A rapid micromethod for measuring theophylline in serum by reverse-phase high-performance liquid chromatography

We describe an assay system for measuring theophylline in 25 microliters of serum. The procedure involves extraction with a 95:5 mixture of chloroform:isopropanol containing beta-hydroxypropyltheophylline as internal standard, and reverse-phase chromatography on a 4 mm x 30 cm column containing "micron Bondapak C18." Theophylline and beta-hydroxypropyltheophylline are eluted with a 90:10 mixture of sodium acetate butter (20 mmoles/litre pH 4.0) and acetonitrile at a flow rate of 1.8 ml/min., are detected by their absorbance at 254 nm, and quantitated by measuring peak areas. Column temperature has not been found to be critical in this analysis. Each analysis requires 9 minutes of chromatography time with a total analysis time of 20 minutes. Analytical recoveries were found to be 71 to 75% for theophylline and 94% for beta-hydroxypropyltheophylline. This difference in recovery is corrected when determining the theophylline concentration in unknown samples. The method has good precision (coefficients of variation between 7.0% and 7.9% for therapeutic and toxic concentrations). The results obtained with this method compare favourably with results obtained by a published cation-exchange high-performance liquid chromatographic method. None of the metabolites of theophylline, common compounds related to theophylline in structure or drugs tested have been found to interfere with the analysis described.     

Age-related variations in hepatic regeneration and erythropoietin production in the rat

Two hundred and thirty-five entrants into the MRC trial for mild to moderate hypertension were matched with control subjects in order to assess the psychological effects of a screening programme and recruitment into a clinical trial. The prevalence and incidence of psychiatric morbidity among the trial participants were compared with those of the controls by means of responses to a self-administered questionnaire and diagnostic psychiatric interviews. No differences among the groups were shown between screening and entry into the trial, but after entry the prevalence of psychiatric morbidity among the trial participants fell. This was due to a greater improvement of those with psychiatric symptoms at entry in this group, the incidence of new morbidity being similar among the groups.     

Gas chromatographic analysis of acetophenone oxime and its metabolites

A gas-liquid chromatographic (GLC) method has been developed for monitoring the metabolic reduction of acetophenone oxime or oxidative metabolism of the corresponding amine, alpha-methylbenzylamine in liver homogenates. The oxime, amine, n-hydroxy-alpha-methylbenzylamine and acetophenone are quantitatively determined after GLC separation of components with temperature programming on an SP-2401-DB-coated column. The first three compounds were silylated with N,O-bis(trimethylsilyl)-acetamide prior to chromatographic analysis to enhance the stability and improve the chromatographic properties of these components. The effluent gas was monitored with flame ionization detection, and permitted quantitation of components at sub-microgram/ml levels with reproducibility between injections of +/-2%. The optimal composition of enantiomeric mixtures of (R,S)-alpha-methylbenzylamines formed during metabolic reduction of acetophenone oximes were determined by conversion to diastereomeric amides and subsequent GLC analysis.     

Effect of antioxidant in endothelial cells exposed to oxidized low-density lipoproteins

Antioxidants such as probucol and alpha-tocopherol have been shown to attenuate the oxidation of low-density lipoproteins (LDL) and atherosclerotic lesions in animal models of atherosclerosis. The purpose of this study is to determine the protection effect of antioxidants on endothelial cells when exposed to oxidized and native LDL. In a cell-free system, we found that probucol, alpha-tocopherol, and ascorbic acid inhibited copper-induced LDL oxidation by a dose-dependent fashion (from 1 microM to 10 mM). In porcine aortic endothelial cells, antioxidants alone did not change basal endothelin-1 (ET-1) secretion. When porcine aortic endothelial cells were exposed to LDL and oxidized-LDL, both of them stimulated ET-1 secretion dose-dependently, whereas oxidized-LDL elicited higher ET-1 secretion. However, probucol, alpha-tocopherol, and ascorbic acid did not prevent LDL or oxidized-LDL induced ET-1 secretion. Furthermore, nimodipine inhibited both of native and oxidized LDL induced ET-1 secretion. Since Ca2+ channel blocker reduced the elevation of induced ET-1 secretion, the [Ca2+]i is possibly involved for the regulation of ET-1 secretion. Our results suggest that antioxidants can only prevent the oxidation of LDL rather than oxidized and native LDL-induced ET-1 secretion in vascular endothelial cells. The increase in the [Ca2+]i of endothelial cells through the opening of voltage-dependent Ca2+ channels may be involved in the LDL-induced ET-1 release.     

Total anomalous pulmonary venous return complicating portoenterostomy for biliary atresia

Cardiovascular anomalies such as absent inferior vena cava and preduodenal portal vein are reported in cases of biliary atresia and make hepatic portoenterostomy a technical challenge. The authors present the case of a severe cardiac anomaly that significantly altered the functional outcome of a Kasai procedure. Baby M., an 8-week-old boy born with total anomalous pulmonary venous return (TAPVR), underwent hepatic portoenterostomy for biliary atresia. Over the next 3 months he remained icteric and febrile, and failed to gain weight. After multiple antibiotic treatments for suspected cholangitis, he underwent reexploration of the portoenterostomy, with no improvement in his overall condition. His prognosis was considered dismal because correction of the cardiac anomaly is associated with a high mortality rate (> 90%). The cardiac surgeon agreed to attempt a cure of the TAPVR, provided liver transplantation is contemplated if the patient survived. Within 48 hours postoperatively, his hepatic function had improved drastically. He became afebrile, had an improved appetite and weight gain, and was finally discharged 203 days after admission. One year later, he is thriving and remains anicteric. The exact reason for this drastic improvement is not well understood, but the right-sided cardiac failure caused by the TAPVR had a significant effect on the functional outcome of the portoenterostomy.     

Enhanced T-cell immunogenicity and protective efficacy of a human immunodeficiency virus type 1 vaccine regimen consisting of consecutive priming with DNA and boosting with recombinant fowlpox virus

The induction of human immunodeficiency virus (HIV)-specific T-cell responses is widely seen as critical to the development of effective immunity to HIV type 1 (HIV-1). Plasmid DNA and recombinant fowlpox virus (rFPV) vaccines are among the most promising safe HIV-1 vaccine candidates. However, the immunity induced by either vaccine alone may be insufficient to provide durable protection against HIV-1 infection. We evaluated a consecutive immunization strategy involving priming with DNA and boosting with rFPV vaccines encoding common HIV-1 antigens. In mice, this approach induced greater HIV-1-specific immunity than either vector alone and protected mice from challenge with a recombinant vaccinia virus expressing HIV-1 antigens. In macaques, a dramatic boosting effect on DNA vaccine-primed HIV-1-specific helper and cytotoxic T-lymphocyte responses, but a decline in HIV-1 antibody titers, was observed following rFPV immunization. The vaccine regimen protected macaques from an intravenous HIV-1 challenge, with the resistance most likely mediated by T-cell responses. These studies suggest a safe strategy for the enhanced generation of T-cell-mediated protective immunity to HIV-1.     

Chromosomes of Damaliscus (Artiodactyla, Bovidae): simple and complex centric fusion rearrangements

G- and C-banded karyotypes of Damaliscus hunteri, D. lunatus and D. pygargus were compared using the standard karyotype of Bos taurus. Chromosomal complements were 2n = 36 in D. lunatus jimela, 2n = 38 in D. pygargus phillipsi and D. p. pygargus, and 2n = 44 in D. hunteri. The fundamental number in all karyotypes was 60. Among the three species of Damaliscus, seven autosomal pairs and the X chromosomes were conserved. Y-chromosome differences were attributed to heterochromatic additions or deletions. Banded karyotypes of the two subspecies of D. pygargus exhibited complete homology. Chromosomal complements of D. pygargus and D. lunatus differed by a simple centric fusion. However, karyotypes of D. pygargus and D. lunatus differed from D. hunteri by numerous centric fusions, several of which were related by monobrachial chain complexes. Between the karyotypes of D. hunteri and D. pygargus or D. lunatus, there were two chain complexes, one involving five chromosomes (chain V) and the other involving 12 in pygargus (chain XII) or 13 in lunatus (chain XIII). There were also two simple centric fusions between D. hunteri and D. lunatus/D. pygargus; acrocentric chromosomes 13, 15, 20 and 22 in D hunteri were fused as 13;15 and 20;22 in D. lunatus and D. pygargus.