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D A Johnson

Publications and source records attributed to D A Johnson.

At least 145 records · Page 8Linked to original sources
PubMed1994

The molecular diversity of the 5S rRNA gene in barley (Hordeum vulgare).

The 5S rRNA genes from several accessions of cultivated barley, Hordeum vulgare L., were amplified by the polymerase chain reaction, cloned, and sequenced. Analysis of the aligned sequences, followed by principal coordinate analysis, support the recognition of at least two distinct classes of 5S rDNA genes. The short repeat class corresponds to the 300-bp tandem repeat defined by E.V. Ananiev as containing several TAG repeating units. The long repeat class contains long tandem repeats and lacks the TAG repeating unit. Sequences in each class can be further subdivided, with the long repeat class containing two groups and the short repeat class containing two and possibly three groups. These results suggest that in cultivated barley the sequence diversity found within the 5S rDNA nontranscribed spacer region may be encoded by three or more loci and may be useful for phylogenetic analyses provided that orthology can be established.

Base Sequence↗
PubMed1994

Ozone, but not nitrogen dioxide, fragments elastin and increases its susceptibility to proteolysis.

The effects of ozone (O3) and nitrogen dioxide (NO2) on the solubility and proteolytic susceptibility of elastin were examined to better understand how these oxidant air pollutants might damage the lung. In vitro O3 exposures at pH 7.4 resulted in the complete solubilization of elastin, but NO2 had no effect on solubility. The initial solubilization rate was 65 micrograms/mumol of O3, which increased to 150 micrograms/mumol in the midregion of a sigmoidal solubilization curve. Peptide fragments of the O3-solubilized elastin ranged in size from 5 to 20 kD. The conversion of insoluble elastin into soluble fragments by O3 was not due to the destruction of desmosine crosslinks. The effect of O3 on the proteolytic susceptibility of elastin was measured using insoluble elastin recovered from exposures that resulted in 5.3%, 12.8%, and 26.3% solubilization. Human neutrophil elastase (HNE) digested the remaining insoluble elastin samples 4.3, 6.0, and 9.8 times faster than unexposed elastin. In contrast, NO2-exposed elastin was no more susceptible to digestion by HNE. Ascorbate, EDTA, and uric acid reduced the proteolytic susceptibility of O3-exposed elastin, but mannitol afforded no protection. These findings indicate that the inhalation of O3 may contribute to lung disease by directly damaging elastin and by increasing its susceptibility to proteolysis, whereas NO2 probably damages lungs via alternative mechanisms.

Air Pollutants↗
PubMed1994

Management of penetrating colon injuries.

The management of colon injuries remains an area of major controversy. Selecting the patients who can undergo primary repair safely remains undefined. To address this issue, 231 consecutive patients with penetrating colon injuries were reviewed to determine those factors that affected outcome. Overall, there were 54 (25.2%) septic complications, with 36 (16.8%) wound infections and 18 (8.4%) intra-abdominal abscesses. There were seven (3.3%) deaths in the entire series. The surgical management method of the colon injury was not significant in wound infections (P > .39), intra-abdominal abscesses (P > .24), or mortality (P > .39). A more aggressive approach of primary repair should be performed for civilian colon injuries.

Abscess↗
PubMed1993

Fluorescence resonance energy transfer within a heterochromatic cAMP-dependent protein kinase holoenzyme under equilibrium conditions: new insights into the conformational changes that result in cAMP-dependent activation.

Previous studies of the ligand regulation of the cAMP-dependent protein kinase have demonstrated the cAMP-mediated dissociation of the holoenzyme by using nonequilibrium techniques; i.e., gel filtration, ion-exchange chromatography, and differential centrifugation. While physically mild, these could have caused weakly associated species to dissociate, thereby providing a potentially flawed interpretation of the mechanism of activation of the protein kinase. To assess this, the activation of the cAMP-dependent protein kinase has been monitored under equilibrium conditions using dipolar fluorescence energy transfer to measure changes in the proximity relations between the catalytic (C) and regulatory (R) subunits that compose the holoenzyme. Specifically, we prepared a heterochromatically labeled protein kinase type II holoenzyme, with the regulatory and catalytic subunits labeled with sulforhodamine and carboxyfluorescein, respectively, and monitored the exchange of electronic excitation energy between the C and R subunits by both donor lifetime and steady-state fluorescence. Biochemically, the heterochromatic holoenzyme was closely identical to the native protein with regard to cAMP-induced increase in catalytic activity, reassociation of C and R subunits, inhibition of catalytic activity by the specific protein kinase inhibitor (PKI), and observed dissociation examined by gel filtration upon cAMP addition. However, under equilibrium conditions, the energy-transfer measurements revealed that the addition of cAMP to this heterochromatic reporter complex promoted an estimated 10-A increase in the distance between the derivatization sites on C and R but not a dissociation of these subunits. Addition of PKI plus cAMP promoted full dissociation of the two subunits.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1993

Quinacrine and ethidium bind to different loci on the Torpedo acetylcholine receptor.

Fluorescence spectroscopy was used to determine whether quinacrine and ethidium, two high-affinity noncompetitive inhibitors of the Torpedo acetylcholine receptor (AcChR), bind to the same loci. The ability of three nitroxide spin-labels, 5-doxylstearate (5-SAL), spin-labeled androstane (ASL), and TEMPO, to quench receptor-bound quinacrine and ethidium fluorescence was measured. When bound to a phencyclidine-displaceable site on the AcChR, quinacrine was 16.9 and 19 times more efficiently quenched than ethidium by the highly lipophilic 5-SAL and ASL, respectively. TEMPO, which has a limited ability to partition into Torpedo plasma membranes (< 1%), was only twice as efficient at quenching receptor-bound quinacrine than ethidium fluorescence. The relative sensitivity of quinacrine and ethidium fluorescence to paramagnetic quenching was examined in three solvents, 1-butanol, sodium phosphate buffer, and acetonitrile, with TEMPO as a quencher. The results from the different solvents demonstrate that quinacrine fluorescence is intrinsically 1.4-3.6 times more sensitive than ethidium fluorescence to quenching by nitroxide spin-labels. Examination of the effect of high concentrations of 5-SAL on ethidium and quinacrine dissociation constants showed that quinacrine but not ethidium binding was competitively inhibited. Together, these results indicate that although quinacrine and ethidium bind in a mutually exclusive manner, the two inhibitors interact at different loci on the AcChR. Whereas the ethidium binding site is at a distance from membrane lipids, probably in or near the lumen, the quinacrine binding site appears to be at a lipid-protein interface in the transmembrane domain and at a distance from the lumen.

Animals↗
PubMed1993

Transverse localization of the quinacrine binding site on the Torpedo acetylcholine receptor.

We demonstrated previously that a phencyclidine-displaceable quinacrine binding site exists at the lipid-protein interface of the Torpedo acetylcholine receptor (AcChR) (Valenzuela, C. F., Kerr, J. A., and Johnson, D. A. (1992) J. Biol. Chem. 267, 8238-8244). In this manuscript, we assess (1) the transverse position of this site in the lipid bilayer by examining the ability of a series of paramagnetic n-doxyl stearates (n-SALs) and iodide to quench receptor-bound quinacrine and membrane-partitioned octadecyl rhodamine B (C18-Rho) fluorescence and (2) the stoichiometry of histrionicotoxin- or phencyclidine-displaceable quinacrine binding. Initial experiments established what fraction of the n-doxyl stearates partitioned into the membranes and that the n-doxyl stearates do not interfere with quinacrine binding to the receptor at the concentrations used in the quenching studies. The n-doxyl stearate quenching experiments indicated relatively small (< 2) differences between the n-doxyl stearates to quench receptor-bound quinacrine fluorescence, with a rank order of 7-SAL > or = 5-SAL > 12-SAL > 16-SAL. This contrasts with the n-doxyl stearate quenching of the membrane-partitioned C18-Rho which showed as much as an 8.6-fold difference between the various isomers with a rank order of quenching efficiencies of 5-SAL > 7-SAL > 12-SAL > or = 16-SAL. Iodide quenching measurements indicated significant solute accessibility to membrane-partitioned C18-Rho but not to receptor-bound quinacrine. The ratios of the bimolecular quenching rate constants for free to bound quinacrine and for free rhodamine B to membrane-partitioned C18-Rho were 53.4 and 6.6, respectively. Direct titration of quinacrine into suspensions of a high concentration of AcChR-associated membranes yielded an upper limit to the binding stoichiometry of 1.4 HTX- or PCP-displaceable quinacrine binding sites/AcChR functional units. The results suggest that there is a single phencyclidine- or histrionicotoxin-displaceable quinacrine binding site located at or somewhat below the level of the C5-C7 in the phospholipid acyl chains at the lipid-protein interface.

Animals↗
PubMed1993

Nitrogen dioxide reactivity with proteins: effects on activity and immunoreactivity with alpha-1-proteinase inhibitor and implications for NO2-mediated peptide degradation.

Nitrogen dioxide (NO2), an air pollutant produced by burning fossil fuels and a component of cigarette smoke, is thought to contribute to the pathogenesis of pulmonary diseases, such as emphysema. In order to gain information on the mechanism by which NO2 damages the lung and proteins vital to its function, as well as its reaction with proteins in general, in vitro exposures of alpha-1-proteinase inhibitor (alpha 1PI), elastin, poly-L-lysine, and poly-L-arginine were performed. The ability of alpha 1PI to inhibit its natural physiological target, human neutrophil elastase (HNE), declined with exposure to 54% of the control value at molar ratios of NO2:alpha 1PI of 400:1 and greater. Exposure of alpha 1PI to NO2 resulted in a 50% loss of immunoreactivity with either monoclonal or polyclonal antibodies in an enzyme-linked immunosorbent assay at molar ratios of NO2:alpha 1PI of 100:1 and greater. The results of parallel O-phthalaldehyde and bicinchoninic acid protein assays as well as amino acid analysis on control and NO2-exposed alpha 1PI suggested a reactivity of NO2 with lysine residues. Elastin and poly-L-lysine were labeled by reductive methylation of amino groups with [3H]HCHO prior to treatment with NO2 in aqueous solutions at physiological pH. NO2 exposure of elastin resulted in the solubilization of 84% of the associated radioactivity of which 79% was identified as [3H]methyllysine by amino acid analysis. After NO2 exposure of poly-L-[3H]lysine, gel filtration chromatography revealed that the 50,000 M(r) poly-L-[3H]lysine had been degraded to small peptides of 1-3000 M(r). Similarly, after NO2 exposure of unlabeled poly-L-arginine, gel filtration chromatography, and total peptide analysis revealed that the 47,500 M(r) peptide was also partially degraded to peptides. These results suggest that NO2 reacts with the epsilon-amino groups of Lys residues (primary amines) and with the amide nitrogen (secondary amines) of surface-exposed Lys and Arg residues in the peptide backbone to result in peptide bond cleavage. These findings are the first indication of NO2-mediated peptide degradation and provide additional data on the potential of NO2 to damage proteins vital to the function of the lung in an in vitro exposure system.

Elastin↗
PubMed1993

Human recombinant interferon-beta SER and tamoxifen: growth suppressive effects for the human breast carcinoma MCF-7 grown in the athymic mouse.

Tamoxifen is the endocrine treatment of choice for breast cancer. However, resistance to therapy and patient relapse inevitably occurs. In future treatment schedules, interferons could be administered with tamoxifen, in an attempt to prevent disease recurrence. Human recombinant interferon-beta SER (rIFN-beta SER) inhibited the growth in vitro of the estrogen receptor (ER) positive breast cancer cell line MCF-7 and the ER negative breast cancer cell line MDA-MB-231. This inhibitory effect was achieved at doses of 50 U/ml and above. The growth of MCF-7 tumors in estradiol-stimulated athymic mice was greatly inhibited by high dose rIFN-beta SER treatment (10(6)U/day). In spite of the impressive antitumor effects upon MCF-7 tumors, rIFN-beta SER had no effect upon ER levels within the tumors at either the RNA or protein level, as measured by Northern blotting and ER-EIA respectively. High dose rIFN-beta SER (10(6)U/day) did result in some inhibition in the growth in vivo of the tamoxifen-stimulated MCF-7 variant MCF-7 TAM, although not to the same extent as was observed with the estradiol-stimulated MCF-7 tumors. rIFN-beta SER was also administered to animals bearing MCF-7 tumors and treated with estradiol and tamoxifen. In the animals undergoing high dose therapy (10(6)U/day), tumor growth was completely suppressed. Furthermore, tumor growth continued to be suppressed in those animals in which the rIFN-beta SER therapy was halted and the tamoxifen capsule removed. No tumors were observed in spite of the environment of estradiol stimulation. Thus, the combination of interferon and tamoxifen was totally growth suppressive for MCF-7 xenografts in nude mice.

Animals↗
PubMed1993

The effects of intermittent progesterone upon tamoxifen inhibition of tumor growth in the 7,12-dimethylbenzanthracene rat mammary tumor model.

The development of endometrial cancer is a potential risk during long-term tamoxifen therapy for breast cancer. In order to protect the uterus, progestin treatment has been proposed for these patients. However, within the 7,12-dimethylbenzanthracene-induced rat mammary model, progesterone is known to reverse the antitumor effects of tamoxifen. This study shows that progesterone administered intermittently still reverses the antitumor effects of tamoxifen in this model. This effect of progesterone is not due to a decrease in the tissue levels of tamoxifen, and may be direct, via the progesterone receptor.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1993

Influence of mastication on saliva, plaque pH and masseter muscle activity in man.

An earlier study showed that frequent gum chewing may enhance parotid gland function and reduce the acidogenicity of dental plaque. The aim now was to determine whether these effects would be observed after a 2-week period of diet altered to increase masticatory effort, and secondarily to assess the effects of chewing gum on masseter muscle activity. Ten subjects took part in the first experiment. Saliva was collected before and after the diet change and the plaque pH response to a sucrose challenge was measured. Subjects completed 3-day diet histories and wore electromyographic (EMG) devices to record masseter activity. In the second experiment, 10 subjects wore EMG devices for 3 days to record masseter activity on three daily regimens: baseline (no gum chewing), hourly gum chewing (sugar-free gum chewed for 10 min every hour) and chewing five sticks of gum each for 20 min during the day. Data were analysed by paired t test or repeated-measures analysis of variance. For the first experiment, EMG data indicated significant increases in chewing activity (p < 0.05), although there were no changes in salivary flow rates or the plaque pH response to sucrose. The second experiment showed that total EMG activity increased significantly on both gum-chewing regimens (p < 0.01), the magnitude of the increase being greater for hourly chewing. Overall, masseter EMG activity was increased 41% by diet alteration, compared to increases of 96 and 152% on the five-stick and hourly gum-chewing regimens, respectively.

Adult↗
PubMed1993

Long-term visual results and complications in children with aphakia. A function of cataract type.

PURPOSE: Previous studies of outcome in children with aphakia have approached the data by grouping patients according to features such as age at surgery, type of procedure, or some other common attribute. The purpose of this study is to identify factors predictive for visual outcome and complications in pediatric patients with cataracts. METHOD: One hundred seventy-four eyes in 118 patients underwent lensectomy and anterior vitrectomy for congenital or juvenile cataracts. All received early optical correction, occlusion therapy when necessary, and follow-up for at least 6 months. Visual outcome and complications were analyzed statistically to determine predictive factors. Features analyzed included cataract type, laterality, age at onset, follow-up, and corneal size. RESULTS: Statistical analysis showed that the most important predictor of long-term visual outcome and complications is cataract type. Visual outcome differed significantly by cataract type, with best results in the lamellar and posterior lentiglobus groups. Unilateral cases had a mean acuity lower than bilateral cases. Other factors, such as age at surgery and corneal size, were less predictive but closely linked to cataract type. Complications, such as aphakic glaucoma, also were more closely linked to the type of cataract than to other variables. CONCLUSION: The results indicate that an important determinant of long-term outcome and complications in aphakic children is cataract type. Other features were found to be closely correlated to cataract type and were not independently significant.

Age Factors↗
PubMed1993

Effects of food mastication on rat parotid gland adrenergic and cholinergic cell surface receptors.

Adult male rats were fed diets of differing texture (liquid, powder, standard pelleted, or bulk pelleted) to alter food mastication. After 2 weeks, the parotid glands were removed and adrenergic and muscarinic-cholinergic cell surface receptor density (fM bound/mg protein) and ligand binding dissociation constants (Kd in nM) were determined by radioligand binding techniques on a crude membrane fraction. For all diets, gland weight increased as the requirement for food mastication increased (i.e., liquid < powder < standard pelleted < bulk pelleted). Among the diets, neither beta-two nor alpha-two receptor density was altered. Beta-one receptor density was directly related to dietary mastication. Compared with the standard pelleted diet, beta-one receptor density was reduced 21% for the liquid diet and 7% for the powdered diet; for the bulk-pelleted diet, beta-one receptor density was increased 11%. With respect to alpha-one receptor density, it was not affected by the liquid or powdered diet when compared with the standard pelleted diet, but alpha-one receptors were increased 14% with the bulk-pelleted diet. Muscarinic-cholinergic receptor density for the liquid diet fed rats was 27% less than for the standard-pelleted diet; powdered diet did not differ from standard pelleted, while that for the bulk-pelleted diet was increased 6%. With but minor exceptions, ligand binding affinity was unaffected by the changes in diet texture. These studies demonstrate that dietary mastication as well as affecting parotid gland weight, cell size, and saliva production also influences autonomic cell surface receptor density.

Animals↗
PubMed1993

Fine-needle aspiration cytology of the thyroid. A 12-year experience with 11,000 biopsies.

The primary purpose of FNA biopsy is to identify benign nodules and avoid unnecessary surgery. Numerous reports confirm that the introduction of FNA reduces thyroid operations by 25% and increases the yield of carcinoma from 14% to at least 30%. With an improvement in surgical selectivity, the yield of carcinoma has significantly increased, and the impact of FNA on thyroid practice has been substantial. The economic impact of FNA biopsy is also considerable and makes this test a useful and cost-effective procedure. Our approach to a patient with nodular thyroid disease is outlined in Figure 1. This scheme rests on thyroid cytology, and FNA biopsy is used as the first diagnostic test. When an experienced clinician performs the aspiration and an experienced cytopathologist reviews the slides, the accuracy of the technique is better than 90%. In agreement with recent FNA series, we believe that FNA biopsy is a safe and reliable procedure and that complications are extremely rare. On the basis of our experience, its continued use as the first step in the diagnostic management of thyroid nodules seems justified.

Biopsy, Needle↗
PubMed1993

Management of extraperitoneal rectal injuries.

Twenty-eight consecutive extraperitoneal rectal injuries for a period of 34 months ending in May 1990 were reviewed retrospectively. All injuries were due to penetrating gunshot wounds. The rectal exam was positive in 75% of patients versus 80.8% with proctosigmoidoscopy. All 28 patients had diversion of the fecal stream. Diverting colostomies were performed in 17 patients, Hartmann's colostomies in 7 patients, and proximal loop colostomies in 4 patients. Presacral drainage was used in 25 patients (89.3%). Distal irrigation was performed in 13 patients (46.4%) and primary repair in 9 patients (32.1%). There was one infectious complication (3.6%) and no deaths (0%). Fecal diversion and presacral drainage are the mainstay of therapy for civilian rectal injuries. The importance of distal irrigation of the rectum has not been established. Primary repair of the rectum has no effect on morbidity and mortality.

Adolescent↗
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↗