Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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 presence of osteogenic progenitors in human skeletal muscle is suggested by the formation of ectopic bone in clinical and experimental conditions, but their direct identification has not yet been demonstrated. The aims of this study were to identify osteogenic progenitor cells in human skeletal muscle tissue and to expand and characterize them in culture. Specimens of gracilis and semitendinosus muscle were obtained from young adults and digested to separate the connective tissue and satellite cell fractions. The cells were cultured and characterized morphologically and immunohistochemically using antibodies known to be reactive with primitive osteoprogenitor cells, pericytes, intermediate filaments, and endothelial cells. Alkaline phosphatase activity and osteocalcin gene expression were also determined. In the early stages of culture, the connective tissue cells obtained were highly positive for primitive osteoprogenitor cell and for pericyte markers. Alkaline phosphatase activity was detectable at early stages of culture and rose as a function of time, whereas primitive osteoprogenitor cell markers declined and osteocalcin mRNA expression became detectable by reverse transcriptase-polymerase chain reaction (RT-PCR). It is shown that human skeletal muscle connective tissue contains osteogenic progenitor cells. Their identification as pericytes, perivascular cells with established osteogenic potential, suggests a cellular link between angiogenesis and bone formation in muscle tissue. These cells are easily cultured and expanded in vitro by standard techniques, providing an alternative source of osteogenic progenitor cells for possible cell-based therapeutic use in certain conditions.
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PMID:Osteoprogenitor cells of mature human skeletal muscle tissue: an in vitro study. 1159 13

Postmenopausal osteoporosis is a contributing factor to alveolar bone atrophy associated with tooth loss in the elderly. The use of dental titanium implants has been increasingly adapted to treat these edentulous patients. This study examines whether female gonadal hormone deficiency interferes with the critical integration process between bone and implants. Two types of experimental titanium implants with acid-treated surfaces were placed in the femurs of ovariectomized (ovx) and sham-operated control rats: T-cell implants with a hollow chamber for histomorphometric and steady-state mRNA expression assays, and unthreaded cylindrical implants for biomechanical push-in tests. At week 2, less bone area was found in the ovx-implant group (p = 0.0495) than in the sham-implant group. The implant push-in test showed that the ovx-implant group had approximately half of the withstanding value of the sham-implant group (p = 0.009). However, these differences between the ovx and sham groups became diminished at week 4. Total RNA samples were examined by a reverse transcriptase-polymerase chain reaction assay for col1a1, col3a1, bone sialoprotein (bSP) II, osteonectin, osteopontin, osteocalcin, integrin beta1 and integrin beta3. In untreated bones and in created bone defects without implant placement, ovx did not affect the steady-state levels of the mRNAs tested. When implants were placed, significant upregulation of these genes was observed in the sham-implant group; however, only osteocalcin and integrins were upregulated in the ovx-implant group. The results suggest a biphasic effect of female gonadal hormone deficiency that may temporarily interfere with the early implant-tissue integration process, and which may be associated with a failure to upregulate a selected set of bone extracellular matrix genes. Once established, however, functional bone-implant integration can be achieved even in ovx rats.
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PMID:Ovariectomy hinders the early stage of bone-implant integration: histomorphometric, biomechanical, and molecular analyses. 1179 76

The effect of standard orthopaedic implant materials on osteoblast proliferation and differentiation was investigated using a human osteoblast cell culture system. Human fetal osteoblasts 1.19 were cultured on stainless steel, cobalt-chrome-molybdenum, and commercially pure titanium for 12 days. Tissue culture polystyrene was used as a control. Cell proliferation was measured by electronic cell counting and by a colorimetric proliferation assay. To assess the degree of differentiation, levels of alkaline phosphatase activity, collagen Type I, and osteocalcin production were measured. Osteocalcin gene expression was measured by reverse transcriptase-polymerase chain reaction. Electronic cell counting and proliferation assays showed lower cell numbers and delayed proliferation on stainless steel and cobalt-chrome-molybdenum compared with titanium and polystyrene. Alkaline phosphatase and osteocalcin were measured higher on titanium than on stainless steel or cobalt-chrome-molybdenum. Differences in collagen Type I production were not found. Reverse transcriptase-polymerase chain reaction showed the highest osteocalcin gene expression on titanium. The human fetal osteoblast cell line 1.19 provides a rapidly proliferating and differentiating system for testing biomaterials in which differences in osteoblast proliferation and differentiation on orthopaedic implant materials could be revealed, suggesting that the chemistry of biomaterials has a dynamic effect on proliferation and differentiation of human osteoblasts.
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PMID:Testing of skeletal implant surfaces with human fetal osteoblasts. 1179 45

Cell- and time-specific gene inactivation should enhance our knowledge of bone biology. Implementation of this technique requires construction of transgenic mouse lines expressing Cre recombinase in osteoblasts, the bone forming cell. We tested several promoter fragments for their ability to drive efficient Cre expression in osteoblasts. In the first mouse transgenic line, the Cre gene was placed under the control of the 2.3-kb proximal fragment of the alpha1(I)-collagen promoter, which is expressed at high levels in osteoblasts throughout their differentiation. Transgenic mice expressing this transgene in bone were bred with the ROSA26 reporter (R26R) strain in which the ROSA26 locus is targeted with a conditional LacZ reporter cassette. In R26R mice, Cre expression and subsequent Cre-mediated recombination lead to expression of the LacZ reporter gene, an event that can be monitored by LacZ staining. LacZ staining was detected in virtually all osteoblasts of alpha1(I)-Cre;R26R mice indicating that homologous recombination occurred in these cells. No other cell type stained blue. In the second line studied, the 1.3-kb fragment of osteocalcin gene 2 (OG2) promoter, which is active in differentiated osteoblasts, was used to drive Cre expression. OG2-Cre mice expressed Cre specifically in bone. However, cross of OG2-Cre mice with R26R mice did not lead to any detectable LacZ staining in osteoblasts. Lastly, we tested a more active artificial promoter derived from the OG2 promoter. The artificial OG2-Cre transgene was expressed by reverse transcriptase-polymerase chain reaction in cartilage and bone samples. After cross of the artificial OG2-Cre mice with R26R mice, we detected a LacZ staining in articular chondrocytes but not in osteoblasts. Our data suggest that the only promoter able to drive Cre expression at a level sufficient to induce recombination in osteoblasts is the alpha1(I)-collagen promoter.
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PMID:Mouse alpha1(I)-collagen promoter is the best known promoter to drive efficient Cre recombinase expression in osteoblast. 1211 77

Understanding the factors that control osteoblastic behavior is centrally important in establishment of successful osseointegration. Pharmacogenetic control of the osteoblast to increase the mineral content around dental implants may offer a unique advantage to clinicians in improving osseointegration success and decreasing time before mechanical loading. This in vitro pilot study has screened for bioactive peptides derived from bone morphogenetic protein 7 (BMP-7) (also called osteogenic protein 1 [OP-1]). Thirteen overlapping peptides of BMP-7 were synthesized and covalently coupled to glass coverslip substrates using silane chemistry. The rate and relative amount of mineralization were compared by von Kossa analysis using primary rat calvarial osteoblastic cell populations during a 7- to 21-day period. In addition, bone sialoprotein (BSP) and osteocalcin (OC) gene expression was measured from osteoblastic cells grown on peptide-immobilized glass coverslips by reverse transcriptase-polymerase chain reaction. Initial results from mineralization studies suggested the BMP-7-derived peptides were able to support mineralization to varying degrees with enhanced peptide-induced mineralization from the C-and N-termini of the BMP-7 molecule. Analysis of these peptide regions indicated that these peptides comprised the finger 1 and 2 domains of CP-1, which contribute toward ligand-receptor interaction. Further analysis of gene expression from select peptide-immobilized substrates indicated that peptides from the C-terminus of BMP-7 were capable of supporting BSP and OC messenger RNA expression. These studies indicate that BMP-7 peptides covalently bound to solid substrates may provide the biological basis to immobilize peptides to titanium implants to induce osteoblastic differentiation and mineralization in a more predictable fashion.
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PMID:In vitro mineralization studies with substrate-immobilized bone morphogenetic protein peptides. 1276 Apr 48

This study investigated the effects of mineral trioxide aggregate on cementoblast growth and osteocalcin production in tissue culture. For cellular morphology studies, cementoblasts on mineral trioxide aggregate, IRM, and amalgam were incubated for 48 h then fixed for scanning electron microscopic analysis. For gene expression on mineral trioxide aggregate and IRM, reverse transcriptase polymerase chain reaction was performed using primer sets for glyceraldehyde-3-phosphate dehydrogenase, type I collagen, alkaline phosphatase, osteocalcin, and bone sialoprotein after 3 and 5 days. In vitro matrix protein expression was evaluated by confocal microscopy for the presence of osteocalcin on MTA after 7 and 12 days. Images were compared with controls to assess qualitative differences. Results suggest that mineral trioxide aggregate permits cementoblast attachment and growth and the production of mineralized matrix gene and protein expression. Our data indicates that mineral trioxide aggregate can be considered cementoconductive.
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PMID:Cementoblasts maintain expression of osteocalcin in the presence of mineral trioxide aggregate. 1281 26

Cloned cementoblasts (OCCMs), periodontal ligament fibroblasts (SV-PDLs), and dental follicle (SV-F) cells obtained from mice were used as a tool to study periodontal tissue engineering. OCCM, SV-PDL, and SV-F cells were seeded onto three-dimensional poly lactic-co-glycolic acid (PLGA) scaffolds and cultured with the use of bioreactors or implanted subcutaneously in severe combined immune deficiency (SCID) mice for up to 6 weeks. We explored the behavior of these cells in porous PLGA sponges by cell growth, expression of mineral-associated genes using reverse transcriptase polymerase chain reaction, and mineralization by histologic analysis in vitro and in vivo. Results indicated that cells attached to PLGA scaffolds under either static or dynamic conditions in vitro. Only OCCM implants, retrieved from both in vitro bioreactors and SCID mice at 3-and 6-weeks post-cell implantation exhibited mineral formation. Types I and XII collagens, osteocalcin, and bone sialoprotein genes were detected in all implants retrieved from SCID mice. These results suggest that delivery of selected cells via PLGA scaffolds may serve as a viable approach for promoting periodontal tissue regeneration.
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PMID:Cementum engineering with three-dimensional polymer scaffolds. 1451 61

Bone defects caused by periapical inflammation can be treated and improved by endodontic therapy. However, the mechanism for osseous healing of periapical lesions after root canal treatment is unclear. In this study we examined whether fibroblastic cells from human periapical granulation tissue could produce calcified matrix in vitro. Periapical lesions from three patients were dissected in endodontic surgery, and fibroblastic cells (HFC) migrating from these lesions in vitro were used in this study. The HFC were cultured with or without beta-glycerophosphate (beta-GP) and ascorbic acid (AA), and the expression of human runt-related transcription factor-2 (Runx2), osterix (Osx), osteopontin (Opn), and osteocalcin (Ocn) mRNA, and alkaline phosphatase (ALPase) was examined by reverse transcriptase-polymerase chain reaction (RT-PCR) or by an enzyme-cytochemical technique. Furthermore, calcification in the cells was investigated by von Kossa staining. At the beginning of the culture, HFC expressed Runx2 mRNA faintly, but neither Opn mRNA nor ALPase activity. Immunocytochemical study also showed HFC expressed Runx2 more weakly, compared to SaOS2. However, the expression levels of ALPase, and Runx2, Osx, and Opn mRNA, were stimulated by 2 mM beta-GP and 50 microg/ml AA. After 4 weeks of culture with 2 mM beta-GP and 50 microg/ml AA, HFC formed von Kossa staining-positive calcified deposits on culture dishes, and also expressed Ocn mRNA. These results suggest that inflamed periapical granulation tissue contains osteogenic cells that have the potential to differentiate into mature osteoblastic or cementoblastic cells, and that such cells might contribute to osseous healing after root canal treatment.
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PMID:Human periapical granulation tissue contains osteogenic cells. 1464 94

This study evaluated the behavior of osteoblast-like cells on multigrooved surfaces consisting of a combination of microgrooves and macrogrooves. A polystyrene substrate was fabricated with multigrooves with 90-degree, V-shaped microgrooves with a 2-microm pitch cut on trapezoidal macrogrooves, which had a 50-microm ridge width, a 50-microm wall width, a 50-microm bottom width, and 25-microm depth. Smooth polystyrene substrates were also prepared as controls. Rat bone marrow cells were cultured as osteoblast-like cells on the substrates for morphological evaluation using a scanning electron microscope, and for biochemical evaluation using the quantitative reverse transcriptase-polymerase chain reaction technique for osteopontin and osteocalcin mRNA expression. After 8 days of incubation, the osteoblast-like cells were aligned parallel to the surface grooves on the multigrooved substrates. After 16 days of incubation, a dense mineralized extracellular matrix (ECM) was produced along the multigrooves. The ECM on the multigrooved surface appeared oriented more in the direction of the grooves than on the smooth surface, and trapezoid-shaped macrogrooves of the ECM were cast upside down. Although there were not significant differences, the osteopontin and osteocalcin mRNA expressions of the osteoblast-like cells on the multigrooved surfaces tended to be higher than on smooth surfaces. These results suggest that multigrooves could be used to control the orientation of mineralized ECM as well as of cells, and also to enhance the production of mineralized ECM.
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PMID:Effects of multigrooved surfaces on osteoblast-like cells in vitro: scanning electron microscopic observation and mRNA expression of osteopontin and osteocalcin. 1470 64

The purpose of the current study was to investigate the effect of different diameters of cylindrical titanium channels on human osteoblasts. Titanium samples having continuous drill channels with diameters of 300, 400, 500, 600, and 1000 microm were put into osteoblast cell cultures that were isolated from 12 adult human trauma patients. Cell migration into the drill channels was investigated by transmitted-light microscopy. The DNA content in the drill channels was measured photometrically, collagen type I production was analyzed by enzyme-linked immunosorbent assay (ELISA) and osteocalcin gene expression by reverse transcriptase-polymerase chain reaction (RT-PCR). Formation of mineralized tissue was assessed by microradiographs of histological sections. Within 20 days, cells grew an average of 838 microm (+/-128 microm) into the drill channels with a diameter of 600 microm and were significantly faster (p < 0.05) than in all other channels. Cells produced significantly more osteocalcin messenger RNA (mRNA) in 600-microm channels (p < 0.05) than they did in 1000-microm channels and demonstrated the highest osteogenic differentiation. The channel diameter did not influence collagen type I production. The highest cell density was found in 300-microm channels (p < 0.05). The DNA content of the channels linearly decreased with increasing channel diameters. After 40 days of culture, the proportion of mineralized tissue at the mouth section amounted to 6% in 300-microm channels and to 9-11% in 400-600-microm channels. In 1000-microm channels, only traces of mineralization were detected. Our data suggest that the diameter of cylindrical titanium channels has a significant effect on migration, gene expression, and mineralization of human osteoblasts.
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PMID:Growth behavior, matrix production, and gene expression of human osteoblasts in defined cylindrical titanium channels. 1470 74


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