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Biomedical subjects

J Oxford

Publications and source records attributed to J Oxford.

At least 19 recordsLinked to original sources

Construction and characterization of a full-length HIV-1(92UG001) subtype D infectious molecular clone.

Here we report the construction, sequencing, and biological characterization of a molecular clone of HIV-1(92UG001), a virus representative of subtype D strains circulating in Uganda. The virus produced by the clone has an aggressive syncytium-inducing phenotype, which matches that of the parental virus. This phenotype may be related to duplication of a binding site for a transcription factor, T cell factor 1alpha (TCF-1alpha), in the long terminal repeat of the virus.

Adult↗

Influenza virus carrying neuraminidase with reduced sensitivity to oseltamivir carboxylate has altered properties in vitro and is compromised for infectivity and replicative ability in vivo.

Oseltamivir phosphate (Tamiflu, Ro 64-0796) is the first orally administered neuraminidase (NA) inhibitor approved for use in treatment and prevention of influenza virus infection in man. Oseltamivir phosphate is the pro-drug of the active metabolite oseltamivir carboxylate (Ro 64-0802). Extensive monitoring throughout the oseltamivir development programme has identified a very low incidence of patients who have carried drug-resistant virus. The predominant mutation seen is the substitution of arginine for lysine at position 292 of the viral NA. The fitness of clinically isolated influenza virus A/Sydney/5/97 (H3N2) carrying this mutation was markedly reduced in animal models of influenza virus infection. The infectivity and replicative abilities of R292K mutant virus were reduced by at least 2 logs in a mouse model of influenza infection and by 2 and 4 logs, respectively, in the ferret model. Pathogenicity of R292K influenza virus A/Sydney/5/97 was reduced in ferrets as measured by inflammatory and febrile responses at least in parallel to the decrease in replicative ability. The data indicate that the R292K NA mutation compromises viral fitness such that virus carrying this mutation is unlikely to be of significant clinical consequence in man.

Acetamides↗

Antiretroviral therapy for HIV-2 infected patients.

OBJECTIVES: To evaluate clinical and RNA load response to antiretroviral therapy amongst patients infected with HIV-2 and to study the development of drug resistance. METHODS: Seven HIV-2 seropositive patients were monitored with clinical examination, CD4 cell count and HIV-2 viral RNA load. Viruses from four subjects were genotyped and in vitro recovery of virus by co-cultivation with PBMCs and HVS T-cells was attempted. Viruses isolated from two subjects were assayed for phenotypic antiviral resistance. The main outcome measures were the relationship between disease stage, viral load, CD4 cell count, viral subtype and the clinical course of HIV-2 infection and the effect of combination antiretroviral therapy on disease progression, CD4 cell count, HIV-2 RNA viral load and drug resistance. RESULTS: The median time of follow-up was 3 years (range 0-8 years). Three patients had AIDS, and one had symptomatic disease. Of the four patients genotyped, three were infected with HIV-2 subtype B and one with subtype A. Viraemia was detectable only at CD4 counts of less than 300 x 10(6)/ml. Two patients with high viral loads failed to respond to antiretroviral therapy although their treatment may not have been optimal. One developed in vitro phenotypic antiviral resistance. The genotype of this patient's viral reverse transcriptase is being analysed. CONCLUSIONS: In contrast to HIV-1, HIV-2 RNA levels were often undetectable despite advanced disease and low CD4 cell counts. However, HIV-2 was clearly capable of causing CD4 cell depletion resulting in symptomatic disease. The principles of highly active antiretroviral therapy seem to apply to HIV-2 and suboptimal therapy may lead to drug resistance. The timing of therapy initiation, monitoring of response and the measurement of resistance remain unresolved issues and conclusions cannot be extrapolated from HIV-1.

Acquired Immunodeficiency Syndrome↗

Construction and biological characterization of an infectious molecular clone of HIV type 1GB8.

Here we report the construction, sequencing, and repair of a molecular clone of HIV-1GB8, a virus representative of HIV-1 subtype B strains circulating in the UK. The phenotype of virus produced by the clone matches that of the parental virus. The molecular clone will be used in the production of attenuated virus stocks for chemical inactivation to allow development of faccines based on killed whole virus preparations.

Base Sequence↗

Electrophoretic analysis of nucleic acids isolated from scrapie-infected hamster brain.

The purpose of this study was to investigate previous reports of a scrapie-specific 1.2 kb single-stranded DNA observed in alkaline agarose electrophoresis gels. Protocols were developed to be as consistent as possible with those used previously. Partial subcellular fractionation was applied to the brains of hamsters clinically affected by the 263K strain of scrapie. Nucleic acids were then isolated, and compared electrophoretically to nucleic acids isolated from equivalent fractions, made from the brains of hamsters inoculated with normal brain. Several modifications to the protocols were suggested. Results obtained by using these modifications were compared to results obtained using the original protocols. Scrapie-specific DNA was not observed in a total of eleven consecutive experiments. Thus, these results do not support the hypothesis that the agents of transmissible spongiform encephalopathy contain a single-stranded 1.2 kb DNA species as previously described.

Animals↗

Does prenatal exposure to influenza in mice induce pyramidal cell disarray in the dorsal hippocampus?

Epidemiological studies point to an association between prenatal exposure to influenza and later schizophrenia. Such studies are consistent with neuropathologic reports demonstrating cytoarchitectural abnormalities in the hippocampus and parahippocampal gyrus suggestive of second trimester developmental anomalies. The hypothesis that prenatal exposure to influenza in the second trimester may induce hippocampal pyramidal cell disarray in mice was investigated. Between days 9-16 of pregnancy, 35 Balb/c mice were intranasally inoculated with either a mouse-adapted or non mouse-adapted pool of Influenza A/Singapore/1/57 (H2N2), and 10 controls were inoculated with normal saline. Offspring were sacrificed on day 21 postpartum. Microscopic examination of the CA1-CA2 junctional areas in the offspring of mice exposed to influenza failed to demonstrate excess pyramidal cell disarray when compared with influenza-free, age matched controls. There was evidence that disarray was greater among those exposed on day 13 of pregnancy. Analyses of the data by sex and severity of maternal infection failed to reveal any significant effects.

Animals↗

Characterization of glucuronic acid conjugates of a novel angiotensin receptor antagonist.

Nanogram quantities of glucuronic acid conjugates of GR117289 in rat and dog bile have been analysed by semi-microbore high-performance liquid chromatography (HPLC)/ionspray mass spectrometry with on-line UV diode array detection. The determination of drug metabolites in bile has often proved problematical due to the large number of endogenous components in this biological matrix, in particular the bile acids. Semi-microbore HPLC is useful for concentrating small quantities of material and, in combination with an on-line diode array detector, for distinguishing between drug related and endogenous components. A novel angiotensin II receptor antagonist, GR117289, had proved difficult to analyse by thermospray mass spectrometry because of its thermal lability. The use of the less thermally dependent technique of ionspray mass spectrometry allowed the characterization of nanogram quantities of glucuronic acid metabolites of GR117289 in bile.

Angiotensin Receptor Antagonists↗

Thermospray liquid chromatography-mass spectrometry for the characterisation of sulphate ester conjugates.

Formation of polar conjugates is a well documented metabolic pathway for xenobiotics containing phenolic hydroxyl groups. This paper describes the analysis of two sulphate ester conjugates by fast atom bombardment mass spectrometry and thermospray liquid chromatography-mass spectrometry. Thermospray liquid chromatography-mass spectrometry proved the more successful technique for obtaining the molecular weight of the intact conjugate, but only by removal of the buffer from the high-performance liquid chromatography eluent.

Albuterol↗

Failure of influenza vaccine to prevent two successive outbreaks of influenza A H1N1 in a school community.

Forty nine of the 149 boys (33%) at a preparatory school fell ill at the beginning of the autumn term 1986 with symptoms of influenza. One hundred and eighty two of the 470 pupils (39%) in the senior part of the same school had similar symptoms of influenza at the beginning of the spring term 1987. A new variant of influenza A H1N1 virus was isolated from both outbreaks and shown to be antigenically similar to A/Taiwan/1/86. The attack rate among pupils who had previously received trivalent influenza vaccine containing A/Chile/1/83 H1N1 antigen was not significantly different from the rate among those who had never been vaccinated. It is concluded that annual vaccination of all boarding school pupils may be inappropriate.

Disease Outbreaks↗

Development and validation of a liquid chromatographic-mass spectrometric assay for the determination of sumatriptan in plasma.

Sumatriptan succinate is a novel compound currently in development for the acute treatment of migraine. During early studies in man a sensitive and selective assay was required, which had to be developed rapidly, to determine plasma concentrations following an intravenous infusion. Thermospray liquid chromatography-mass spectrometry combined with the advanced automated sample processor was selected to achieve this. Although the assay was required quickly criteria for intra- and inter-assay accuracies and precisions of +/- 10% had to be achieved. These were obtained only by using a co-eluting deuterium-labelled internal standard. Attempts to use a homologue as an internal standard, which did not co-elute with sumatriptan, gave inferior results. The assay was linear over the calibration range 2-50 ng/ml with a limit of quantification of 2 ng/ml. The application of the technique to the analysis of samples from a volunteer study is demonstrated.

Chemical Phenomena↗

Automated sample preparation on-line with thermospray high-performance liquid chromatography-mass spectrometry for the determination of drugs in plasma.

The combination of a solid-phase extraction module, the AASP, on-line with thermospray high-performance liquid chromatography-mass spectrometry for the automated determination of drugs in plasma is described. The technique was evaluated successfully using, as an example, the determination of labetalol in human plasma. [2H7]Labetalol was used as an internal standard to compensate for changes in ionization efficiencies between analyses. The chromatographic and mass spectrometric conditions were optimized for labetalol. The combined technique was demonstrated as being robust and reliable for the analysis of plasma samples from a clinical study.

Autoanalysis↗

Qualitative and quantitative analysis of ranitidine and its metabolites by high-performance liquid chromatography-mass spectrometry.

Reversed-phase high-performance liquid chromatography systems for the separation of ranitidine and its metabolites ranitidine-N-oxide, ranitidine-S-oxide, and desmethylranitidine have been developed for use in high-performance liquid chromatography-mass spectrometry. A direct liquid introduction-high-performance liquid chromatography-mass spectrometry system to analyse qualitatively and quantitatively solutions containing ranitidine and its metabolites by reversed-phase chromatography is described. A sample of urine collected from a subject given an oral dose of 75 mg of ranitidine and 75 mg of tris-deuterated ranitidine was analysed by this system. Ranitidine and its metabolites were identified by the ion doublets in the mass spectra which were 3 a.m.u. apart.

Biotransformation↗

Automated extraction of drugs from biological fluids.

An automated continuous flow liquid-liquid extraction procedure is described for the separation of the H2-antagonist loxtidine from plasma samples containing two metabolites which interfere in the radioimmunoassay of the drug. The extraction of the bronchodilator salbutamol was studied using the DuPont Prep I automated liquid solid extraction apparatus, with a 12 cartridge capacity, and a vacuum extraction box designed in this laboratory to hold 30 Sep-pak C-18 (Waters Associates) cartridges. Twenty-four plasma samples per hour can be automatically processed with the Prep I. Although the vacuum box is not fully automated 45 plasma samples per hour can be processed. The Prep I can only be used with DuPont XAD, strong cation and anion exchange cartridges. Cartridges containing alumina, silica, florisil, cation and anion exchange resins and reverse phase packings can all be used with the vacuum extraction box. The latter costs only a fraction of the Prep I and therefore each analyst can have his own unit.

Acetates↗

Modulation of human natural killer cytotoxicity by influenza virus and its subunit protein.

The influence of intact influenza virus and purified detergent solubilized haemagglutinin (HA) subunits from these viruses on human natural killer (NK) cell activity was examined. Effector cells incubated with whole influenza virus for 18 hr initiated the production of alpha interferon which was associated with the enhancement of NK cell activity. In contrast, purified influenza virus HA suppressed NK activity in a dose-dependent manner, when added at the onset of the cytotoxicity assay, or when used to pre-treated effector cells prior to assay for cytotoxicity against K562 target cells. Effector cells exposed to influenza HA for 90 min, washed and re-incubated in fresh medium for up to 18 hr, failed to regain their cytotoxicity. Suppression of NK cell cytotoxicity could not be ascribed to direct toxicity of HA preparations or residual detergent and preservative in these preparations. The augmented cytotoxicity of activated human effector cells was also susceptible to suppression by virus HA, and pretreatment of human PBL effector cells with HA for 90 min, prior to exposure to human alpha interferon caused NK effector cells to become refractive to the enhancing effects of HIFN. That direct interaction between influenza virus HA and effector cells was a requirement for suppression of activity was shown in experiments using Bromelain-released influenza HA, which would not be expected to bind to cells and which failed to suppress NK cell activity.

Bromelains↗