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)

Amplification of the enterically-transmitted non-A, non-B hepatitis virus (HEV) RNA using conventional reverse transcriptase reactions followed by the polymerase chain reaction (PCR) of the cDNA has not been successful. However, after application of two different RNA capture/extraction methods we were able to amplify HEV nucleic acid from clinical samples and specimens from experimentally infected animals. The first procedure, adapted from an immune electron microscopy (IEM) technique, incorporated an immunocapture step with concentration of the virus-antibody complexes by pelleting in a Beckman airfuge. In the second method, glass powder (or size-fractionated silicon dioxide) was used to capture the RNA from its surrounding milieu by adsorption of the nucleic acid to the silicate particles. Since conventional immunoassays for HEV antigen or antibody are not currently available, the use of these RNA extraction methods, coupled with PCR techniques, will be valuable in screening clinical specimens and in further defining the course of disease using animal infectivity studies.
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PMID:Application of two RNA extraction methods prior to amplification of hepatitis E virus nucleic acid by the polymerase chain reaction. 181 58

Heteropolyoxotungstates of the Keggin class containing different heteroatoms were tested for inhibition of two strains of human immunodeficiency virus 1 (HIV-1); they exhibited varying antiviral activity. Compounds containing boron were inactive, only one of those containing phosphorus showed selective anti-viral activity, whereas all silicon-containing compounds exhibited significant anti-viral activity in C8166 cells infected with the IIIB strain. Their effectiveness was some 10-fold higher in JM cells with selectivity indices of about 2000. The silicotungstates were effective inhibitors of HIV reverse transcriptase, showing greater inhibition with RNA/DNA template primers than with DNA/DNA template.primer. Kinetic analysis demonstrated that they inhibit the enzyme by different mechanisms, as, of the four compounds examined, two competed with template.primer and two competed with deoxynucleoside triphosphate. Inhibition of DNA polymerase activity by these compounds was compared using polymerases from different sources, including human; although not necessarily most specific for HIV-1 reverse transcriptase, they did not inhibit all DNA polymerases to a similar degree.
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PMID:Anti-(human immunodeficiency virus) activity of polyoxotungstates and their inhibition of human immunodeficiency virus reverse transcriptase. 753 11

We have established two distinct human cervical cell lines, NCC16 and NCE16, after transfecting human papillomavirus type 16 (HPV16) DNA into normal human ecto-cervical and endo-cervical epithelial cells, respectively. Both lines expressed HPV16 E6 and E7 as detected by reverse transcriptase-polymerase chain reaction and northern blot hybridization. These cells have been passaged for over 100 population doublings and express strong telomerase activity. Neither cell line was tumorigenic in athymic nu/nu mice. However, both NCC16 and NCE16 developed abnormally stratified architectures following implantation with a silicon membrane sheet in the back of athymic nude mice. The former cells were pathohistologically similar to carcinoma, while the latter produced Alcian-blue positive cells, suggesting the occurrence of metaplastic changes. These distinct cell lines offer a useful model system for the study of cervical carcinogenesis and of its regulatory mechanism after HPV infection in different regions of the uterine cervix.
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PMID:Two distinct human uterine cervical epithelial cell lines established after transfection with human papillomavirus 16 DNA. 931 Jan 37

A rapid, efficient and inexpensive method was developed to concentrate poliovirus type 1 (PV1), rotavirus (RV) and hepatitis A virus (HAV) from artificially spiked samples of tap and surface water. The method consists of adsorbing the viruses to silicon dioxide (SiO2) in the presence of 0.5 mM AlCl3 and adjustment of the pH to 3.5. The silica-adsorbed virus was collected by low speed centrifugation. Viral RNA was then extracted with guanidium thiocyanate (GT), and environmental nucleases and inhibitors of reverse transcriptase and Taq polymerase were further eliminated from concentrates by sequential treatment with GT, ethanol and acetone. Subsequent RT-PCR allowed the detection of as few as 1 to 10 TCID50 of PV1, RV, and HAV in seeded 1 liter samples of tap water. The same protocol was then used with effluents from two local sewage treatment plants. These samples, found to be free of HAV, were most commonly contaminated with enteroviruses and rotaviruses. Addition of 1000 TCID50 of HAV, PV1 or RV to a second 1 liter sample, taken at the same time from the corresponding surface waters allowed detection of the input virus without discernible inhibition by amplification inhibitors. The newly established method seems amenable to scaling up and promising for virus monitoring in different water types. The method is rapid and results can be obtained within 24 to 36 hours.
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PMID:Use of silica as a carrier to recover and prepare waterborne enteric viruses for detection by RT-PCR. 963 82

This study was designed to determine the effect of Si(3)N(4) disks and particulates on human osteoblast-like MG-63 cells in vitro. The MG-63 (10(5)/mL) cells were plated onto 24-well polystyrene plates fitted with either sintered reaction-bonded (SRBSN) or reaction-bonded (RBSN) 15-mm disks. Controls consisted of wells without Si(3)N(4) disks. Cells propagated at 37 degrees C, 5% CO(2) for 48 h on Si(3)N(4) disks and control polystyrene surfaces exhibited similar proliferative capacities (7000 and 4000 cpm/10(5) cells, respectively, p > 0.05). Cells incubated with 1, 10, or 100 microgram/ml of Si(3)N(4) particles (<1.00 to 5.00 micrometer) for 24 h did not exhibit a decrease in DNA synthetic activity: 12 +/- 1.3 x 10(4), 10.5 +/- 1.5 x 10(4), and 11.0 +/- 1.7 x 10(4) cpm, respectively, compared to 11.6 +/- 2.6 x 10(4) cpm/10(5) for the control cells, as indicated by (3)H-thymidine uptake. Cells propagated on RBSN displayed increased expression of cytokines IL-1beta and TNF-alpha compared to the control cells, as shown by reverse transcriptase-polymerase chain reaction (RT-PCR). In contrast, cells propagated on SRBSN surfaces expressed the same level of IL-1beta and TNF-alpha as that of control cells. Incubation of MG-63 cells with 1-10 microgram/mL of particles did not increase IL-1beta expression. However, at 100 microgram/mL, TNF-alpha expression was greater than that of the control cells. Silicon nitride, evaluated here as disks or as particulates (1-10 microgram/mL), is biocompatible and does not hinder the proliferation or induce proinflammatory cytokine expression of human osteoblast-like MG-63 cells in vitro.
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PMID:Proinflammatory cytokine expression of IL-1beta and TNF-alpha by human osteoblast-like MG-63 cells upon exposure to silicon nitride in vitro. 1064 62

We have developed a reverse transcriptase polymerase chain reaction (RT-PCR)-based assay to detect influenza A in guano samples as part of our program to investigate ancient viral RNA from under Antarctic Adelie penguin (Pygoscelis adeliae) colonies. Of five extraction protocols tested (RNeasy, GTC TRIZOL, GTC Silica, Rnaid, and AGPC), AGPC proved to be the most consistent and sensitive to low concentrations of influenza, but further purification with commercial viral nucleic acid spin filter system was still required to remove remaining PCR inhibitors. RT-PCR was then performed on the eluent and 650 bases of the M1 gene were amplified. The assay was found to be able to detect as little as 100 microl of 0.1 hemagglutination units (HU)/ml influenza.
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PMID:Development of a highly sensitive screen for influenza A in guano and its application in the search for ancient RNA preserved under Antarctic Adelie penguin colonies. 1457 42

Intratracheal instillation studies have shown that exposure to silicon carbide whiskers (SiCW), an asbestos substitute, produces pulmonary fibrotic changes, suggesting that SiCW might have fibrogenic potential. It has been theorized that Clara cell secretory protein (CCSP) plays a critical role in regulating the acute inflammatory response in the lung. The present study was conducted to investigate the time course of the expression of CCSP in lungs exposed to SiCW in vivo. Male Wistar rats were administered 2 mg or 10 mg of SiCW suspended in saline by a single intratracheal instillation and were sacrificed at 3 days, 1 week, 1 month, 3 months and 6 months of recovery time. The expression of CCSP was observed by reverse transcriptase-polymerase chain reaction (RT-PCR), Western blot and immunostaining. Exposure to 10 mg of SiCW decreased in levels of CCSP mRNA at 3 days, 1 week, 1 month and 6 months following intratracheal instillation. The protein level of CCSP in SiCW-exposed rats was decreased at 1 day, 3 days and 1 month after a single instillation of 2 and 10 mg. These findings suggest that CCSP are involved not only in the acute injury but also in the chronic injury of the lung exposed to SiCW.
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PMID:Expression of Clara cell secretory protein in the lungs of rats exposed to silicon carbide whisker in vivo. 1458 Aug 98

Bioactive materials have previously been used to coat implants. In a new development for bioactive materials, a silica-ceramic mixture was found to alleviate pain (Lee, Poster presented at the Ninth World Congress of Gynecological Endocrinology, Hongkong, 2001. Poster session (p47)). Here, we hypothesized that silica-ceramic can reduce the expression and activity of cyclooxygenase 2 (COX2) or cytokines associated with inflammation. The production of COX2 and proinflammatory cytokines was investigated by reverse transcriptase (RT)-PCR and ELISA assay in macrophages stimulated by lipopolysaccharide (LPS). Silica-ceramic had no effect of COX2 expression and prostaglandin production in macrophages. However, silica-ceramic suppressed the synthesis of cytokines involved in inflammation, in particular, the expression of IL-1beta and IL-6 was reduced at the transcriptional and translational levels. The involvement of NF-kappaB in the suppression of cytokines by silica-ceramic was examined by luciferase reporter assay. The NF-kappaB activity stimulated by LPS was inhibited by 20-60% with silica-ceramic compared with treatment with LPS alone. We suggest that inhibition of NF-kappaB activity by silica-ceramic might cause the attenuation of proinflammatory cytokine expression in macrophages. In conclusion, silica-ceramic could be an alternative approach to regulate the inflammation process.
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PMID:Silica-ceramic suppresses the expression of proinflammatory cytokines induced by lipopolysaccharide in macrophages. 1713 51

In this study we purposed an alternative method to study the angiogenic and invasive potential of neuroblastoma cell suspensions implanted on the chick embryo chorioallantoic membrane (CAM) surface. Neuroblastoma cells were seeded in Matrigel and thereafter the suspension was pipetted onto the CAM surface at day 8 of incubation inside a silicon ring previously loaded onto the CAM surface. Four days after implantation, the silicon ring was removed and the angiogenic and invasive response were studied morphologically at macroscopic and microscopic levels and by reverse transcriptase-polymerase chain reaction (RT-PCR) by using human and chicken primers for several angiogenic cytokines, namely vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor-2 (FGF-2), angiopoietin-1 (ANG-1), hypoxia inducible factor-2alpha (HIF-2alpha), and for an endogenous angiostatic molecule, namely endostatin. Results showed that: (1) Neuroblastoma cells induced an angiogenic response in the CAM assay comparable to that induced by FGF-2; (2) neuroblastoma cells are packed inside Matrigel or are recognizable in the CAM mesenchyme; (3) Angiogenic activity of neuroblastoma cells is associated to an high expression of the transcripts of human VEGF-A, FGF-2, ANG-1 and HIF-2alpha and to a low expression in the transcript of a human endostatin while in the control specimens there is no expression of both angiogenic and angiostatic molecules; and (4) the expression of the transcripts of the same chicken angiogenesis stimulators and inhibitor is unmodified in treated and control specimens. Overall, these data indicate that neuroblastoma cells growth on the chick CAM express characteristics of the human disease. This experimental model could be employed for further research on human tumor progression and anti-angiogenic molecules screening.
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PMID:An alternative in vivo system for testing angiogenic potential of human neuroblastoma cells. 1915 May 83

The objective of this study was to investigate potential role of taurine and niacin supplementation, and their combination, in an in vitro model of silica-induced, macrophage-mediated pulmonary fibroblast proliferation. Human monocytic cell line (THP-1 cell) was primed to differentiation into macrophages by phorbol myristate acetate (PMA). PMA-primed THP-1 cells were subjected to silicon dioxide exposure. Other PMA-primed THP-1 cells incubated with taurine and niacin concentration gradients, respectively, and then were treated with silicon dioxide for 6 hours. Collected THP-1 supernatants preconditioned with taurine and niacin gradients were added to human pulmonary WI-38 cells to evaluate proliferative activity. Transforming growth factor (TGF)-beta1 mRNA in macrophages and protein level in supernatant were determined by reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Taurine- and niacin-preconditioned macrophages were more resistant to silica-induced TGF-beta1 up-regulation than macrophages without precondition. Furthermore, medium conditioned with supernatant from silica-exposed macrophages following taurine and niacin pretreatment could facilitate inhibition of pulmonary fibroblast proliferation. Moreover, the above effects could be accentuated by the combination of taurine and niacin. Down-regulation of TGF-beta 1 expression in macrophages by taurine and niacin could attenuate silica-induced pulmonary fibroblasts proliferation in vitro, which may be of therapeutic potential for early stage silicosis.
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PMID:Attenuation of silica-induced pulmonary fibroblasts proliferation by taurine and niacin in vitro. 1933 3


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