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)

Cutaneous eruptions related to hepatitis C virus (HCV), a major cause of hepatitis in the setting of blood transfusion, intravenous drug abuse, organ transplantation, and hemodialysis, are typically reported as isolated cases. We encountered 35 cases of HCV infection associated with cutaneous eruptions. The present study evaluates paraffin-embedded, formalin-fixed tissue sections stained with hematoxylin and eosin from biopsy specimens of skin lesions from 35 patients seropositive for HCV. In 20 cases, reverse transcriptase polymerase chain reaction (RT-PCR) was performed using a probe for HCV RNA; the RNA was detected through the action of alkaline phosphatase on the chromogen nitroblue tetrazolium and bromochloroindolyl phosphate. The clinical spectrum comprised dermatomyositis-like photodistributed eruptions, palpable purpura, folliculitis, violaceous and perniotic acral lesions, ulcers, nodules, and urticaria. Lesions were also classified histopathologically by the dominant reaction pattern: vasculopathies of neutrophilic, lymphocytic, and granulomatous vasculitis and pauci-inflammatory subtypes (15 patients); palisading granulomatous inflammation (3 patients); sterile neutrophilic folliculitis (5 patients); dermatitis herpetiformis (1 patient); lobular panniculitis composed of neutrophilic lobular panniculitis in 2 patients and benign cutaneous polyarteritis nodosa in 1 patient; neutrophilic dermatoses, including neutrophilic urticaria, neutrophilic eccrine hidradenitis, and pyoderma gangrenosum (3 patients); interface dermatitis (3 patients); and low-grade lymphoproliferative disease of B-cell lineage representing marginal zone lymphoma in 1 patient and a clonal plasmacellular infiltrate in another patient. In most cases, whereas 1 of the aforementioned disorders defined the dominant reaction pattern, there was an accompanying secondary reaction pattern, defining a hybrid picture. Endothelial changes including endothelial cell enlargement and effaced heterochromatin with margination of the chromatin to the nuclear membrane were seen in several cases; in some cases similar cytopathic changes also involved the supporting pericytes, eccrine ductular cells, or keratinocytes. The RT-PCR analyses in 8 of 20 cases examined revealed HCV RNA expression in a focal, weak fashion in endothelia and perivascular inflammatory cells in those cases showing vasculopathic changes. Viral parasitism of endothelia may be important in cutaneous lesional propagation in the setting of HCV infection. Cross-reactivity between endogenous and viral antigens, leading to cellular and/or type II immune reactions; viral tropism to B lymphocytes, resulting in B cell expansion with resultant autoantibody production; and circulating immune complexes containing monoclonal cryoglobulins may also be of pathogenetic importance. Tropism of the virus to B lymphocytes provides a mechanism for the development of low-grade clonal B cell lymphoproliferative disease in this setting.
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PMID:The dermatopathologic manifestations of hepatitis C infection: a clinical, histological, and molecular assessment of 35 cases. 1282 11

The purpose of this study was to investigate pulp cell responses during hypoxia and reoxygenation. Pulp tissues obtained from beagle dogs were cultured. In the control group, pulp cells were incubated in normoxic conditions (20% O2) for 1-4 d. In the hypoxia group, pulp cells were incubated under hypoxic conditions (2% O2) for 1-4 d. In the reoxygenation group, pulp cells were first incubated under hypoxic conditions for 24 h, and were then incubated in normoxic conditions (20% O2) for one to three additional days. Cell viability, MTT (3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) reduction assay, cellular proliferation, and alkaline phosphatase (ALPase) activity were determined. Expression of heat shock protein 70 (HSP70) and vascular endothelial growth factor (VEGF) was analysed by Western blotting. Hypoxia inducible factor-1alpha (HIF-1alpha) in pulp cells was analysed by reverse transcriptase polymerase chain reaction (RT-PCR). The cell growth rate and ALPase activity were significantly higher in the hypoxia group than in the control group. After reoxygenation, cellular proliferation and ALPase activity decreased to the level of the control group while HSP70 expression increased. Hypoxia inducible factor-1alpha expression was detected in pulp cells, and VEGF expression (which is regulated by HIF-1alpha) increased under hypoxic conditions. These results suggest that dynamic responses to hypoxia and reoxygenation occur in pulp cells.
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PMID:Pulp cell responses during hypoxia and reoxygenation in vitro. 1288 99

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

The theoretical possibility that exposure of a solid malignancy to high-intensity focused ultrasound (US), or HIFU, could lead to an increased rate of metastasis still remains. Using reverse transcriptase polymerase chain reaction (RT-PCR), the potential risk of hematogenous dissemination was assessed in HIFU-treated patients with solid malignancy. RT-PCR can demonstrate the presence or absence of specific RNA fragments. On the day before HIFU ablation, 5-mL peripheral blood samples were collected, and again 5 to 7 days after HIFU, from 26 enrolled patients (hepatocellular carcinoma, HCC: 10; osteosarcoma: 16). Total RNA was isolated and RT-PCR was performed to analyze the mRNA expression of (alpha-fetoprotein (AFP) and bone-specific alkaline phosphatase (BALP) genes. Positive AFP mRNA expression was preoperatively detected in 8 of 10 patients with HCC. In the postoperative specimens, positive expression was also detected in 8 of 10 patients. In 2 patients, circulating tumor cells were found preoperatively, but not postoperatively. Conversely, 2 patients with no circulating tumor cells preoperatively were found to have circulating tumor cells after HIFU. Of 16 osteosarcoma patients, 12 patients had circulating tumor cells and 4 had none. After HIFU treatment, 2 of the 12 patients had converted from presence to absence of circulating cells and the remaining 4 patients remained negative. It is concluded that patients undergoing complete HIFU ablation may demonstrate conversion from presence to absence of circulating tumor-specific marker mRNA, and that HIFU would not enhance the potential risk of metastasis in patients with malignant diseases.
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PMID:Circulating tumor cells in patients with solid malignancy treated by high-intensity focused ultrasound. 1512 Dec 53

A quantitative reverse transcriptase polymerase chain reaction (RT-PCR) method, employing internal standard (IS) RNA and a simplified chemiluminometric hybridization assay, is described for the determination of prostate-specific membrane antigen (PSMA) mRNA. The recombinant RNA IS has the same binding sites and size as the amplified PSMA mRNA. Biotinylated PCR products (263 bp) from PSMA mRNA and RNA IS are captured in microtiter wells coated with streptavidin, and hybridized with alkaline phosphatase-conjugated probes. The bound alkaline phosphatase (AP) is measured by using a chemiluminogenic substrate. The ratio of the luminescence values obtained for PSMA mRNA and the RNA IS is a linear function of the initial amount of PSMA mRNA present in the sample before RT-PCR. The linear range extends from 500 to 5,000,000 PSMA mRNA copies and the overall reproducibility of the assay, including RT-PCR and hybridization, ranges from 7.4 to 16.6%. Samples containing total RNA from PSMA-expressing LNCaP cells give luminescence ratios linearly related to the number of cells in the range 0.5-5,000 cells.
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PMID:High-throughput chemiluminometric determination of prostate-specific membrane antigen mRNA in peripheral blood by RT-PCR using a synthetic RNA internal standard. 1527 42

Owing to their plasticity and high proliferation capacity in vitro, mesenchymal stem cells (MSC) isolated from human bone marrow are promising candidates for use in tissue engineering approaches for the repair or replacement of mesenchymal tissues such as bone, cartilage or tendon. In keeping with the tissue engineering concept, these cells are cultivated on three-dimensional (3D) scaffolds to replace 3D tissue defects. Among the scaffolds tested for tissue engineering of bone, those containing phosphorus and calcium, as natural bone does, are the most promising candidates for this purpose. In this study, MSC from five patients were isolated from bone marrow. After in vitro expansion, cells were cultivated and differentiated towards the osteogenic lineage on mineralized collagen sponges and alpha-tricalcium phosphate (alpha-TCP). To analyze how appropriate these scaffolds would be for tissue engineering purposes, we established an in vitro characterization system to describe seeding efficiency, cell distribution and proliferation behavior on each scaffold. Real-time reverse transcriptase polymerase chain reaction quantification of important genes involved in osteogenic differentiation [e.g. bone sialoprotein (BSP), bone morphogenic protein 2, alkaline phosphatase and osteocalcin] was used to monitor the differentiation process of cells seeded on mineralized collagen and alpha-TCP. Using this in vitro characterization, we were able to demonstrate effective 3D growth of MSC on both scaffolds investigated. Improved osteogenic differentiation was observed on the scaffolds as compared to control monolayers. Of the two matrices, mineralized collagen was superior to alpha-TCP with regard to seeding efficacy (98 vs. 67%, p = 0.0003), increase in osteogenic marker genes (BSP expression on day 24, Pcollagen vs. TCP = 0.046) and 3D cell alignment (cell infiltration up to 500 vs. 200 microm). In conclusion, our data suggest that mineralized collagen is a promising candidate for use as a scaffold in tissue engineering of bone.
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PMID:Evaluation of mineralized collagen and alpha-tricalcium phosphate as scaffolds for tissue engineering of bone using human mesenchymal stem cells. 1529 81

Mesenchymal progenitor or stem cells (MPCs) isolated from fetal blood, liver, and bone marrow are a population of multipotential cells that can proliferate and differentiate into multiple mesodermal tissues including bone, cartilage, muscle, ligament, tendon, fat, and stroma. The objective of this study was to isolate and characterize MPCs in the human umbilical cord. The suspensions of endothelial and subendothelial cells in cord vein were collected and cultured in M199 supplemented with 10% fetal bovine serum (FBS). Of 50 umbilical cord samples, 3 had numerous fibroblastoid cells morphologically distinguishable from endothelial cells. Fibroblastic cells displayed lack of expression of vWF, Flk-1, and PECAM-1, indicating the endothelial cell-specific marker. To investigate the differentiation potentials, the cells were cultured in adipogenic or osteogenic medium for 2 weeks. Fibroblast-like cells treated with adipogenic supplementation showed Oil red O-positive staining and expressed adipsin, FABP4, LPL, and PPARgamma2 genes by reverse transcriptase polymerase chain reaction (RT-PCR). In osteogenic differentiation, alkaline phosphatase activity and calcium accumulation were detected. RT-PCR studies determined that Cx43, osteopontin, and Runx2 genes were expressed in the osteogenic cultures. Among three cell lines cultured continuously for passage 10, two had normal karyotypes; however, one retained a karyotype of mos 46,XY[19]/47,XY,+mar[3]. These observations suggest that MPCs are present in human umbilical cord and possess several typical traits of MPCs.
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PMID:Mesenchymal progenitor cells in the human umbilical cord. 1537 3

MEPE, 56.6 kDa protein isolated from tumors associated with hypophosphatemic osteomalacia, increases renal phosphate excretion and is expressed in normal human bone cells. AC-100, a central 23-amino acid fragment of MEPE, contains motifs that are important in regulating cellular activities in the bone microenvironment. Thus, we assessed in vitro effects of AC-100 on multipotential normal human marrow stromal (hMS) cells that have the capacity to differentiate into mature osteoblasts. Proliferation was quantified by [H3]thymidine uptake and cell counting and differentiation by the levels of mRNA for the alpha2-chain of type I procollagen (COL1A2), alkaline phosphatase (AP), and osteocalcin (OC) measured using real time reverse transcriptase PCR (RT-PCR) and by the formation of mineralized nodules. AC-100 increased proliferation by 257 +/- 89% (P < 0.005), increased gene expression of COL1A2 by 339 +/- 85% (P < 0.005), AP by 1,437 +/- 40% (P < 0.001), and OC by 1,962 +/- 337% (P < 0.001). In addition, it increased mineralized nodule formation by 81 +/- 14% (P < 0.001) in a dose- and time-dependent fashion. In equimolar dosages, the parent compound, MEPE, had the full activity of the AC-100 fragment. AC-100 elicited a comparable response to both IGF-I and BMP-2 with respect to proliferation and differentiation of hMS cells. Using gene expression microarray analysis, we demonstrated that AC-100 increased (by approximately 3-fold) the mRNA for cyclooxgenase-2 (COX-2), an inducible enzyme required for prostaglandin synthesis. Moreover, NS-398, a specific inhibitor of COX-2 action completely blocked AC-100-induced increases in proliferation and differentiation. Thus, AC-100 has potent anabolic activity on osteoblast precursor cells in vitro and these effects require the induction of COX-2.
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PMID:A fragment of the hypophosphatemic factor, MEPE, requires inducible cyclooxygenase-2 to exert potent anabolic effects on normal human marrow osteoblast precursors. 1544 21

Although beta-alanyl-L-histidinato zinc (AHZ) can promote osteoblast differentiation, the molecular mechanism responsible is not fully understood. The purpose of this study was to determine the effect of AHZ on undifferentiating mesenchymal cells. C2C12, a typical pluripotential mesenchymal cell line, was used. The cells were cultured in 5% serum-containing medium to induce differentiation, either with or without the addition of AHZ. Cell lineage was determined by immunostaining of type II myosin heavy chains, alkaline phosphatase (ALPase) activity, mRNA expression of cellular phenotype-specific markers using semi-quantitative reverse transcriptase-polymerase chain reaction, and core binding factor alpha1/runt-related transcription factor-2 (Cbfa1/Runx2) protein synthesis using Western blot analysis. C2C12 cells cultured in the presence of AHZ were strongly inhibited from developing into myoblasts, and showed high ALPase activity that was approximately double that in the vehicle. The expression of mRNA for Cbfa1/Runx2, ALPase, Sox9 and type X collagen was increased markedly by the AHZ-stimulated medium, whereas that of desmin and MyoD mRNA was drastically decreased. AHZ increased Cbfa1/Runx2 protein expression substantially. These results provide clear evidence that AHZ converts the differentiation pathway of C2C12 cells to the osteoblast and/or chondroblast lineage.
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PMID:Effect of beta-alanyl-L-histidinato zinc on the differentiation of C2C12 cells. 1555 64

In the present study, we investigated the protective effect of Lycium chinense Miller (Solanaceae) fruit (LFE) against CCl(4)-induced hepatotoxicity and the mechanism underlying these protective effects in rats. The pretreatment of LFE has shown to possess a significant protective effect by lowering the serum aspartate and alanine aminotransferase (AST and ALT) and alkaline phosphatase (ALP). This hepatoprotective action was confirmed by histological observation. In addition, pretreatment of LFE prevented the elevation of hepatic malondialdehyde (MDA) formation and the depletion of reduced glutathione (GSH) content and catalase activity in the liver of CCl(4)-injected rats. The LFE also displayed hydroxide radical scavenging activity in a dose-dependent manner (IC(50) = 83.6 microg/ml), as assayed by electron spin resonance (ESR) spin-trapping technique. The expression level of cytochrome P450 2E1 (CYP2E1) mRNA and protein, as measured by reverse transcriptase-polymerase chain reaction (RT-PCR) and western blot analysis, was significantly decreased in the liver of LFE-pretreated rats when compared with that in the liver of control group. Based on these results, it was suggested that the hepatoprotective effects of the LFE might be related to antioxidative activity and expressional regulation of CYP2E1.
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PMID:Protective effect of Lycium chinense fruit on carbon tetrachloride-induced hepatotoxicity. 1561 74


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