Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The modification of titanium (Ti) surfaces by ion-implantation has previously been reported to enhance osseointegration in vivo. However, the mechanisms underlying the apparently improved biocompatibility of these novel implant materials are unknown. The aim of this study is, therefore, to determine the precise effects of calcium ion-implanted Ti on the functional activity of bone cells in vitro. Flow cytometry (FCM) and the reverse transcriptase polymerase chain reaction (RT-PCR) were used to measure the response of bone-derived cells to key bone-associated components, including alkaline phosphatase (ALP), bone morphogenetic protein receptor-1B (BMPR-1B), bone sialoprotein (BSP), osteonectin (ON), and osteopontin (OPN). FCM analysis showed that BMPR-1B, BSP and particularly OPN were significantly up-regulated in MG-63 cells cultured on Ca-implanted Ti compared with control nonimplanted Ti. Moreover, the effects of this novel Ca-Ti surface were found to be mediated, at least partly, via gene activation, since RT-PCR demonstrated the presence of notably elevated levels of OPN mRNA transcripts in the MG-63 cells. These findings thus show that Ti surfaces implanted with Ca ions can enhance the expression of certain bone-associated components in vitro, and suggest that this effect could be the cause of the potential benefit of this material on bone in vivo.
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PMID:Effects of calcium ion-implantation of titanium on bone cell function in vitro. 1743 6

In this study, we delineate the sequential expression of selected growth factors associated with bone formation in vitro. Mineralization, osteocalcin, and alkaline phosphatase (ALP-2) were measured to monitor the differentiation and maturation of osteoprogenitor cells collected from C57BL mice. Bone-related growth factors, including transforming growth factor beta (TGF-beta), fibroblast growth factor 2 (FGF-2), platelet-derived growth factor (PDGF), insulinlike growth factor (IGF)-1, vascular endothelial growth factor (VEGF), bone morphogenetic protein (BMP)-2, and BMP-7, were selected. Enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR) were used to measure growth factors at the protein and messenger ribonucleic acid (mRNA) level, respectively. The results found that ALP-2 expression increased progressively over time, whereas mineralization and osteocalcin did not become evident until culture day 14. VEGF and IGF-1 were upregulated early during proliferation. PDGF and TGF-beta mRNA expression was bimodal. FGF-2 and BMP-2 mRNAs were expressed only later in differentiation. FGF-2 mRNA signal levels were highest at day 14 and remained prominent through day 28 of culture. BMP-2 showed a similar profile as FGF-2. BMP-7 was not detectable using RT-PCR or ELISA. Strong correlations existed for the expression patterns between several early-response growth factors (VEGF, TGF-beta, and IGF-1) and were also evident for several late-response growth factors (BMP-2, PDGF, and FGF-2). Differential expression for grouped sets of growth factors occurs during the temporal acquisition of bone-specific markers as osteoprogenitor cell maturation proceeds in vitro.
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PMID:The sequential expression profiles of growth factors from osteoprogenitors [correction of osteroprogenitors] to osteoblasts in vitro. 1752 79

Our objective was to establish the role of fibroblasts in medial vascular calcification, a pathological process known to be associated with elastin degradation and remodeling. Rat dermal fibroblasts were treated in vitro with elastin degradation products and transforming growth factor (TGF)-beta1, factors usually present in deteriorated matrix environments. Cellular changes were monitored at the gene and protein level by reverse transcriptase-polymerase chain reaction, enzyme-linked immunosorbent assay, immunofluorescence, and von Kossa staining for calcium deposits. By 21 days, multicellular calcified nodules were formed in the presence of elastin degradation products and TGF-beta1 separately and to a significantly greater extent when used together. Before mineralization, cells expressed alpha-smooth muscle actin and large amounts of collagen type I and matrix metalloproteinase-2, characteristic features of myofibroblasts, key elements in tissue remodeling and repair. Stimulated cells expressed increased levels of core-binding factor alpha1, osteocalcin, alkaline phosphatase, and osteoprotegerin, representative bone-regulating proteins. For most proteins analyzed, TGF-beta1 synergistically amplified responses of fibroblasts to elastin degradation products. In conclusion, elastin degradation products and TGF-beta1 promote myofibroblastic and osteogenic differentiation in fibroblasts. These results support the idea that elastin-related calcification involves dynamic remodeling events and suggest the possibility of a defective tissue repair process.
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PMID:Osteogenic responses in fibroblasts activated by elastin degradation products and transforming growth factor-beta1: role of myofibroblasts in vascular calcification. 1759 59

The aim of the present study was to test the osteogenic effects of BMP-2 (bone morphogenetic protein-2) gene transfer in BMSCs (bone-marrow stromal cells) and rabbit calvarial bone defects. The pBMP-2-cDNA3.1 plasmid was constructed by subcloning hBMP-2 (human BMP-2) cDNA into the plasmid pcDNA3.1. BMSCs were transfected with a pBMP-2-cDNA3.1-Lipofect-amine complex. Transfected cells were observed for localization of the BMP-2 coding plasmid. Also, the level of BMP-2 in the culture medium of transfected cells was measured. The culture medium was collected and we tested whether this medium could induce non-transfected BMSCs to express ALP (alkaline phosphatase) and osteocalcin. The pBMP-2-cDNA3.1 complexes were incorporated into the collagen scaffold and the plasmid-loaded collagen scaffolds were then grafted into rabbit calvarial defects. After 2 weeks, granulation tissue at the grafted site was obtained and mRNA of BMP-2 was examined via RT (reverse transcriptase)--PCR. After 4 and 8 weeks, the animals were killed and the calvarial tissue was excised. After specimen preparation, optical microscopical examination was performed to evaluate bone formation. The results show that transfected cells were able to incorporate the BMP-2 gene into their nuclei. Also, the level of expressed and secreted BMP-2 was significantly higher in transfected cells than in untransfected cells (P<0.01). The retrieved culture medium could induce the expression of ALP and osteocalcin in non-transfected BMSCs. hBMP-2 mRNA was detected at the granulation tissue of experimental animals, but not in control animals after 2 weeks. At both 4 and 8 weeks, experimental groups showed significantly more newly formed bone area than the control group (P<0.01). Therefore pBMP-2-cDNA3.1 gene delivery could induce BMSCs into osteoblastic phenotype cells and enhance bone regeneration in rabbit calvarial bone defects.
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PMID:Osteogenic effects of bone-morphogenetic-protein-2 plasmid gene transfer. 1760 24

Osteoporotic women exhibit high frequency of alveolar bone loss and low bone density. Estrogen deficiency, which is vital in the pathogenesis of postmenopausal osteoporosis, has received increasing attention in the studies related to the periodontal diseases. Similar to most hormones, estrogen exerts its influence by binding to specific receptors, estrogen receptor (ER)-alpha and -beta. The periodontal ligament cells (PDLcs) are very important in maintaining the integrity of the periodontal tissue, which is the connective tissue located between the alveolar bone and the root surface of tooth. In this study, we evaluated the effects of estrogen deficiency on the alveolar bone in ovariectomized rats by histometric measurement of attachment level in vivo. Using the reverse transcriptase polymerase chain reaction (RT-PCR) and Western-blot procedure, we also detected mRNA and protein products of ERs and investigated the effects of estrogen on bone-forming capability by monitoring alkaline phosphatase (ALP) activity and osteocalcin production in cultured human PDLcs. Our results demonstrated that both ER-alpha and -beta were expressed in PDLcs. Moreover, when exposed to 17-beta estradiol, PDLcs exhibited positive modulation on ALP activity and osteocalcin production. The study suggests that estrogen and ERs may play an important role in periodontal diseases.
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PMID:The expression of estrogen receptors and the effects of estrogen on human periodontal ligament cells. 1780 34

Embryonic stem cell (ESC) culture is fragmented and laborious and involves operator decisions. Most protocols consist of 3 individual steps: maintenance, embryoid body (EB) formation, and differentiation. Integration will assist automation, ultimately aiding scale-up to clinically relevant numbers. These problems were addressed by encapsulating undifferentiated murine ESCs (mESCs) in 1.1% (w/v) low-viscosity alginic acid, 0.1% (v/v) porcine gelatin hydrogel beads (d = 2.3 mm). Six hundred beads containing 10,000 mESCs per bead were cultured in a 50-mL high-aspect-ratio vessel bioreactor. Bioreactor cultures were rotated at 17.5 revolutions per min, cultured in maintenance medium containing leukemia inhibitory factor for 3 days, replaced with EB formation medium for 5 days followed by osteogenic medium containing L-ascorbate-2-phosphate (50 microg/mL), beta-glycerophosphate (10 mM), and dexamethasone (1 microM) for an additional 21 days. After 29 days, 84 times as many cells per bead were observed and mineralized matrix was formed within the alginate beads. Osteogenesis was confirmed using von Kossa, Alizarin Red S staining, alkaline phosphatase activity, immunocytochemistry for osteocalcin, OB-cadherin, collagen type I, reverse transcriptase polymerase chain reaction, microcomputed tomography (micro-computed tomography) and Fourier transform infrared spectroscopic imaging. This simplified, integrated, and potentially scaleable methodology could enable the production of 3-demensional mineralized tissue from ESCs for potential clinical applications.
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PMID:Integrated 3-dimensional expansion and osteogenic differentiation of murine embryonic stem cells. 1798 91

The use of light for medical treatment has been studied previously. In this study, we examined the effect of light from a red light-emitting diode on osteogenic differentiation of mouse mesenchymal stem cells (D1 cells) which were cultured in the presence of osteogenic differentiation medium (ODM) for 3 days, then exposed to a red light-emitting diode (LED) light of 647 nm wavelength once for 10 s, 30 s or 90 s with radiation energies of 0.093 J, 0.279 J and 0.836 J, respectively. D1 cells in the presence of ODM differentiated into osteoblasts, and this process was enhanced on exposure to LED light in ODM medium. This effect was confirmed by increased Alizarin red staining, higher alkaline phosphatase (ALP) activity, higher mRNA expressions of osteocalcin, collagen type I, osteopontin and Runt-related transcription factor2 (Runx2), and higher levels by reverse transcriptase-polymerase chain reaction (RT-PCR) and by increased immunofluorescence staining against cluster of differentiation 44 (CD44) by immunofluorescence microscopy, confocal microscopy and flow cytometric analysis. These data suggest that osteogenic differentiation of mesenchymal stem cells (MSCs) in ODM is enhanced by LED light exposure.
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PMID:Red light of 647 nm enhances osteogenic differentiation in mesenchymal stem cells. 1962 Dec 50

The biocompatibility of nanoparticles is the prerequisite for their applications in biomedicine but can be misleading due to the absence of criteria for evaluating the safety and toxicity of those nanomaterials. Recent studies indicate that mesoporous silica nanoparticles (MSNs) can easily internalize into human mesenchymal stem cells (hMSCs) without apparent deleterious effects on cellular growth or differentiation, and hence are emerging as an ideal stem cell labeling agent. The objective of this study was to thoroughly investigate the effect of MSNs on osteogenesis induction and to examine their biocompatibility in hMSCs. Uptake of MSNs into hMSCs did not affect the cell viability, proliferation and regular osteogenic differentiation of the cells. However, the internalization of MSNs indeed induced actin polymerization and activated the small GTP-bound protein RhoA. The MSN-induced cellular protein responses as believed to cause osteogenesis of hMSCs did not result in promotion of regular osteogenic differentiation as analyzed by cytochemical stain and protein activity assay of alkaline phosphatase (ALP). When the effect of MSNs on ALP gene expression was further examined by reverse transcriptase polymerase chain reaction, MSN-treated hMSCs were shown to have significantly higher mRNA expression than control cells after 1-hour osteogenic induction. The induction of ALP gene expression by MSNs, however, was absent in cells after 1-day incubation with osteogenic differentiation. Together our results show that the internalization of MSNs had a significant effect on the transient protein response and osteogenic signal in hMSCs, thereby suggesting that the effects of nanoparticles on diverse aspects of cellular activities should be carefully evaluated even though the nanoparticles are generally considered as biocompatible at present.
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PMID:Internalization of mesoporous silica nanoparticles induces transient but not sufficient osteogenic signals in human mesenchymal stem cells. 1851 41

Researchers in our laboratory have previously shown that ghrelin, a gastric peptide hormone, may regulate mesenchymal cell differentiation into adipocytes and myocytes. Here we show that ghrelin promotes osteogenesis of intramembranous bone and improves the repair of calvarial bone defects in rats. Rats with a 9 mm full-thickness calvarial bone defect received either Bio-Oss (control group) or Bio-Oss mixed with 20 mug ghrelin (treatment group), followed by local administration of saline or ghrelin (10 microg), respectively, on days 5, 10 and 15. After 6 and 12 weeks, new bone formation was assessed. Animals treated with ghrelin showed a significant increase in new bone formation as demonstrated by an increment in bone mineral density and fluorescence labelling of tetracycline relative to the control group. At 6 weeks, bone mineral density increased from 54 +/- 7 (control group) to 78 +/- 9 mg cm(-2) in the treatment group, while the tetracycline fluorescence labelling increased by 61 +/- 15%. A similar increment was observed at 12 weeks. Quantitative reverse transcriptase-polymerase chain reaction showed that expression of alkaline phosphatase (ALP), osteocalcin and collagen type I was elevated. Relative to the control animals, mRNAs for ALP, osteocalcin and collagen type I increased 2.4 +/- 0.4-, 4.7 +/- 1.9- and 4.0 +/- 1.7-fold, respectively, in animals treated with ghrelin for 6 weeks (P < 0.05). At 12 weeks, mRNA levels of ALP, osteocalcin and collagen type I showed a decline relative to levels at 6 weeks but still remained significantly higher than in the control group, with fold changes of 2.4 +/- 0.8, 2.4 +/- 1.2 and 2.1 +/- 0.7, respectively (P < 0.05). This study demonstrated that ghrelin stimulates intramembranous osteogenesis.
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PMID:Stimulation of intramembranous bone repair in rats by ghrelin. 1856 76

Alendronate inhibits osteoclastic activity. However, some studies suggest alendronate also has effects on osteoblast activity. We hypothesized alendronate would enhance osteoblastic differentiation without causing cytotoxicity of the osteoblasts. We evaluated the effect of alendronate on the osteogenic differentiation of mouse mesenchymal stem cells. D1 cells (multipotent mouse mesenchymal stem cells) were cultured in osteogenic differentiation medium for 7 days and then treated with alendronate for 2 days before being subjected to various tests using MTT assays, Alizarin Red, enzyme-linked immunosorbent assay, energy-dispersive xray spectrophotometry, reverse transcriptase-polymerase chain reaction, confocal microscopy, and flow cytometric analysis. D1 cells differentiated into osteoblasts in the presence of osteogenic differentiation medium as confirmed by positive Alizarin Red S staining, increased alkaline phosphatase activity and osteocalcin mRNA expression, a calcium peak by energy-dispersive xray spectrophotometry, and by positive immunofluorescence staining against CD44. Osteogenic differentiation was enhanced after treatment with alendronate as confirmed by Alizarin Red S staining, elevated alkaline phosphatase activity and osteocalcin mRNA expression, a greater calcium peak by energy-dispersive xray spectrophotometry, and by immunofluorescence staining against CD44 by flow cytometric analysis. These data suggest alendronate enhances osteogenic differentiation when treated with mouse mesenchymal stem cells in osteogenic differentiation medium.
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PMID:Alendronate enhances osteogenic differentiation of bone marrow stromal cells: a preliminary study. 1866 32


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