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1.
Antisense oligonucleotide inhibition of hepatitis C virus gene expression in transformed hepatocytes
R Hanecak V Brown-Driver MC Fox RF Azad S Furusako C Nozaki C Ford H Sasmor KP Anderson 《Canadian Metallurgical Quarterly》1996,70(8):5203-5212
Genetic and biochemical studies have provided convincing evidence that the 5' noncoding region (5' NCR) of hepatitis C virus (HCV) is highly conserved among viral isolates worldwide and that translation of HCV is directed by an internal ribosome entry site (IRES) located within the 5' NCR. We have investigated inhibition of HCV gene expression using antisense oligonucleotides complementary to the 5' NCR, translation initiation codon, and core protein coding sequences. Oligonucleotides were evaluated for activity after treatment of a human hepatocyte cell line expressing the HCV 5' NCR, core protein coding sequences, and the majority of the envelope gene (E1). More than 50 oligonucleotides were evaluated for inhibition of HCV RNA and protein expression. Two oligonucleotides, ISIS 6095, targeted to a stem-loop structure within the 5' NCR known to be important for IRES function, and ISIS 6547, targeted to sequences spanning the AUG used for initiation of HCV polyprotein translation, were found to be the most effective at inhibiting HCV gene expression. ISIS 6095 and 6547 caused concentration-dependent reductions in HCV RNA and protein levels, with 50% inhibitory concentrations of 0.1 to 0.2 microM. Reduction of RNA levels, and subsequently protein levels, by these phosphorothioate oligonucleotides was consistent with RNase H cleavage of RNA at the site of oligonucleotide hybridization. Chemically modified HCV antisense phosphodiester oligonucleotides were designed and evaluated for inhibition of core protein expression to identify oligonucleotides and HCV target sequences that do not require RNase H activity to inhibit expression. A uniformly modified 2'-methoxyethoxy phosphodiester antisense oligonucleotide complementary to the initiator AUG reduced HCV core protein levels as effectively as phosphorothioate oligonucleotide ISIS 6095 but without reducing HCV RNA levels. Results of our studies show that HCV gene expression is reduced by antisense oligonucleotides and demonstrate that it is feasible to design antisense oligonucleotide inhibitors of translation that do not require RNase H activation. The data demonstrate that chemically modified antisense oligonucleotides can be used as tools to identify important regulatory sequences and/or structures important for efficient translation of HCV. 相似文献
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RE O'Carroll A Moffoot KP Ebmeier C Murray GM Goodwin 《Canadian Metallurgical Quarterly》1993,23(2):341-347
Eighteen patients suffering from Alcoholic Korsakoff's Syndrome participated in a placebo-controlled double-blind cross-over trial of clonidine 0.3 mg b.d. for two weeks versus matched placebo for two weeks. A detailed neuropsychological assessment was carried out at the end of each treatment phase and staff ratings of behaviour were also obtained. Clonidine treatment resulted in no significant improvement over placebo on any of the cognitive measures employed. The results contradict previous smaller studies which had suggested that chronic treatment with clonidine had a memory-enhancing effect in Korsakoff's syndrome. 相似文献
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In rhodopsin, the 11-cis-retinal chromophore forms a complex with Lys296 of opsin via a protonated Schiff base. Absorption of light initiates the activation of rhodopsin by cis/trans photoisomerization of retinal. Thermal relaxation through different intermediates leads into the metarhodopsin states which bind and activate transducin (Gt) and rhodopsin kinase (RK). all-trans-Retinal also recombines with opsin independent of light, forming activating species of the receptor. In this study, we examined the mechanism by which all-trans-retinal activates opsin. To exclude other amines except active site Lys296 from formation of Schiff bases, we reductively methylated rhodopsin (PM-rhodopsin), which we then bleached to generate PM-opsin. Using spectroscopic methods and a Gt activation assay, we found that all-trans-retinal interacted with PM-opsin, producing a noncovalent complex that activated Gt. The residual nucleotide exchange in Gt catalyzed by opsin was approximately 1/250 lower relative to that of photoactivated rhodopsin (pH 8.0, 23 degrees C). Addition of equimolar all-trans-retinal led to an occupancy of one-tenth of the putative retinal binding site(s) of opsin and enhanced the Gt activation rate 2-fold. When the concentration of all-trans-retinal was increased to saturation, the Gt activation rate of the opsin/all-trans-retinal complex was approximately 1/33 lower compared to that of photoactivated rhodopsin. We conclude that all-trans-retinal can form a noncovalent complex with opsin that activates Gt by different mechanisms than photolyzed rhodopsin. 相似文献
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The tentative standard impact test procedure, adopted by the Porcelain Enamel Institute, was used to test and compare cylinders coated with several experimental and commercial enamels. The important facts noted were (1) the effect of cubic thermal expansion on the impact resistance, (2) the importance of the ground coat to the resistance of any one cover enamel, and (3) the wide variation between cover enamels in resistance to impact. 相似文献
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KP Fennelly 《Canadian Metallurgical Quarterly》1997,18(1):1-17
The present study investigated the simultaneous occurrence of emergent stimulus-response relations (functional equivalence) and stimulus-stimulus relations (stimulus equivalence). After being pretrained and tested on two symbolic match-to-sample tasks (X1-Y1, X2-Y2), 20 4- and 5-year-old children were trained to emit specified responses to pairs of stimuli (A1-R1, B1-R1, A2-R2, B2-R2) in one setting (original training) and to emit other responses to one member of each pair (A1-R3, A2-R4) in another setting (reassignment training). Of the 18 children who responded correctly on all trained tasks, 15 emitted the novel responses also in the presence of the nonreassigned stimuli (B1-R3, B2-R4). Eleven of these children also matched same-class stimuli with one another (A1-B1, A2-B2, and vice versa). Additional tests with four of these children documented the formation of conditional response-stimulus relations (R3-B1, R4-B2) in all four children, and of conditional response-response relations (R1-R3, R2-R4, and vice versa) in two of them. Children who did not show stimulus control transfer also failed to match same-class stimuli with one another. Present findings, together with those obtained in animal research, suggest that functional equivalence can imply but does not require stimulus equivalence. 相似文献
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Characterization of the epidermal growth factor receptor in pig oviduct and endometrium 总被引:1,自引:0,他引:1
K Wollenhaupt U Tiemann R Einspanier F Schneider W Kanitz KP Brüssow 《Canadian Metallurgical Quarterly》1997,111(2):173-181
Two different plaque variants of Japanese encephalitis virus were selected from a wild-type Taiwanese isolate using Vero cells. One variant was found to exhibit small plaque morphology with retarded virus replication kinetics in Vero cells, and was demonstrated to be resistant to monoclonal antibody (mAb) E3.3 neutralization. The other variant showed large plaque morphology, was sensitive to mAb E3.3 neutralization, and manifested reduced virulence in mice on both intracranial and intraperitoneal inoculations. These two variants propagated in Vero cells retained high levels of infectivity but had relatively low HA titers as compared with the parent strain. The envelope sequences of these two variants showed four amino acid differences at residues E-85 (Glu/Arg), E-306 (Glu/Gly), E-331 (Ser/Arg), and E-387 (Met/Arg). Our results indicated the neutralizing epitope of Japanese encephalitis virus did not overlap with virus virulence determinant. 相似文献
9.
CM Wheeler CA Parmenter WC Hunt TM Becker CE Greer A Hildesheim MM Manos 《Canadian Metallurgical Quarterly》1993,20(5):286-289
OBJECTIVES: To confirm the risk factors for genital human papillomavirus (HPV) infection. GOAL OF THIS STUDY: To investigate risk factors for HPV detection apart from the correlated risk factors for cervical neoplasia. STUDY DESIGN: Cervical human papillomavirus (HPV) DNA was assessed in 357 cytologically normal women attending the University of New Mexico student health center. Cervical swab samples were obtained for HPV DNA detection and typing using a PCR-based DNA amplification system. Possible determinants of cervical HPV were examined including age, ethnicity, history of sexually transmitted disease, oral contraceptive use, smoking, age at first intercourse, lifetime number of sex partners, marital status, and history of pregnancy. RESULTS: A 44.3% overall prevalence of cervical HPV was observed. On univariate analysis, factors associated with increasing HPV prevalence included higher lifetime number of sex partners and single marital status. After adjustment for potential confounding variables, we found that HPV prevalence increased with higher lifetime number of sexual partners. CONCLUSION: These findings, along with those from the companion reports in this issue of the journal, support the sexual route of transmission of the virus. 相似文献
10.
Salinger Walter L.; Ladrow Pamela; Wheeler Catherine 《Canadian Metallurgical Quarterly》2003,117(6):1257
Reeler (rl/rl) and reeler/wild-type (+/rl) mice synthesize Reln at subnormal rates, as do patients with schizophrenia, bipolar disorder, and autism, thereby forming the basis for a Reln hypothesis for vulnerability to these psychopathologies and justifying attention to the behavioral phenotypes of Reln-deficient mice. Tests of gait, emotionality, social aggression, spatial working memory, novel-object detection, fear conditioning, and sensorimotor reflex modulation revealed the behavioral phenotype of rl/rl, but not +/rl, mice to be different from that of wild-type (+/+) mice. These results reveal no effect of Reln gene dosage and provide significant challenges to both the Reln and the neurodevelopmental hypotheses of the etiology of major psychopathologies. (PsycINFO Database Record (c) 2010 APA, all rights reserved) 相似文献