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Biomedical subjects

D A Johnson

Publications and source records attributed to D A Johnson.

At least 19 recordsLinked to original sources
PubMed1992

A deuterium NMR and steady-state fluorescence anisotropy study of the effects of cholesterol on the lipid membrane-disordering actions of ethanol.

We examined the effects of cholesterol on the membrane-disordering action of ethanol by using deuterium nuclear magnetic resonance (2H-NMR) and fluorescence spectroscopy. Specifically, the effects of ethanol were measured on the 2H-NMR spectra of di(perdeuteropalmitoyl)phosphatidylcholine (DPPC-d62) and on the steady-state emission anisotropy of diphenylhexatriene (DPH) incorporated into hydrated egg phosphatidylcholine (eggPC)/cholesterol dispersions. Analysis of the 2H-NMR spectra of DPPC-d62 incorporated into eggPC liposomes showed that the addition of cholesterol up to 30 mol% enhanced the ability of ethanol to disorder methylene groups all along the phospholipid acyl chains. This effect was somewhat greater toward the terminal methyl groups. However, above 30 mol% cholesterol, the bilayer-disordering action of ethanol on both the upper and lower portions of the acyl chains decreased to an apparent constant change up to the highest cholesterol content examined (50 mol%). Analysis of the fluorescence anisotropy of DPH, on the other hand, suggested that cholesterol attenuated the ability of ethanol to disorder the bilayers, which is in agreement with a previous EPR study [Chin and Goldstein, Mol Pharmacol 19: 425-431, 1981]. Re-analysis of our previous fluorescence anisotropy results with DPH incorporated into dispersions of brain-lipid extracts as a percent change [Johnson et al., Mol Pharmacol 15: 739-746, 1979] indicated that the chemical composition of the lipid bilayers also affects the apparent ability of cholesterol to modulate the membrane-disordering action of ethanol, because the addition of cholesterol to brain-lipid extracts had no significant effect on the membrane-disordering action of ethanol. Given the greater likelihood that the 2H-NMR probes accurately monitor bulk phospholipid properties, some caution is required in the analysis of the membrane-disordering actions of drugs using EPR and fluorescence spectroscopy.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1992

Medical therapy for gastroesophageal reflux disease.

Gastroesophageal reflux disease (GERD) remains a ubiquitous problem, although therapeutic options continue to evolve. Effective therapy calls for understanding the pathogenesis. Key factors associated with GERD include incompetence of the lower esophageal sphincter, esophageal clearance, gastric contents, tissue resistance, and potency of the refluxate. Phase-type directed therapy remains the best treatment approach and histamine (H2)-receptor antagonists are now the cornerstone of therapy for patients not responsive to conservative measures. In a subset of patients with severe esophagitis who do not respond to conventional H2-receptor antagonist therapy, efficacy has been demonstrated with high-dose therapy. The acid suppressant omeprazole, highly effective in erosive esophagitis, is the drug of choice for esophagitis resistant to H2-receptor antagonists. Despite effective forms of therapy, relapse rates are high in patients with severe GERD, and maintenance therapy typically is required. With near uniformity, efficacy end points for these agents have been directed toward relief of heartburn, regurgitation, and dyspepsia. Few data exist correlating relief of GERD and improvement of chest pain. Although therapeutic strategies for treating GERD have improved, empiric treatment of suspected GERD in the patient with noncardiac chest pain does not appear to be the optimal approach and should be reserved for cases where diagnostic testing is limited or unavailable.

Chest Pain↗
PubMed1992

Molecular cloning and characterization of mouse mast cell chymases.

Mouse mast cell chymases are granule-associated serine proteinases with chymotrypsin-like substrate specificities. cDNAs for two new chymases were isolated from a cDNA library constructed using mRNA from ABFTL-6 mouse mast cells by screening with a rat mast cell proteinase cDNA. The deduced amino acid sequence of mouse chymase 1 consists of a 226 amino acid catalytic portion and a 21 amino acid preprosequence. Chymase 1 is unusual in that an Asn occurs in the substrate binding pocket, a feature that has not been observed in any other serine proteinase. Also, chymase 1 is expected to have a large positive charge (+13) at physiological pH. A partial cDNA for chymase 2 encodes 177 residues of the carboxy terminal portion of a second proteinase distinct from chymase 1. Chymase 2 cDNA contains a highly conserved intron/exon junction, a high positive charge (+17) and a novel, second potential N-glycosylation site. Transcripts for both chymases are found in ABFTL-6 mast cells, but only chymase 2 mRNA is in mouse connective tissue mast cells. These data suggest that these chymases have distinct enzymatic properties and tissue-specific patterns of gene expression.

Amino Acid Sequence↗
PubMed1992

Quinacrine binds to the lipid-protein interface of the Torpedo acetylcholine receptor: a fluorescence study.

It has been argued both that there is a high affinity noncompetitive inhibitor binding site in the lumen of the acetylcholine receptor and that this lumen exists on the central axis of the receptor. Such a site would be expected to be 20-40 A from the membrane lipids. We tested whether, in fact, quinacrine, a potent fluorescent noncompetitive inhibitor, binds to such a site. We measured quenching of receptor-bound quinacrine fluorescence by fluorescence dipolar energy transfer to lipid probes, 5-(N-dodecanoylamino)eosin and N-(3-sulfopropyl)-4-(p-didecylaminostyryl)pyridinium, or by collision with paramagnetic lipid probes 2,2,6,6-tetramethylpiperidine-1-oxyl and 3-doxyl-17 beta-hydroxy-5 alpha-androstane (spin-labeled androstane). Initial control experiments established that in the presence of carbamylcholine, quinacrine binds to a phencyclidine-sensitive site on the Torpedo receptor with a Kd equal to 0.14 microM and with a quantum yield of 0.18. Fluorescence energy transfer from receptor-bound quinacrine had a magnitude consistent with quinacrine being less than 10 A from the lipid fluorescent probes. 2,2,6,6-Tetramethylpiperidine-1-oxyl and spin-labeled androstane were two to five times more effective at quenching receptor-bound quinacrine fluorescence than the fluorescence from membrane-partitioned 5-(dodecanoylamino)fluorescein. These results suggest that the quinacrine binding site is too close to the lipid domain to be in the lumen of the receptor, and therefore it is probably located on the outer surface of the membrane-spanning domain of the acetylcholine receptor.

Animals↗
PubMed1992

Mast cell tryptases: examination of unusual characteristics by multiple sequence alignment and molecular modeling.

Tryptases are trypsin-like serine proteinases found in the granules of mast cells. Although they show 40% sequence identity with trypsin and contain only 20 or 21 additional residues, tryptases display several unusual features. Unlike trypsin, the tryptases only make limited cleavages in a few proteins and are not inhibited by natural trypsin inhibitors, they form tetramers, bind heparin, and their activity on synthetic substrates is progressively inhibited as the concentration of salt increases above 0.2 M. Unique sequence features of seven tryptases were identified by comparison to other serine proteinases. The three-dimensional structures of the tryptases were then predicted by molecular modeling based on the crystal structure of bovine trypsin. The models show two large insertions to lie on either side of the active-site cleft, suggesting an explanation for the limited activity of tryptases on protein substrates and the lack of inhibition by natural inhibitors. A group of conserved Trp residues and a unique proline-rich region make two surface hydrophobic patches that may account for the formation of tetramers and/or inhibition with increasing salt. Although they contain no consensus heparin-binding sequence, the tryptases have 10-13 more His residues than trypsin, and these are positioned on the surface of the model. In addition, clustering of Arg and Lys residues may also contribute to heparin binding. Putative Asn-linked glycosylation sites are found on the opposite side of the model from the active site. The model provides structural explanations for some to the unusual characteristics of the tryptases and a rational basis for future experiments, such as site-directed mutagenesis.

Amino Acid Sequence↗
PubMed1992

Correlation between basal acid output and daily ranitidine dose required for therapy in Barrett's esophagus.

We prospectively evaluated basal gastric acid secretion in 42 consecutive patients with Barrett's esophagus to determine the optimal dose requirement for an H2-receptor antagonist in relation to the gastric acid secretory status of each patient. All patients with Barrett's esophagus had pyrosis and 31 of the 42 patients had erosive esophagitis. Mean extension of Barrett's epithelium was 6.9 cm (range 2-17 cm). Mean basal acid output for the patients with Barrett's esophagus was 8.0 +/- 5.2 meq/hr, which was significantly different compared to a group of 65 normal subjects with mean basal acid output of 3.0 +/- 2.7 meq/hr (P less than 0.001). There was no correlation between basal acid output and extension of Barrett's epithelium. All patients with Barrett's esophagus were treated with ranitidine, with 24 requiring standard-dose (300 mg/day) and 18 requiring increased doses (mean 1170 mg/day, range 600-2400 mg/day) for complete healing of esophagitis and disappearance of pyrosis. There was a significant correlation between basal acid output and daily ranitidine dose required for therapy (r = 0.52, P less than 0.001). Fifteen of the 42 patients with Barrett's esophagus (36%) had gastric acid hypersecretion. There was a significant association between gastric acid hypersecretion defined as a basal acid output of greater than 10.0 meq/hr and a requirement for increased daily ranitidine doses (greater than 300 mg/day) (P less than 0.0002). No side effects occurred with any of these high doses of ranitidine. We conclude that as a group, patients with Barrett's esophagus have significantly higher basal acid outputs than normal subjects and many require increased therapeutic doses of ranitidine.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1992

Caffeine potentiates the enhancement by choline of striatal acetylcholine release.

We investigated the effect of peripherally administered caffeine (50 mg/kg), choline (30, 60, or 120 mg/kg) or combinations of both drugs on the spontaneous release of acetylcholine (ACh) from the corpus striatum of anesthetized rats using in vivo microdialysis. Caffeine alone or choline in the 30 or 60 mg/kg dose failed to increase ACh in microdialysis samples; the 120 mg/kg choline dose significantly enhanced ACh during the 80 min following drug administration. Coadministration of caffeine with choline significantly increased ACh release after each of the choline doses tested. Peak microdialysate levels with the 120 mg/kg dose were increased 112% when caffeine was additionally administered, as compared with 54% without caffeine. These results indicate that choline administration can enhance spontaneous ACh release from neurons, and that caffeine, a drug known to block adenosine receptors on these neurons, can amplify the choline effect.

Acetylcholine↗
PubMed1992

Mohs histographic surgery and ophthalmic plastic reconstruction.

A retrospective analysis of 43 consecutive cases of primary and recurrent basal cell carcinomas is presented. Each patient's tumor was first managed by Mohs histographic surgery followed by opthalmic plastic reconstruction within 24 h. A review of the age distribution and gender of patients included in this study reveals a sizeably younger population and a increased incidence of females as compared to previous studies. The success rate was high and approached the experience of other authors. Invariably the Mohs failures were among those patients that required six to eight sequential layers but failures do occur in primary cases outside the medial canthus.

Adult↗
PubMed1992

Head injured children and education: a need for greater delineation and understanding.

There is continuing professional misunderstanding within education and psychology regarding the effects of head injury upon the immature brain. Children who sustain repeated mild, or single severe head injuries in falls, road accidents, or as victims of physical abuse, may look good but perform badly. There is no acceptable evidence that such children achieve a good or full recovery. Conclusions about good prognosis cannot be substantiated in the present state of inadequate assessment including the perpetual reliance on inadequate or inappropriate global estimates, such as IQ, or Glasgow Outcome scores. The basis of many problems for head injured children within education is the apparent failure to adequately delineate the variables of mechanism and severity of injury, age at insult and time since injury.

Achievement↗
PubMed1992

Structure of the 5S rRNA genes in birch (Betula papyrifera) and alder (Alnus incana).

Hybridization of a 5S rDNA probe to Southern transfers of birch (Betula papyrifera) or alder (Alnus incana) DNA digested with BamH1 reveals similar triple-band "ladder-like" patterns. The sizes of sequenced 5S repeat units from both plants ranges only from 471 to 490 base pairs, suggesting that the complexity detected by Southern analysis is not due to different size classes of 5S repeats as found in other species. Within the intercistronic spacer region, conservation of large blocks of sequence between birch and alder 5S is observed implying a close evolutionary relationship between these two species. In both species, a duplication of part of the coding sequence including a restriction site for BamH1 introduces a second BamH1 site into the repeat unit. Differential methylation of the two BamHI restriction sites can account for the observed triple-band pattern.

Base Sequence↗
PubMed1992

Catecholamines in cerebrospinal fluid are increased by behavioral arousal and myocardial ischemia.

To study the central neural mechanisms involved in malignant ventricular arrhythmia, concentrations of norepinephrine in the cerebrospinal fluid were measured during behavioral stimulation and during coronary artery occlusion. Pigs were instrumented via thoracotomy with catheters to measure mean arterial pressure and plasma catecholamines and with silk snares around the left anterior descending coronary artery for occlusion after recovery from surgery. Cannulas were placed in the lateral ventricle of the brain to sample cerebrospinal fluid. Behavioral arousal was induced by lifting the pig in a canvas sling for 5 min. Mean arterial pressure, heart rate, and both plasma and cerebrospinal fluid norepinephrine concentrations increased significantly after lifting stimulation. In a separate experiment, 5 min after coronary artery occlusion, both plasma catecholamines and norepinephrine in cerebrospinal fluid were significantly elevated. Furthermore, pigs in which ventricular fibrillation occurred after occlusion had significantly higher concentrations of norepinephrine in cerebrospinal fluid before coronary artery occlusion.

Animals↗
PubMed1992

Control of ventricular fibrillation after coronary artery occlusion via intracerebroventricular injections.

The central nervous system (CNS) may play a larger role than previously thought in the development of ventricular fibrillation after coronary artery occlusion. The probability of ventricular fibrillation after complete, permanent occlusion of the left anterior descending coronary artery was 52% in conscious control pigs. After the administration into the lateral cerebral ventricle of tyrosine, the amino acid precursor of the catecholamine neurotransmitters, 100% of the animals developed ventricular fibrillation. After tyrosine plus propranolol, a beta-adrenoceptor antagonist, only 9% of pigs developed ventricular fibrillation. Treatment with propranolol alone did not affect the outcome. Catecholamine synthesis in the CNS may be associated with the development of ventricular fibrillation after coronary artery occlusion.

Animals↗
PubMed1992

Modulation of phencyclidine-sensitive ethidium binding to the Torpedo acetylcholine receptor: interaction of noncompetitive inhibitors with carbamylcholine and cobra alpha-toxin.

Ethidium is one of two fluorescent ligands known to bind to the noncompetitive inhibitor (NCI) site in the central ion channel of the Torpedo acetylcholine receptor with a micromolar dissociation constant. To further characterize heterotropic allosteric regulation of ligand binding in general, and of ethidium binding in particular, to the Torpedo receptor, we measured the effects of three liquid anesthetics (diethyl ether, halothane, and butanol), two barbiturates (secobarbital and thiamylal), and urethane. The phencyclidine-sensitive chromatic shift and the quantum yield increase associated with ethidium binding to the channel NCI site were used as indicators of ethidium binding. In the absence of other ligands, halothane, diethyl ether, and butanol increased the affinity of ethidium toward the channel NCI site to the same extent as carbamylcholine (400-600-fold), whereas the barbiturates and urethane were without effect. Cobra alpha-toxin blocked anesthetic-induced ethidium binding, confirming that cobra alpha-toxin stabilizes the AcChR in the resting-like state. In the presence of carbamylcholine, when ethidium was bound to the channel NCI site, several ligand-dependent effects were observed. 1) Without affecting further the affinity of ethidium for the NCI binding site, diethyl ether and halothane increased and butanol had no effect on the fluorescence emission of channel-bound ethidium. This indicated that there is little relation between the affinity and the quantum yield of the channel-bound ethidium. 2) Addition of secobarbital and thiamylal had no effect, beyond the effect of carbamylcholine, on ethidium binding to the channel NCI site, indicating that the barbiturates did not bind to the channel NCI site. 3) Urethane inhibited carbamylcholine-induced ethidium binding to the channel NCI binding site, suggesting direct interaction of urethane with the channel NCI binding site, at least when the receptor is in a desensitized state. The results confirm the conformational sensitivity of ethidium binding to the channel NCI binding site and demonstrate at least three different modes of action of anesthetics to inhibit the Torpedo receptor noncompetitively.

Anesthetics↗
PubMed1991

Rotational mobility of high-affinity epidermal growth factor receptors on the surface of living A431 cells.

The rotational diffusion of epidermal growth factor (EGF) bound to its specific receptor on the surface of human carcinoma A431 cells was studied by means of time-resolved phosphorescence anisotropy measurements. The rotational mobility was measured on the total population of EGF receptors by using a saturating concentration of EGF conjugated with a phosphorescent label, erythrosin, or on the subpopulation of high-affinity EGF receptors by using a low concentration of labeled EGF. At 4 degrees C, the rotational correlation times for both the high-affinity and total (mostly low affinity) receptor populations were in the range of 60-100 microns. Elevation of the temperature to 37 degrees C resulted in a lengthening of the rotational correlation time of the total receptor population to 200-300 microns, confirming a previous study of receptor microaggregation. The high-affinity EGF receptors were completely immobilized at 37 degrees C (rotational correlation time greater than 500 microns). The data are consistent with a model involving association of the cytoskeleton with the high-affinity receptors at 37 degrees C, but not at 4 degrees C.

Animals↗