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1.
J. appl. oral sci ; 25(6): 689-699, Nov.-Dec. 2017. graf
Artículo en Inglés | LILACS, BBO | ID: biblio-893665

RESUMEN

Abstract Periodontal regeneration is still a challenge in terms of predictability and magnitude of effect. In this study we assess the biological effects of combining chemical root conditioning and biological mediators on three relevant cell types for periodontal regeneration. Material and Methods: Bovine dentin slices were conditioned with 25% citric acid followed by topical application of basic fibroblast growth factor (bFGF, 10 and 50 ng). We used ELISA to assess the dynamics of bFGF release from the dentin surface and RT-qPCR to study the expression of Runx2, Col1a1, Bglap and fibronectin by periodontal ligament (PDL) fibroblasts, cementoblasts and bone marrow stromal cells (BMSC) grown onto these dentin slices. We also assessed the effects of topical application of bFGF on cell proliferation by quantification of genomic DNA. Results: Acid conditioning significantly increased the release of bFGF from dentin slices. Overall, bFGF application significantly (p<0.05) increased cell proliferation, except for BMSC grown on non-conditioned dentin slices. Dentin substrate discretely increased expression of Col1a1 in all cell types. Expression of Runx2, Col1a1 and Fn was either unaffected or inhibited by bFGF application in all cell types. We could not detect expression of the target genes on BMSC grown onto conditioned dentin. Conclusion: Acid conditioning of dentin improves the release of topically-applied bFGF. Topical application of bFGF had a stimulatory effect on proliferation of PDL fibroblasts, cementoblasts and BMSC, but did not affect expression of Runx2, Col1a1, Bglap and fibronectin by these cells.


Asunto(s)
Animales , Bovinos , Ligamento Periodontal/efectos de los fármacos , Regeneración/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Dentina/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Ligamento Periodontal/metabolismo , Expresión Génica , Factor 2 de Crecimiento de Fibroblastos/administración & dosificación
2.
J. appl. oral sci ; 24(2): 153-161, Mar.-Apr. 2016. graf
Artículo en Inglés | LILACS | ID: lil-779903

RESUMEN

ABSTRACT Objective Biocompatible materials such as interconnected porous hydroxyapatite ceramics (IP-CHA) loaded with osteogenic cells and bioactive agents are part of an evolving concept for overcoming craniofacial defects by use of artificial bone tissue regeneration. Amongst the bioactive agents, melatonin (MEL) and basic fibroblast growth factor (FGF-2) have been independently reported to induce osteoblastic activity. The present in vitro study was undertaken to examine the relationship between these two bioactive agents and their combinatory effects on osteoblastic activity and mineralization in vitro. Material and Methods Mouse preosteoblast cells (MC3T3-E1) were seeded and cultured within cylindrical type of IP-CHA block (ø 4x7 mm) by vacuum-assisted method. The IP-CHA/MC3T3 composites were subjected to FGF-2 and/or MEL. The proliferation assay, alkaline phosphatase enzyme activity (ALP), mRNA expressions of late bone markers, namely Osteocalcin (OCN) and Osteopontin (OPN), and Alizarin Red staining were examined over a period of 7 days. Results FGF-2 mainly enhanced the proliferation of MC3T3-E1 cells within the IP-CHA constructs. MEL mainly induced the mRNA expression of late bone markers (OCN and OPN) and showed increased ALP activity of MC3T3 cells cultured within IP-CHA construct. Moreover, the combination of FGF-2 and MEL showed increased osteogenic activity within the IP-CHA construct in terms of cell proliferation, upregulated expressions of OCN and OPN, increased ALP activity and mineralization with Alizarin Red. The synergy of the proliferative potential of FGF-2 and the differentiation potential of MEL showed increased osteogenic activity in MC3T3-E1 cells cultured within IP-CHA constructs. Conclusion These findings indicate that the combination of FGF-2 and MEL may be utilized with biocompatible materials to attain augmented osteogenic activity and mineralization.


Asunto(s)
Animales , Ratones , Osteoblastos/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Durapatita/farmacología , Sustitutos de Huesos/farmacología , Melatonina/farmacología , Factores de Tiempo , Ensayo de Materiales , Calcificación Fisiológica/efectos de los fármacos , Microscopía Electrónica de Rastreo , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proliferación Celular/efectos de los fármacos , Fosfatasa Alcalina/análisis
3.
Yonsei Medical Journal ; : 246-252, 2013.
Artículo en Inglés | WPRIM | ID: wpr-17422

RESUMEN

PURPOSE: Cell transplantation of myelin-producing exogenous cells is being extensively explored as a means of remyelinating axons in X-linked adrenoleukodystrophy. We determined whether 3,3',5-Triiodo-L-thyronine (T3) overexpresses the ABCD2 gene in the polysialylated (PSA) form of neural cell adhesion molecule (NCAM)-positive cells and promotes cell proliferation and favors oligodendrocyte lineage differentiation. MATERIALS AND METHODS: PSA-NCAM+ cells from newborn Sprague-Dawley rats were grown for five days on uncoated dishes in defined medium with or without supplementation of basic fibroblast growth factor (bFGF) and/or T3. Then, PSA-NCAM+ spheres were prepared in single cells and transferred to polyornithine/fibronectin-coated glass coverslips for five days to determine the fate of the cells according to the supplementation of these molecules. T3 responsiveness of ABCD2 was analyzed using real-time quantitative polymerase chain reaction, the growth and fate of cells were determined using 5-bromo-2-deoxyuridine incorporation and immunocytochemistry, respectively. RESULTS: Results demonstrated that T3 induces overexpression of the ABCD2 gene in PSA-NCAM+ cells, and can enhance PSA-NCAM+ cell growth in the presence of bFGF, favoring an oligodendrocyte fate. CONCLUSION: These results may provide new insights into investigation of PSA-NCAM+ cells for therapeutic application to X-linked adrenoleukodystrophy.


Asunto(s)
Animales , Ratas , Transportadoras de Casetes de Unión a ATP/metabolismo , Adrenoleucodistrofia/genética , Animales Recién Nacidos , Bromodesoxiuridina , Diferenciación Celular , Factor 2 de Crecimiento de Fibroblastos/farmacología , Fibronectinas/metabolismo , Inmunohistoquímica , Moléculas de Adhesión de Célula Nerviosa/genética , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Ácidos Siálicos/metabolismo , Células Madre , Hormonas Tiroideas/metabolismo , Triyodotironina/farmacología
4.
Experimental & Molecular Medicine ; : 133-137, 2005.
Artículo en Inglés | WPRIM | ID: wpr-90138

RESUMEN

In the course of screening of angiogenesis inhibitor from natural products, cryptotanshinone from Salvia miltiorrhiza was isolated as a potent small molecule inhibitor of angiogenesis. Cryptotanshinone inhibits bFGF-induced angiogenesis of BAECs at ten micromolar ranges in vitro without cytotoxicity. Tanshinone IIA, another tanshinone isolated from S. miltiorrhiza, which is structurally very similar to cryptotanshinone except C-15 position of dihydrofuran ring does not inhibit angiogenesis induced by bFGF. These results demonstrate that cryptotanshinone is a new anti-angiogenic agent and double bond at C-15 position of the dihydrofuran ring plays a crucial role in the activity.


Asunto(s)
Animales , Bovinos , Humanos , Inhibidores de la Angiogénesis/química , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Medicamentos Herbarios Chinos/química , Células Endoteliales/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Fenantrenos/química , Raíces de Plantas/química , Salvia miltiorrhiza/química
5.
Experimental & Molecular Medicine ; : 52-56, 2004.
Artículo en Inglés | WPRIM | ID: wpr-190974

RESUMEN

Human central neurocytoma is a kind of the brain tumors that are usually found in anterior part of the lateral ventricles. In this study, we established conditions that allowed proliferation of neurocytoma cells culture and analyzed characteristics of neurocytoma cells in vitro. For in vitro, a condition that used for culturing neural stem cells and contained basic fibroblast growth factor (bFGF) provided high proliferation. RT-PCR analaysis showed that nestin was found in neurocytoma cells, indicating that the neurocytomas possess neural stem cell properties. Interestingly, treatment of neurocytoma cells with forskolin increased expression of glial fibrillary acidic protein with a concomitant decrease in the nestin expression. Forskolin also induced morphological changes of neurocytoma cells to adopt an astrocyte-like phenotype. The results suggest that neurocyotma cells may have properties of multipotent neural stem cells.


Asunto(s)
Animales , Humanos , Astrocitos/citología , Diferenciación Celular/efectos de los fármacos , Proliferación Celular , Forma de la Célula , Factor 2 de Crecimiento de Fibroblastos/farmacología , Colforsina/farmacología , Proteínas de Filamentos Intermediarios/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Neurocitoma/tratamiento farmacológico , Células Tumorales Cultivadas
6.
Korean Journal of Ophthalmology ; : 121-131, 2004.
Artículo en Inglés | WPRIM | ID: wpr-94536

RESUMEN

We evaluated the effect of a basic fibroblast growth factor (bFGF) and saporin conjugate (bFGF-SAP) on proliferation, migration and tubule formation in bovine choriocapillary endothelial cells (BCECs). Cell proliferation and MTS assays were done with 0.01, 0.1, 1, 10, and 100 nM bFGF-SAP, and an equimolar concentration of bFGF and saporin. TUNEL assay was performed to confirm apoptosis. Cells were treated with 1, 10, and 100 nM bFGF-SAP and migration assay and tubule formation assay were done. Results were evaluated with image analysis. All experiments were performed in triplicate and repeated three times. Viable cells (ID50 = 0.62) and cell proliferation by MTS assay (ID50 = 0.75 nM) were inhibited. Saporin caused cytotoxicity and inhibition of proliferation at high concentration. DNA fragmentation was identified by TUNEL assay. Migration and tubule formation were also inhibited. All mechanisms responsible for neovascularization were inhibited, and this could be applied in the management of subretinal choroidal neovascularization (SRN).


Asunto(s)
Animales , Bovinos , Apoptosis/efectos de los fármacos , Recuento de Células , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Estudio Comparativo , Citotoxinas/farmacología , Endotelio Vascular/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Etiquetado Corte-Fin in Situ , Neovascularización Fisiológica/efectos de los fármacos , Proteínas de Plantas/farmacología , Proteínas Recombinantes de Fusión/farmacología
7.
Experimental & Molecular Medicine ; : 197-202, 1999.
Artículo en Inglés | WPRIM | ID: wpr-158705

RESUMEN

Endostatin, a carboxyl-terminal fragment of collagen XVIII is known as an anti-angiogenic agent, that specifically inhibits the proliferation of endothelial cell and the growth of several primary tumor. We report here the purification and characterization of the recombinant murine endostatin (rmEndostatin) which was expressed in a prokaryotic expression system. This rmEndostatin has similar physiochemical properties of yeast-produced recombinant endostatin, and it also specifically inhibits the proliferation and migration of bovine capillary endothelial cells stimulated by basic fibroblast growth factor. The biological activity of rmEndostatin was also shown by its anti-angiogenic ability on the chorioallantoic membrane of chick embryo in vivo. In this article, we demonstrate the refolding and purification of rmEndostatin, expressed using E. coli system, to a biologically active and soluble form. In addition, these results confirm the activity of endostatin as a potent anti-angiogenic agent. Copyright 2000 Academic Press.


Asunto(s)
Bovinos , Embrión de Pollo , Ratones , Inhibidores de la Angiogénesis/farmacología , Inhibidores de la Angiogénesis/aislamiento & purificación , Inhibidores de la Angiogénesis/genética , Animales , Western Blotting , Movimiento Celular/efectos de los fármacos , Corion/patología , Corion/efectos de los fármacos , Dicroismo Circular , Colágeno/farmacología , Colágeno/aislamiento & purificación , Colágeno/genética , Electroforesis en Gel de Poliacrilamida , Endotelio Vascular/efectos de los fármacos , Endotelio Vascular/citología , Escherichia coli/genética , Factor 2 de Crecimiento de Fibroblastos/farmacología , Neovascularización Fisiológica/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Fragmentos de Péptidos/aislamiento & purificación , Fragmentos de Péptidos/genética , Pliegue de Proteína , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/genética , Solubilidad , Levaduras/genética
8.
Experimental & Molecular Medicine ; : 203-209, 1999.
Artículo en Inglés | WPRIM | ID: wpr-158704

RESUMEN

Angiostatin is a potent angiogenesis inhibitor that is composed of the first four kringles of plasminogen fragment. Angiostatin with one less kringle molecule (kringle 1 to 3) was recently demonstrated to be an effective angiogenic inhibitor. To determine whether recombinant plasminogen kringle 1-3 (rPK1-3) can inhibit the corneal neovascularization induced by potent angiogenic factors; angiogenin, bFGF, or VEGF, hydron polymer discs each containing 2.0 microg of angiogenin, 500 ng of bFGF, or 500 ng of VEGF respectively were implanted into the corneal stroma of 138 rabbit eyes, and then discs each containing 10 microg, 12.5 microg, 20 microg or 30 microg of rPK1-3 were implanted randomly. Discs containing phosphate buffered saline were also implanted as a control. The angiogenesis score on number and length of newly formed vessels on the each of the rabbit's cornea were recorded daily by two observers (blinded). The treated corneas were also examined histologically. Recombinant PK1-3 treated corneas showed less neovascularization induced by all angiogenic factors (p < 0.05). and the extent of inhibition of neovascularization was proportional to the concentration of rPK1-3 (p < 0.05). Histologic examination showed leukocyte infiltration into the corneal stroma on the PBS treated eyes whereas rPK1-3 treated eyes showed only traces of leukocytes. These results of the effective rPK1-3 inhibition of corneal neovascularization induced by angiogenin, bFGF, or VEGF suggest that this angiostatin related fragment, rPK1-3, may be useful in the treatment of various neovascular diseases. Copyright 2000 Academic Press.


Asunto(s)
Embrión de Pollo , Conejos , Inhibidores de la Angiogénesis/farmacología , Inhibidores de la Angiogénesis/genética , Animales , Corion/efectos de los fármacos , Corion/irrigación sanguínea , Córnea/patología , Córnea/efectos de los fármacos , Córnea/irrigación sanguínea , Factores de Crecimiento Endotelial/farmacología , Factor 2 de Crecimiento de Fibroblastos/farmacología , Kringles/genética , Linfocinas/farmacología , Microscopía/métodos , Neovascularización Patológica/tratamiento farmacológico , Plasminógeno/farmacología , Plasminógeno/genética , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/genética , Ribonucleasa Pancreática/farmacología
9.
Indian J Biochem Biophys ; 1997 Feb-Apr; 34(1-2): 90-6
Artículo en Inglés | IMSEAR | ID: sea-28392

RESUMEN

Ganglioside (GG) and neurotrophic growth factor (GF) interactions in retinal neuronal and glial cells have been very little studied. Rat retinas were mechanically separated into outer (photoreceptor or PR) and inner (other neurons, IR) halves by planar vibratome sectioning and retinal Müller glial (RMG) cells were isolated and cultured according to previously published methods. The distribution on a percent molar basis of individual GG was different between the two halves: PR were dominated by GD3 (48% total GG) and contained only trace amounts (< 4%) of complex species (GT1b, GQ); IR was more typical of mature brain tissue, exhibiting substantial amounts (approximately 25%) of more complex GG. The GG profile of RMG cells was also simple, dominated by GM3 (60%) and GD1a (20%). A single addition to the medium of 500 pM bFGF or EGF for 48 hr to cultured RMG cells led to significant increases in total GG levels of 30-40%. Such treatments by both growth factors induced increases in GM3, whereas longer exposure (96 hr) of confluent RMG to these factors additionally stimulated synthesis of more complex GG. Incubations of RMG with [3H]-glucosamine showed that GG synthesis was 2-fold stimulated by growth factors. We also tested the effect of GM3 on one of the bFGF receptor transduction pathways, namely PI-3 kinase activation. To our knowledge these data constitute the first demonstration of neurotrophic factor stimulation of GG levels in cells of CNS in vitro. Such complex interactions may have particularly important consequences for neural physiopathology.


Asunto(s)
Animales , Células Cultivadas , Factor de Crecimiento Epidérmico/farmacología , Factor 2 de Crecimiento de Fibroblastos/farmacología , Gangliósidos/metabolismo , Metabolismo de los Lípidos , Factores de Crecimiento Nervioso/farmacología , Neuroglía/efectos de los fármacos , Neuronas/efectos de los fármacos , Ratas , Retina/citología
10.
Korean Journal of Ophthalmology ; : 1-7, 1996.
Artículo en Inglés | WPRIM | ID: wpr-77092

RESUMEN

We evaluated the effect of the conjugate of basic fibroblast growth factor (FGF2) and saporin (FGF2-SAP) on proliferation of cultured keratocytes. Cultured rabbit and human keratocytes were incubated in medium containing 0.01 to 100 nM of chemical conjugate of EGF2 conjugated by disulfide bond to saporin (CCFS1), FGF2 genetically fused to saporin (rFGF2-SAP), FGF2, or saporin for three hours or four days and cell proliferation was quantified four days after the drug treatment. Proliferation of rabbit and human keratocytes was effectively inhibited by three hour and by four day exposure to CCFS1 and rFGF2-SAP in a dose-dependent manner, whereas it was affected minimally by four day exposure to saporin. Their inhibitory effects were detected at concentrations above 0.1 or 1 nM, and were most prominent in serum-stimulated rabbit keratocytes. These results suggest a potential role for FGF2-SAP in limiting proliferation of keratocytes during corneal wound healing.


Asunto(s)
Animales , Humanos , Conejos , Antineoplásicos Fitogénicos/farmacología , División Celular/efectos de los fármacos , Línea Celular , Células Cultivadas , Sustancia Propia/citología , Relación Dosis-Respuesta a Droga , Factor 2 de Crecimiento de Fibroblastos/farmacología , Inmunotoxinas/farmacología , N-Glicosil Hidrolasas , Proteínas de Plantas/farmacología , Proteínas Inactivadoras de Ribosomas Tipo 1
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