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Biomedical subjects

A M Harper

Publications and source records attributed to A M Harper.

At least 37 records · Page 2Linked to original sources
PubMed1985

Cytochrome b-245 from human neutrophils is a glycoprotein.

Cytochrome b-245 is a glycoprotein. It runs as a broad band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and its apparent Mr varies with the concentration of acrylamide. It stained positively with Schiff reagent and with silver stains after oxidation with periodic acid. It preferentially bound the lectin of Phaseolus vulgaris (type III), and cleavage of carbohydrate with endoglycosidase F resulted in a sharp band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis with an apparent Mr of 55000 G.l.c. analysis of carbohydrate showed this to account for about 15% of the Mr and N-acetylglucosamine and galactose to be the major sugars.

Amino Acids↗
PubMed1985

Effect of pretreatment with the calcium antagonist nimodipine on local cerebral blood flow and histopathology after middle cerebral artery occlusion.

We used the [14C]iodoantipyrine autoradiography technique to study the effect of pretreatment with the calcium antagonist nimodipine on local cerebral blood flow (lCBF) in rats that underwent middle cerebral artery (MCA) occlusion. In untreated control animals there were profound localized reductions in 1CBF 30 minutes after MCA occlusion. These were most pronounced in neocortical areas and in the caudate nucleus ipsilateral to the MCA occlusion. In animals pretreated with nimodipine (1 microgram X kg-1 X min-1 for 30 minutes before and 30 minutes after MCA occlusion), the ipsilateral decrease in 1CBF in cortical regions was significantly less than that in control animals. The drug did not appear to alter 1CBF in the ipsilateral caudate nucleus. Neuropathological quantification of the ischemic damage present 3 hours after occlusion showed that nimodipine pretreatment reduced the volume and extent of cellular damage in the periphery but not in the core of the lesion.

Acid-Base Equilibrium↗
PubMed1985

Effect of the calcium antagonist nimodipine on local cerebral blood flow and metabolic coupling.

The effects of a continuous infusion of the calcium antagonist nimodipine (1 microgram kg-1 min-1) on local CBF (LCBF) and local cerebral glucose utilisation (LCGU) were studied, using the quantitative autoradiographic [14C]iodoantipyrine and [14C]2-deoxyglucose techniques in 34 anatomically discrete regions of the brain in lightly restrained, conscious rats. The infusion of nimodipine at this concentration produced only a small (8%) reduction in the MABP. The administration of nimodipine did not alter the rate of glucose utilisation in any of the regions examined. By contrast, in 24 regions, CBF was increased significantly by 39-84% from control levels (for example, cerebral cortices, hippocampus, hypothalamus, and most thalamic nuclei). In vehicle-treated animals, there was an excellent correlation (p less than 0.01) between the local levels of CBF and glucose utilisation, with the ratio of flow to glucose use being approximately 1.5 ml mumol-1 in each brain region. During nimodipine treatment, there was a similarly excellent correlation (p less than 0.01) between LCBF and LCGU, but the median ratio between local flow and glucose use increased to 2.5 ml mumol-1.

Animals↗
PubMed1985

Serotonin depression of local cerebral glucose utilisation after monoamine oxidase inhibition.

The major catabolic enzyme for serotonin, monoamine oxidase (MAO), is present in the endothelium of cerebral vessels. We report the effects of the intracarotid administration of serotonin on local cerebral glucose utilisation in rats following MAO inhibition with the drug clorgyline. It was found that saline, clorgyline, or serotonin alone produced no significant changes in glucose utilisation. Following the infusion of clorgyline, the administration of serotonin produced significant decreases in glucose utilisation in cortical areas of between 12 and 33% and in the caudate nucleus of 16%.

Animals↗
PubMed1985

The uniformity of liver perfusion as measured by krypton-85 clearance.

The uniformity of regional liver perfusion was studied in eight anaesthetised dogs by measuring the clearance of 85Kr with three beta detectors, and the results were compared with independent measurements of mean perfusion by electromagnetic flowmetry. The intra-hepatic flow distribution was found to be heterogeneous, with a coefficient of variation between locations which averaged 17%. The pattern of flow varied from liver to liver, and there was no consistent tendency for any one of a limited number of locations to be better or less well perfused than any other. Discrepancies between 85Kr clearance and flowmeter measurements were of a similar magnitude to inter-regional variations in flow, and there was no evidence of a systematic difference between the two measurement techniques. Individual 85Kr clearance measurements can misrepresent the perfusion of the liver as a whole, and averaging several measurements performed in different locations is likely to yield a better estimate of mean flow.

Animals↗
PubMed1984

Pyrolysis mass spectrometry of complex organic materials.

Pyrolysis mass spectrometry in combination with computerized multivariate statistical analysis enables qualitative and quantitative analysis of nonvolatile organic materials containing molecular assemblies of a complexity and size far beyond the capabilities of direct mass spectrometry. The state of the art in pyrolysis mass spectrometry techniques is illustrated through specific applications, including structural determination and quality control of synthetic polymers, quantitative analysis of polymer mixtures, classification and structural characterization of fossil organic matter, and nonsupervised numerical extraction of component patterns from complex biological samples.

Biochemical Phenomena↗
PubMed1984

Purification of cytochrome b-245 from human neutrophils.

The low potential cytochrome b (b-245) of the microbicidal oxidase of phagocytic cells has been purified from neutrophils from patients with chronic myeloid leukaemia. Cells were homogenized in the presence of proteinase inhibitors and centrifuged to remove the cytoplasm. The pellets containing membranes, granules and other organelles (15 mg/ml) were then washed with buffered sodium cholate (5 mg/ml). Residual pellets were subsequently solubilized with the non-ionic detergent Triton N 101 (10 mg/ml) which extracted about 60% of the cytochrome b. About 10% of the cytochrome b was of mitochondrial origin which was removed on a column of n-amino-octyl-Sepharose that did not adsorb cytochrome b-245. Cytochrome b-245 was chromatographed on a column of heparin-agarose and eluted with NaCl to give a peak specific content of 11-16 nmol of cytochrome b-245/mg of protein, representing a 140-200-fold purification with a recovery of 15%. This technique results in the purification of approx. 100-150 nmol of highly purified cytochrome b-245 from (3-5) X 10(11) cells within 4 days. The most purified material gave a broad band with an apparent Mr of between 68 000 and 78 000 on sodium dodecyl sulphate/polyacrylamide gel electrophoresis, but gel filtration indicated an aggregated form of the protein in Triton N101 . Purified protein (14 nmol of haem/mg of protein) did not contain FAD or FMN and had no NADPH-dependent O2--generating activity.

Chromatography, Affinity↗
PubMed1984

Effect of the calcium antagonist nimodipine on local cerebral blood flow: relationship to arterial blood pressure.

The effect of the calcium antagonist nimodipine on local CBF in 31 regions of the CNS was studied with the [14C]iodoantipyrine autoradiographic technique in lightly anaesthetised, mechanically ventilated rats. Continuous intravenous infusion of nimodipine (1, 2, or 4 micrograms kg-1 min-1) produced a dose-dependent reduction in MABP (reduced by 26 +/- 2% after 30 min of nimodipine, 4 micrograms kg-1 min-1) and a significant elevation in plasma glucose concentration (increased by 44 +/- 2% after 30 min of nimodipine, 4 micrograms kg-1 min-1). Local CBF was increased significantly during infusions of nimodipine (1 microgram kg-1 min-1) in 9 of the 31 regions examined (including the cerebral cortex, hippocampus, hypothalamus, and most thalamic nuclei). In contrast to the increases in CBF observed in forebrain regions, no significant increases in CBF were observed during nimodipine infusions in regions of the lower brainstem, cerebellum, and pons or in myelinated fibre tracts. The proportionately greatest increases in local CBF were observed during infusions of the lowest dosage of nimodipine (1 microgram kg-1 min-1), suggesting either that this dosage provokes maximum cerebrovascular relaxation or that effects of increasing concentrations are counteracted by the concomitant systemic hypotension.

Animals↗
PubMed1983

Iodination by stimulated human neutrophils. Studies on its stoichiometry, subcellular localization and relevance to microbial killing.

Myeloperoxidase of phagocytic leucocytes is thought to utilize H2O2 to oxidize halides, which then react with and kill ingested microbes. This hypothesis was based largely on the incorporation of radiolabelled iodide into cells that had phagocytosed bacteria. The present studies investigated the stoichiometry of these reactions and the subcellular localization and electrophoretic pattern of the cellular components that became iodinated. 1. The stoichiometry of the reactions are such that only a small proportion (less than 0.3%) of the total oxygen consumed is utilized for iodination. Iodination after stimulation with the soluble stimulus phorbol myristate acetate (PMA), which is not known to involve the azurophil granules and their contained myeloperoxidase, was comparable with that occurring after bacterial ingestion. 2. Analytical subcellular fractionation of cells that had phagocytosed bacteria localized about 25% of the radioactivity to the membranes, and most of the residual radioactivity distributed with the bacteria and dense granules. In cells stimulated with PMA, more of the radioactivity was associated with the membranes, but about half was still associated with the dense granules. 3. Autoradiographs after dodecyl sulphate/polyacrylamide-gel electrophoresis of cells stimulated with opsonized bacteria gave a similar distribution of iodinated components to that obtained with cells that had been stimulated with PMA or iodinated with Iodogen. These patterns of iodination were very different from those obtained when bacteria alone were iodinated with Iodogen or myeloperoxidase and H2O2. Preparations in which bacteria had been phagocytosed did not show evidence of iodination of bacterial proteins or coating opsonins. Thus positive evidence for the iodination of bacteria has not been produced, and the role of iodination in the microbicidal process of neutrophils remains to be established.

Autoradiography↗
PubMed1983

The effects of serotonin on local cerebral blood flow.

This study was undertaken to measure the effects of serotonin administration on local cerebral blood flow following blood-brain barrier (BBB) disruption with hypertonic urea. Rats were anesthetized with halothane in nitrous oxide and oxygen (70%:30%). In some animals urea (3.5 M) was infused retrogradely through an external carotid catheter, followed after 10 min by serotonin (50 ng kg-1 min-1) or physiological saline. Local cerebral blood flow was measured using the 14C-iodoantipyrine quantitative autoradiographic technique of Sakurada et al. (1978). The administration of saline or urea alone had only minimal effects on local cerebral blood flow. When the BBB was intact, serotonin produced a significant fall in regional blood flow only in the caudate nucleus. Following BBB disruption, however, serotonin produced a marked decrease in local perfusion in a number of discrete brain areas that are supplied by blood from the internal carotid artery. On the other hand, there were increases in local perfusion in areas not supplied by the internal carotid artery.

Animals↗
PubMed1983

Antigen shared by human hemopoietic precursor cells and T and B lineage cells.

In this study, we investigated antigens present at the surface of acute myeloblastic leukemia (AML) cells by using the monoclonal antibody (MAb) approach. The MAb AGF43 reacted with acute myeloid and lymphoid leukemia cells and chronic lymphocytic leukemia cells and was unreactive against chronic myeloid leukemia cells. A large proportion of AML blasts showing minimal or no differentiation (AML-M1) were intensely labeled by AGF43 in contrast to a smaller percentage of blasts showing partial differentiation (AML-M2 and acute myelomonocytic leukemia). The AGF43 antigen is expressed by bone marrow lymphoid (TdT+) and myeloid (CFU-GM) progenitor cells, 95% of B cells and 65% of T cells in the blood and absent from monocytes. Only 17% of normal myeloblasts were weakly stained by AGF43. Sections of tonsil and spleen were used to confirm that, unlike antibodies to MHC class II antigens, AGF43 stained a majority of T cells and macrophages were unreactive. In conclusion, the MAb AGF43 identifies a new precursor cell antigen. The distribution of this antigen during normal myelopoiesis and on AML cells support the suggestion that acute myeloid leukemias originate in pluripotent or closely related myeloid stem cells.

Antibodies, Monoclonal↗
PubMed1983

Interaction of histamine with noradrenergic constrictory mechanisms in cat cerebral arteries and veins.

We examined responses of pial arteries and veins in situ to noradrenergic stimuli in the presence of histamine. Electrical stimulation of sympathetic nerves and perivascular microapplication of norepinephrine in mock cerebrospinal fluid produced constriction of arteries and veins in anesthetized cats. During simultaneous perivascular injection of histamine, these noradrenergic responses were attenuated or reversed. In both arteries and veins, constriction from sympathetic nerve stimulation was prevented by simultaneous application of the histamine receptor agonists, pyridylethylamine (H1) or impromidine (H2), results that suggest interference involving both types of histamine receptors. In arteries, impromidine, but not pyridylethylamine, inhibited constriction resulting from exogenous norepinephrine. Our findings indicate that histamine may have an inhibitory influence, exerted through both receptor types, on noradrenergic mechanisms in cerebral vessels.

Animals↗
PubMed1982

The enzymic reduction and kinetics of oxidation of cytochrome b-245 of neutrophils.

1. The absorption coefficient of human neutrophil plasma-membrane reduced-minus-oxidized cytochrome b-245 was determined [delta epsilon (mM; 559-540 nm) = 21.6 cm-1]. 2. Neutrophil polymorphonuclear leucocytes (neutrophils) were prepared from human, ox, horse and pig blood. In each case plasma-membrane fractions were found to contain low-potential cytochrome b. When membranes from horse neutrophils were incubated anaerobically with either NADH or NADPH the cytochrome b became reduced. Prior stimulation of the cells with phorbol myristate acetate did not increase the rate or extent of cytochrome b reduction in isolated membranes, but did increase both the rate and extent of reduction by NADPH in Triton-treated cells. 3. A cytochrome b was present also in the specific granule fraction of human neutrophils. Its Em (pH 7.0) was found to be -248 mV, very similar to that of the plasma-membrane cytochrome b. 4. The rate of oxidation of reduce cytochrome b-245 by air-saturated buffer, was determined by using stopped-flow techniques. In intact membranes t 1/2 for oxidation was 4.7 ms. This rate is sufficiently rapid to support the view that cytochrome b-245 is the oxidase in the respiratory burst of neutrophils. 5. Plasma-membrane cytochrome b of human neutrophils formed a complex with CO. At room temperature and 1 atm of CO approx. 40% of the cytochrome formed a complex; approx. 60% binding was measured at the increased concentration of dissolved CO achieved at 5 degrees C. The concentration of CO giving 50% binding was 1.18 mM.

Animals↗
PubMed1982

Regional effect of mepyramine on cerebral glucose utilization in conscious rats.

We examined effects of the antihistamine, mepyramine, which occupies cerebral H1-receptors, on regional brain metabolism of glucose in conscious rats. Although glucose utilization by most structures was not significantly affected, in high doses, mepyramine did cause modest, localized reductions in brain areas with both high (ventromedial nucleus of hypothalamus) and low (medial forebrain bundle) densities of histamine H1-receptors.

Aminopyridines↗
PubMed1982

The action of cells from patients with chronic granulomatous disease on Staphylococcus aureus.

Contrary to current opinion, neutrophil leucocytes from patients with the inherited immunodeficiency syndrome chronic granulomatous disease (CGD) killed 80% of ingested Staphylococcus aureus. Bacterial killing was not impaired by increasing the ratio of bacteria to cells from 1:1 to 10:1. The organisms that survived within the patients' cells did not themselves appear to constitute an unduly resistant subpopulation because they were killed when exposed to fresh cells, and no growth phase of a synchronous culture was found to be particularly resistant. The pH within the phagocytic vacuoles of CGD neutrophils and monocytes is abnormally low and methylamine, which has been shown to normalise this vacuolar pH, improved killing. Clumped bacteria appeared to be more resistant to killing than dispersed ones, suggesting that organisms near the centre of a clump might be protected from the toxicity of the compromised killing systems in cells of these patients.

Granulomatous Disease, Chronic↗
PubMed1982

Intra-arterial histamine increases blood-brain transport in rats.

The neurotransmitter histamine is located in multiple compartments in the brain and may influence cerebral vessels during some conditions. We made measurements of cerebrovascular transport of labeled sucrose, alpha-aminoisobutyric acid (AIB), and horseradish peroxidase during sustained infusion of histamine into the internal carotid artery of anesthetized rats. Histamine increased the rates of transport of sucrose and AIB up to 237% in several brain regions and in different areas of cerebral cortex. The effect was dose dependent, was reversible within 2 h, and was mediated primarily by histamine H2-receptors. An increase in cortical water content occurred, and electron microscopic examination revealed that perivascular astroglial processes were swollen. There was a twofold increase in the number of endothelial pits and vesicles in cortical capillaries perfused by histamine. We conclude that histamine can influence the function of the blood-brain barrier.

Animals↗
PubMed1981

Impromidine, a potent dilator of cat pial arterioles.

Impromidine (SKF 92676) has been demonstrated to have highly potent and selective activity at histamine H2-receptors in a variety of physiological systems. We compared responses produced by perivascular microapplication of histamine and impromidine in cat pial arterioles in situ. Impromidine caused twice as much maximal dilation as histamine, and was ten times more potent. These observations emphasize the pronounced effects that can result from stimulation of histamine H2-receptors in brain arterioles.

Animals↗