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)

Certain osteoclastic markers (multinucleation and tartrate-resistant acid phosphatase) were induced in human leukemia HL-60 cells by treatment with 10(-7) M 1,25-dihydroxy-vitamin D3 [1,25(OH)2D3] for 10 days. However, no formation of pits on a bone substrate by vitamin-treated HL-60 cells was detected. Expression of calcitonin receptors (CTR), another osteoclastic marker, was examined by means of the reverse transcriptase polymerase chain reaction. The human CTR-cDNA (T47D isotype) was amplified from untreated HL-60 cells, but not from cells treated with 1,25(OH)2D3. The CTR mRNA disappeared within 24 h after the treatment. Thus, 1,25(OH)2D3-differentiated HL-60 cells failed to show two intrinsic characteristics of osteoclasts, pit formation on a bone substrate and expression of CTR. We then examined the expression of CTR on established human leukemia cell lines. The CTR mRNA was expressed in myeloblastic ML-1 and promyelocytic HL-60 leukemia cells but not in more mature macrophage-like cell lines, U-937 and THP-1 cells. Neither B cell leukemia BALL-1, T cell leukemia Jurkat, promegakaryoblastic leukemia Meg-J, nor cervix uteri carcinoma HeLa S3 cells amplified the CTR products. The cDNA of BIN67-isotype CTR, that has an additional 16-amino acid insert in the putative first intracellular loop of T47D-type CTR [Kuestner et al. (1994) Mol. Pharmacol. 46, 246-255], was amplified by neither strain tested. It was suggested that the T47D-type CTR is a novel differentiation antigen of immature myeloid lineage cells.
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PMID:Expression of calcitonin receptors on human myeloid leukemia cells. 854 84

Osteocytes have been proposed to be the cells primarily responsible for sensing the effects of mechanical loading in bone. Osteocytes respond to loading in vivo, and have been shown to express osteotropic agents and their receptors, and cell/matrix adhesion molecules in vitro, but the functional significance of such findings is not clear. One obstacle to increased understanding of the role of osteocytes in the regulation of bone mass is that the cells are not easily accessible for study. In situ studies are difficult, and although it is possible to extract and culture osteocytes from neonatal bones, the responses of such cells might be very different from those in older bones in situ. We have developed a technique to investigate osteocyte gene expression in vivo, using the reverse transcriptase linked polymerase chain reaction (PCR), and have shown that they express mRNA for beta-actin (beta-ACT), osteocalcin (OC), connexin-43 (Cx43), insulin-like growth factor I (IGF-I), c-fos and c-jun, but not tumor necrosis factor alpha (TNF-alpha) or tartrate-resistant acid phosphatase (TRAP). The principle behind the method is that after removal of the periosteum, tangential cryostat sections of a tubular bone contain RNA only from osteocytes and a very small number of endothelial cells as long as the marrow cavity is not broached. Using this method, we have investigated gene expression in cells from rat ulnar cortical bone under forming and resorbing bone surfaces. In addition, we have investigated the effect on gene expression of mechanical loading which, if repeated daily, initiates new bone formation on quiescent or resorbing surfaces. Although the expression of the genes we have studied in osteocytes is different from those expressed by the periosteal surfaces overlying the cortex, we have not detected loading-related changes in osteocyte gene expression in any cortical bones. This may be because of the extreme sensitivity of the PCR technique which can only resolve large differences in expression. The use of quantitative methods in the future may allow demonstration of regulated gene expression in osteocytes.
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PMID:Constitutive in vivo mRNA expression by osteocytes of beta-actin, osteocalcin, connexin-43, IGF-I, c-fos and c-jun, but not TNF-alpha nor tartrate-resistant acid phosphatase. 885 45

Pigmented villonodular synovitis (PVNS) and the histologically related lesion giant cell tumor of tendon sheath (GCTTS) are idiopathic, proliferative lesions that can induce osteolysis and formation of bone cysts. These lesions contain two predominant cell types: mononuclear polyhedral cells and multinucleated cells (MNCs). Previous studies demonstrated that the mononuclear cells exhibit phenotypic features consistent with derivation from a monocyte/macrophage lineage. The cell lineage of the MNCs and their relationship to osteoclasts are not known. To characterize the MNCs in these lesions and to establish the relationship of these MNCs to osteoclasts, histological sections from six cases of PVNS and two cases of GCTTS were studied. Mononuclear cells expressed CD14 and HLA-DR, in keeping with their relationship to cells of the monocyte/macrophage lineage. Characterization of the MNCs revealed features associated with an osteoclast phenotype. Seven of the eight specimens contained MNCs that were intensely tartrate-resistant acid phosphatase positive; approximately 5% of the mononuclear cells were tartrate-resistant acid phosphatase positive, and these tended to surround MNCs. MNCs in both lesions reacted strongly with the 23C6 monoclonal antibody that recognizes the alpha V beta 3 integrin (the vitronectin receptor), as did several mononuclear cells surrounding the MNCs. Most MNCs did not express CD14 or HLA-DR. Expression of receptors for calcitonin, a marker for osteoclasts, was detected on MNCs after incubation of sections with 125I-labeled salmon calcitonin and emulsion autoradiography. MNCs in four of six PVNS and two of two GCTTS samples demonstrated specific calcitonin binding. Expression of mRNA for calcitonin receptor was confirmed in all cases by reverse transcriptase polymerase chain reaction. These results demonstrate that MNCs in PVNS and GCTTS express phenotypic features of authentic osteoclasts and suggest that osteoclast-like multinucleated cells can arise in synovial soft tissues remote from bone.
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PMID:Multinucleated cells in pigmented villonodular synovitis and giant cell tumor of tendon sheath express features of osteoclasts. 909 94

An in vitro culture system to generate human osteoclasts (OC) was recently described in which OC precursors in the human peripheral blood mononuclear cell (PBMC) population differentiate in the presence of murine ST-2 stromal cells. We used this culture system to define the cytokine environment in which human OC form and to determine the separate contributions of the stromal and hematopoietic elements. We designed a panel of reverse transcriptase-polymerase chain reaction (RT-PCR) primers that specifically amplify the respective murine or human mRNA species that correspond to cytokines and their cognate receptors previously shown to promote or inhibit OC differentiation. ST-2 cells were cocultured with human PBMC for up to 21 days in the presence of 1alpha,25(OH)(2) vitamin D(3), dexamethasone, and recombinant human macrophage-colony stimulating factor (M-CSF). OC formation was monitored by the appearance of cells that were positive for tartrate-resistant acid phosphatase (TRAP) and able to form resorption lacunae on slices of dentine. We found that the ST-2 cells in these cultures express messenger RNA (mRNA) encoding a repertoire of many of the reported osteoclastogenic factors (interleukins [IL]-1/IL-1R1, IL-11, IL-6/IL-6R, and IL-17 transforming growth factor [TGF]-beta), as well as the recently described OC differentiation factor (ODF/TRANCE/RANKL). The stromal cells also expressed mRNA encoding two molecules shown to be inhibitory to osteoclastogenesis, osteoprotegerin (OPG) and IL-18. OPG, IL-1, IL-1R1, IL-6, IL-6R, IL-11R, IL-17, IL-18, IL-18R, TGF-beta, and M-CSF were expressed by both the stromal cells and the PBMC. Expression of mRNA encoding RANK, IL-1R2, and c-fms, was specific for the PBMC. In addition, PBMC were found to express sIL-6R, granulocyte macrophage (GM)-CSF, GM-CSFRalpha, and tumor necrosis factor (TNF)-alpha. Whereas this indicated that human OC formation occurs in a complex environment of many positive and negative influences, we identified three apparent features of the cytokine environment that may be a characteristic of normal osteoclast formation. First, the ratio of mouse ODF:OPG mRNA was found to increase during the cocultures, consistent with a key role for ODF in the promotion by stromal cells of OC formation. Second, we found that mRNA encoding IL-1 and IL-17, as well as IL-6 and sIL-6R, were coordinately expressed by the PBMC. Third, analysis of the culture medium showed that the PBMC secreted IL-1, IL-6, and TNF-alpha protein only in coculture with ST-2 cells during the first few days of osteoclast development. We conclude that human OC formation occurs in a complex environment of many positive and negative influences; however, these are likely to be strictly regulated by a coordinated cytokine response of both stromal and hematopoietic cells.
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PMID:Coordinated cytokine expression by stromal and hematopoietic cells during human osteoclast formation. 1083 38

Calcifying tendinitis of rotator cuff tendons is a common and painful condition caused by ectopic calcification in humans. To examine the involvement of osteopontin (OPN), a potent regulator of calcium deposition on connective tissues, localization and expression of OPN protein and messenger (m)RNA were investigated in human tissue samples of calcified rotator cuff tendons. Immunohistochemistry demonstrated that OPN was localized in cells surrounding the calcified area. OPN was localized in two distinct cell types, i.e., fibroblast-like cells negative for CD68 and tartrate-resistant acid phosphatase (TRAP) and multinucleated macrophages positive for CD68 and TRAP. In situ hybridization revealed that the mRNA expression of OPN in these cells coincided with the immunohistochemistry results, and these results were supported by reverse transcriptase polymerase chain reaction analysis using human OPN-specific oligonucleotides. Cells located away from the calcified area did not express OPN. The present findings indicate the involvement of OPN in the process of calcification of rotator cuff tendons and suggest that OPN plays a role in such painful disorders through the actions of at least two cell types.
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PMID:Localization and expression of osteopontin in the rotator cuff tendons in patients with calcifying tendinitis. 1146 94

Conventional hormone replacement therapy acts primarily by preserving bone, but cannot restore lost bone in women with established osteoporosis. Studies in rodents have shown that high doses of estrogens have anabolic skeletal effects, and recent observations in a group of women treated long term with high doses of estrogen indicated that similar effects occur in humans. This study examines the hypothesis that locally produced growth factors, including transforming growth factor-beta (TGF-beta) and platelet-derived growth factors (PDGFs), are involved in mediating the anabolic effects of high-dose estrogen. Transiliac-crest bone biopsies were taken from ten women, aged 52-67 years (mean 58 years), who had been treated with high-dose estrogen for 15 years. Control samples were obtained from four age-matched postmenopausal women not receiving estrogen therapy. TGF-betas and PDGFs were analyzed for mRNA and protein expression by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. Results showed both TGF-beta1 and -beta2 mRNA, expressed as a ratio to GAPDH, were increased in the estrogen-treated group with an eightfold increase for TGF-beta1 (0.258 +/- 0.246 [mean +/- SD] vs. 0.032 +/- 0.053 in the control group, p = 0.02) and a twofold increase for TGF-beta2 (p = n.s.). TGF-beta3 analysis showed only negligible amounts in both groups. Protein expression levels for TGF-beta1, -beta2, -betaRI and -RII were higher in the estrogen-treated group than in controls, the most marked effects being seen for TGF-beta1. PDGF-A protein expression was also significantly higher in osteoblasts and osteocytes in women treated with estrogen, whereas PDGF-B showed only modest differences. The percentage of bone surface occupied by osteoclasts, as determined by tartrate-resistant acid phosphatase (TRAP) staining, was significantly reduced in the estrogen-treated group (p = 0.001). These results demonstrate that high-dose estrogen therapy is associated with increased TGF-beta, TGF-betaR, and PDGF synthesis and decreased osteoclast activity, consistent with the hypothesis that these growth factors may mediate the actions of estrogen in bone.
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PMID:Mechanisms by which high-dose estrogen therapy produces anabolic skeletal effects in postmenopausal women: role of locally produced growth factors. 1155 64

A variety of metabolic complications have been reported to be associated with highly active antiretroviral therapy (HAART), including osteopenia and osteoporosis. In this study, we determine the effects of zidovudine (AZT), a nucleoside reverse transcriptase inhibitor, on osteoclastogenesis in a cultured mouse macrophage preosteoclast cell line (RAW264.7), in mouse primary bone marrow macrophage-monocyte precursors, and on bone mineral density in mice. The results indicate that AZT induces an increase in osteoclastogenesis in the mouse preosteoclast cell line and in mouse bone marrow osteoclast precursors in the presence of RANKL. This increased osteoclastogenesis is dependent upon the concentration of AZT. AZT increases the promoter activity of tartrate-resistant acid phosphatase (TRAP) and the binding and function of the nuclear transcription protein, NF-kappaB, in RAW264.7 cells. Therefore, the effect of AZT is mediated, at least in part, by enhancing RANKL-mediated osteoclastogenesis. Bone mineral density (BMD) in AZT-treated mice is decreased and histopathology shows marked osteopenia. These results support an important role of AZT-stimulated osteoclastogenesis in HAART-induced osteopenia.
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PMID:AZT enhances osteoclastogenesis and bone loss. 1524 37

Tibial dyschondroplasia (TD) is a metabolic cartilage disease of young poultry in which endochondral bone formation is disrupted leading to the retention of a non-calcified, avascular plug of cartilage in the tibial growth plate. Chicks aged 7 days were fed either a control diet or one containing thiram 100 ppm for 48 h to induce TD. Cell multiplication in the growth plate was determined thereafter with bromodeoxyuridine (BrdU) labelling, and metabolic changes by measuring alkaline phosphatase (ALP), tartrate-resistant acid phosphatase (TRAP), and glutathione (GSH) activities. The effect on chondrocyte maturation was examined by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis of gene expression. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) and DNA fragmentation were used to determine the effects of thiram on cell survival. The results showed that thiram-induced TD was not due to the multiplication of cells in the post-proliferative zones. Thiram did not affect ALP activity, which would have indicated a loss of calcification potential, but it reduced both TRAP and the glutathione concentrations, suggesting that the growth plate metabolism and remodelling functions were adversely affected. Thiram appeared to have no effect on the expression of type X collagen, transglutaminase, RUNX2, or matrix metalloproteinase-2 (MMP) genes suggesting that it did not alter the maturation potential of chondrocytes. On the contrary, the expressions of MMP-13 and vascular endothelial growth factor (VEGF) genes were "up-regulated," suggesting that thiram has pro-angiogenic activity. However, TUNEL assay showed that thiram induced endothelial cell apoptosis in the capillary vessels of the growth plates, as early as 10 days of age, when TD was not visually evident. The vascular death increased on subsequent days accompanied by massive death of chondrocytes in the transition zone of the growth plate. The induction of apoptosis in the growth plate was also demonstrated by DNA fragmentation. It was concluded that thiram induced TD not through an increase in the multiplication of chondrocytes in the transition zone and not by altering the expression of genes causing the arrest of chondrocytes in a prehypertrophic state, but by creating a metabolic dysfunction which led to the destruction of blood capillaries in the transition zone chondrocytes.
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PMID:Changes in the tibial growth plates of chickens with thiram-induced dyschondroplasia. 1589 90

Nasal inverted papilloma is a rare benign tumor of epithelial origin with aggressive evolution, bone destruction, recurrence, and malignant transformation. Msx2 is a homeobox gene implicated in organ development, bone metabolism, and tumorigenesis. Using reverse transcriptase-polymerase chain reaction and immunohistochemistry, Msx2 expression was examined in nasal inverted papilloma and in nontumorigenic tissue counterparts. For the first time, Msx2 was detected in all inverted papillomas but not in the nasal polyps or in the normal mucosa. The protein expression level was directly and significantly associated with tumor recurrence. Furthermore, Msx2 was associated with bone resorption markers receptor activator of nuclear factor-kappa B ligand and tartrate-resistant acid phosphatase, suggesting a role in osteolysis. In conclusion, Msx2 expression may represent a useful prognostic marker in inverted papilloma.
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PMID:Nasal inverted papilloma expresses the muscle segment homeobox gene Msx2: possible prognostic implications. 1818 85

The present study was a molecular analysis of the initial differentiation of osteoclast precursor RAW264.7 cells on titanium specimens. RAW264.7 cell line was cultured on titanium specimens of which the surfaces were finished by wet grinding with 2000-, 1200-, 600-, or 180-grit waterproof abrasive paper. Total RNA was extracted from cells cultured in the presence or absence of Receptor Activator of NF-kappaB Ligand (RANKL), prior to cDNA synthesis for real-time quantitative reverse transcriptase-polymerase chain reaction analysis. Titanium surfaces initially enhanced the expression of osteoclast differentiation markers including tartrate-resistant acid phosphatase and cathepsin K in RAW264.7 cells cultured with RANKL stimulation, in a roughness-dependent manner. The mRNA expressions of both RANKL receptor, RANK, and its adapter protein TNF receptor-associated factor 6 (TRAF6) increased when RAW264.7 cells were cultured on titanium specimens with roughened surfaces, as compared with that of control specimen with a polished surface. These results, taken together, suggested that titanium surface roughness facilitated osteoclast differentiation through the activation of the RANK-TRAF6 signaling network.
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PMID:Titanium surface roughness accelerates RANKL-dependent differentiation in the osteoclast precursor cell line, RAW264.7. 1820 77


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