difficult to study under normal tissue culture conditions, resulting in the development of numerous methods and specialized equipment to simulate the in vivo mechanical environment (1-4). These techniques have led to the engineering of bioartificial muscles (organoids), which display many of the characteristics of in vivo muscle, including parallel arrays of postmitotic fibers organized into fascicle-like structures with tendon-like ends. They are contractile, express adult isoforms of contractile proteins, perform directed work, and can be maintained in culture for long periods.

8.
Hum Gene Ther ; 9(17): 2555-64, 1998 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-9853522

RESUMO

Skeletal muscle wasting is a significant problem in elderly and debilitated patients. Growth hormone (GH) is an anabolic growth factor for skeletal muscle but is difficult to deliver in a therapeutic manner by injection owing to its in vivo instability. A novel method is presented for the sustained secretion of recombinant human GH (rhGH) from genetically modified skeletal muscle implants, which reduces host muscle wasting. Proliferating murine C2C12 skeletal myoblasts stably transduced with the rhGH gene were tissue engineered in vitro into bioartificial muscles (C2-BAMs) containing organized postmitotic myofibers secreting 3-5 microg of rhGH/day in vitro. When implanted subcutaneously into syngeneic mice, C2-BAMs delivered a sustained physiologic dose of 2.5 to 11.3 ng of rhGH per milliliter of serum. rhGH synthesized and secreted by the myofibers was in the 22-kDa monomeric form and was biologically active, based on downregulation of a GH-sensitive protein synthesized in the liver. Skeletal muscle disuse atrophy was induced in mice by hindlimb unloading, causing the fast plantaris and slow soleus muscles to atrophy by 21 to 35% ( < 0.02). This atrophy was significantly attenuated 41 to 55% (p < 0.02) in animals that received C2-BAM implants, but not in animals receiving daily injections of purified rhGH (1 mg/kg/day). These data support the concept that delivery of rhGH from BAMs may be efficacious in treating muscle-wasting disorders.


Assuntos
Órgãos Artificiais , Hormônio do Crescimento Humano/uso terapêutico , Músculo Esquelético/patologia , Atrofia Muscular/prevenção & controle , Animais , Linhagem Celular , Clonagem Molecular , Hormônio do Crescimento Humano/biossíntese , Hormônio do Crescimento Humano/genética , Masculino , Camundongos , Camundongos Endogâmicos C3H , Músculo Esquelético/transplante , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/uso terapêutico
10.
Hum Gene Ther ; 7(17): 2195-200, 1996 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-8934233

RESUMO

Genetically modified murine skeletal myoblasts were tissue engineered in vitro into organ-like structures (organoids) containing only postmitotic myofibers secreting pharmacological levels of recombinant human growth hormone (rhGH). Subcutaneous organoid implantation under tension led to the rapid and stable appearance of physiological sera levels of rhGH for up to 12 weeks, whereas surgical removal led to its rapid disappearance. Reversible delivery of bioactive compounds from postmitotic cells in tissue engineered organs has several advantages over other forms of muscle gene therapy.


Assuntos
Terapia Genética/métodos , Hormônio do Crescimento/genética , Músculo Esquelético/química , Animais , Análise Química do Sangue , Células Cultivadas , Sistemas de Liberação de Medicamentos/métodos , Regulação da Expressão Gênica , Hormônio do Crescimento/análise , Humanos , Camundongos , Fibras Musculares Esqueléticas , Músculo Esquelético/citologia , Técnicas de Cultura de Órgãos/métodos , Transplante de Órgãos , Radioimunoensaio
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...