Biomedical subjects

D Wade

Publications and source records attributed to D Wade.

At least 19 recordsLinked to original sources

PubMed1992

Shortened cecropin A-melittin hybrids. Significant size reduction retains potent antibiotic activity.

We have earlier reported two 26-residue antibacterial peptides made up from different segments of cecropin A (CA) and melittin (M). We now report a substantial reduction in size at the C-terminal section of the highly active hybrid CA(1-8)M(1-18), leading to a series of 20-, 18- and 15-residue analogs with antibiotic properties similar to the larger molecule. In particular, the 15-residue hybrids CA(1-7)M(2-9), CA(1-7)M(4-11) and CA(1-7)M(5-12) are the shortest cecropin-based peptide antibiotics described so far, with antibacterial activity and spectra similar or better than cecropin A and a 60% reduction in size. Their reduced size and highly alpha-helical structure require an alternative mechanism for their interaction with bacterial membranes.

Amino Acid Sequence↗

PubMed1992

Antibacterial peptides designed as analogs or hybrids of cecropins and melittin.

Eight new analogs of cecropin A, two new analogs of melittin and 30 hybrid peptides containing sequences from cecropins and melittin have been synthesized. The lengths of the peptides have varied from 37 residues (the length of cecropin A) to 18 residues. The peptides have been assayed for lysis of sheep red blood cells and for antibacterial activity against two Gram negative and three Gram positive bacteria. The best analogs of cecropin A maintained the anti-Escherichia coli activity of the parental peptide, and were not lytic for red blood cells. Melittin and its replacement analogs were all lytic for red blood cells, but an analog with transposed segments was not. Several of the hybrid peptides were found to be both non-hemolytic and highly active against all test bacteria. The data were used to define the structural requirements for antibacterial activity.

Amino Acid Sequence↗

PubMed1990

Hepatotropin mRNA expression in human foetal liver development and in liver regeneration.

A 569 bp probe against the beta-chain of hepatotropin was used to examine expression of RNA for this growth factor in human adult and foetal liver, foetal kidney and pancreas, and rat liver after partial hepatectomy. Low level expression of a 6 kb RNA occurred in human adult and normal rat liver. 70% hepatectomy increased expression, peaking at 10 h and returning to near normal levels 24 h after resection. The 6 kb band was strongly expressed in human foetal liver, as compared with adult, but not in foetal kidney or pancreas, suggesting a major role for hepatotropin in both foetal development and regeneration of the liver.

Animals↗

PubMed1990

Role of hepatic gamma-glutamyltransferase in the degradation of circulating glutathione: studies in the intact guinea pig perfused liver.

The role of hepatic gamma-glutamyltransferase in the breakdown of circulating glutathione was studied in the perfused guinea pig liver. Hepatic gamma-glutamyltransferase activity in the guinea pig is sevenfold higher than in the rat and is comparable to its activity in man. Guinea pig livers were found to remove, in a single pass, 50% to 90% of glutathione (10 to 50 mumol/L) added to the portal perfusate. Removal of portal glutathione was totally dependent on the activity of gamma-glutamyltransferase and led to the near quantitative appearance of cysteinyl-glycine and cysteine in the caval perfusate. Glutathione removal by the intact liver followed saturation with a Michaelis constant (Km) of 59 mumol/L for glutathione and a maximum velocity of 235 nmol glutathione/min/gm of liver weight. The capacity of the guinea pig liver to remove circulating glutathione was estimated to be sevenfold to 10-fold higher than its net rate of output of glutathione into the circulation. Inhibition of gamma-glutamyltransferase activity in the perfused liver led to threefold to sixfold increases in the hepatic output of glutathione into the circulation, indicating that more than two thirds of glutathione transported extracellularly is broken down. Data obtained demonstrate a major role of hepatic gamma-glutamyltransferase, both in the removal of portally carried glutathione and in the degradation of glutathione molecules released by the liver itself into the sinusoids. These findings suggest the existence of an intraorgan transport of glutathione in the liver, whereby periportal cells could provide glutathione precursors to pericentral cells.

Animals↗

PubMed1990

Persistent metabolic sequelae of severe head injury in humans in vivo.

Six patients who had suffered severe non-penetrating high velocity head injuries were investigated with phosphorus (31P) magnetic resonance spectroscopy (MRS) to determine, non-invasively, long-term alteration in intracellular biochemistry. The normal subjects were found to have a constant intracellular pH (pHi, 7.03 +/- 0.03) with depth into the brain. The adenosine triphosphate (ATP, 3.46 +/- 0.66 mmol/L of brain tissue), inorganic phosphate (Pi, 1.15 +/- 0.41 mmol/L) and phosphomonester (PME, 2.76 +/- 1.0 mmol/L) tissue concentrations did not alter significantly with depth into normal brain. The phosphocreatine (PCr, 2 cm = 5.21 +/- 1.25, 5 cm = 4.85 +/- 1.49 mmol/L) was slightly reduced, whilst phosphodiesters (PDE, 2 cm = 9.53 +/- 2.6, 5 cm = 14.41 +/- 4.2 mmol/L) rose significantly between tissue comprising mainly of gray (2 cm) and white matter (5 cm). In comparison the contra-lateral hemisphere to the side of worst spasticity showed significant changes a considerable time after injury (6-18 months). The intracellular metabolite tissue concentrations were all reduced by 30% (ATP 2.53 +/- 1.0 mmol/L, PCr 3.44 +/- 0.8 mmol/L) with PDE reduced most significantly at depth (5 cm = 8.4 +/- 3.4 mmol/L), compatible with the cerebral atrophy seen in these patients. In white matter the pHi also decreased with depth (2 cm = 7.03 +/- 0.03, 5 cm = 6.89 +/- 0.05). The reduction in pHi so long after injury is difficult to explain in these steady-state conditions. A structural abnormality, such as a disorder in the blood brain barrier or accumulation of large acidic lysosomes, could cause these pHi changes. There may also be a failure in blood flow regulation, with near critical fluctuations in blood flow both with time and space.

Accidents, Traffic↗

PubMed1990

All-D amino acid-containing channel-forming antibiotic peptides.

The D enantiomers of three naturally occurring antibiotics--cecropin A, magainin 2 amide, and melittin--were synthesized. In addition, the D enantiomers of two synthetic chimeric cecropin-melittin hybrid peptides were prepared. Each D isomer was shown by circular dichroism to be a mirror image of the corresponding L isomer in several solvent mixtures. In 20% hexafluoro-2-propanol the peptides contained 43-75% alpha-helix. The all-D peptides were resistant to enzymatic degradation. The peptides produced single-channel conductances in planar lipid bilayers, and the D and L enantiomers caused equivalent amounts of electrical conductivity. All of the peptides were potent antibacterial agents against representative Gram-negative and Gram-positive species. The D and L enantiomers of each peptide pair were equally active, within experimental error. Sheep erythrocytes were lysed by both D- and L-melittin but not by either isomer of cecropin A, magainin 2 amide, or the hybrids cecropin A-(1-13)-melittin-(1-13)-NH2 or cecropin A-(1-8)-melittin-(1-18)-NH2. The infectivity of the bloodstream form of the malaria parasite Plasmodium falciparum was also inhibited by the D and L hybrids. It is suggested that the mode of action of these peptides on the membranes of bacteria, erythrocytes, plasmodia, and artificial lipid bilayers may be similar and involves the formation of ion-channel pores spanning the membranes, but without specific interaction with chiral receptors or enzymes.

Amino Acid Sequence↗

PubMed1990

Assessing motor impairment after stroke: a pilot reliability study.

Two short tests of motor function, the Motricity Index (MI) and the Trunk Control Test (TCT), were assessed at regular intervals after stroke and compared with a detailed physiotherapy test, the Rivermead Motor Assessment (RMA). The MI and TCT were valid and reliable tests which were usually quicker to perform than the RMA. The TCT was of predictive value when related to eventual walking ability. All three tests appeared to be of equal sensitivity in detecting change.

Adolescent↗

PubMed1989

Antibacterial and antimalarial properties of peptides that are cecropin-melittin hybrids.

Solid phase synthesis was used to produce 5 hybrid peptides containing sequences from the antibacterial peptide, cecropin A, and from the bee venom toxin, melittin. Four of these chimeric peptides showed good antibacterial activity against representative Gram-negative and Gram-positive bacterial species. The best hybrid, cecropin A(1-13)-melittin(1-13) was 100-fold more active than cecropin A against Staphylococcus aureus. It was also a 10-fold better antimalarial agent than cecropin B or magainin 2. Sheep red cells were lysed by melittin at low concentrations, but not by the hybrid molecules, even at 50 times higher concentrations.

Amino Acid Sequence↗

PubMed1987

Deuterium isotope effect on denitrosation and demethylation of N-nitrosodimethylamine by rat liver microsomes.

In an attempt to elucidate the molecular basis for the decrease in rat liver carcinogenicity and DNA-alkylating ability that accompanies deuteration of N-nitrosodimethylamine (NDMA), NDMA and its fully deuterated analogue ([2H6]NDMA) were incubated with acetone-induced rat liver microsomes. Rates for the competing metabolic routes, denitrosation and demethylation, were determined from colorimetric data on nitrite and formaldehyde generation, respectively. The Vmax calculated for demethylation of NDMA was 7.9 nmol/min/mg, while that for denitrosation was 0.83 nmol/min/mg. Deuteration of NDMA did not significantly change the Vmax for either pathway, but it did increase the Km for demethylation from 0.06 to 0.3 mM. The Km for denitrosation was also increased from 0.06 to 0.3 mM on deuteration, as determined by incubating an equimolar mixture of amino-15N-labeled NDMA with [2H6]NDMA and measuring the methyl[15N]amine:[2H3]methylamine ratio by derivatization-gas chromatography-mass spectrometry. The fact that the Km values for denitrosation were so similar to those for demethylation suggested that the two pathways were catalyzed by the same enzyme. The isotope effects calculated from these data [VmaxH/VmaxD approximately 1 and (Vmax/Km)H/(Vmax/Km)D approximately 5] show that microsomal metabolism of NDMA is not significantly shifted from demethylation to denitrosation on deuteration of substrate and may indicate a low commitment to catalysis for the enzyme. The results are consistent with the view that the metabolism of NDMA is initiated by formation of an alpha-nitrosamino radical which either combines with a hydroxyl radical to form the alpha-hydroxynitrosamine as the initial product of the demethylation pathway or fragments to nitric oxide and N-methylformaldimine as the first products of denitrosation.

Animals↗

PubMed1987

Nature of N-nitrosodimethylamine demethylase and its inhibitors.

The present study was undertaken to examine the nature of the low Km (KmI) form of rat liver microsomal N-nitrosodimethylamine demethylase (NDMAd) and its inhibition by organic compounds which are commonly present in the assay mixture. Using radiometric and colorimetric assay methods with an NADPH-generating system consisting of 0.4 mM NADP, 10 mM glucose-6-phosphate, and glucose-6-phosphate dehydrogenase (0.4 units/ml), Km values of 40-50 microM were obtained. These Km values were lower than the values of 60-80 microM reported previously. This decrease was due to the elimination of inhibitors such as glycerol in the assay mixture. Glycerol was a competitive inhibitor, and this observation explained in part why purified P-450ac (acetone-inducible form of P-450), displayed a higher Km value in a reconstituted NDMAd system, which contained glycerol, than in microsomes. Semi-carbazide which had been used in many previous assays of NDMAd was also found to be a competitive inhibitor of this enzyme. Other inhibitors studied include the commonly used solvents dimethylsulfoxide, acetone, ethylene glycol, dimethylformamide, ethyl acetate, benzene, and hexane as well as thiol compounds dithiothreitol and mercaptoethanol. Although very low Km values (10-20 microM) for N-nitrosodimethylamine metabolism were reported in studies with perfused liver, liver slices, and isolated liver cells, we believe that the KmI form of liver NDMAd is responsible for the metabolism and activation of N-nitrosodimethylamine in the rat liver.

1-Propanol↗

PubMed1987

Concurrent generation of methylamine and nitrite during denitrosation of N-nitrosodimethylamine by rat liver microsomes.

With the goal of identifying the organic amine product(s) of enzymatic N-nitrosodimethylamine (NDMA) denitrosation, 4 mM NDMA was incubated with liver microsomes from ethanol-treated rats. The concentrations of dimethylamine and methylamine were determined by derivatization with 2,4-dinitrofluorobenzene followed by gas chromatography-mass spectrometry. There was no net increase in the concentration of dimethylamine during incubation, but the yield of methylamine was equimolar with that of nitrite. Additional incubations of NDMA using acetone-induced microsomes, 1 mM and 0.1 mM substrate, gave methylamine/nitrite ratios of 0.9 and 0.7, respectively, confirming the quantitative linkage between these two products. Control incubations conducted with pure methylamine or dimethylamine indicated that the secondary amine is not a significant intermediate in the metabolic generation of the primary amine. Experiments with 15N-labeled NDMA showed that the methylamine nitrogen came from the amino moiety of the nitrosamine. The results suggest that NDMA metabolism is best viewed as a competition between at least two important pathways, demethylation and the presumably deactivating denitrosation route, a formulation which seems to account for the previously reported detection of methylamine as a urinary metabolite of NDMA and for the production of less than theoretical yields of labeled dinitrogen gas during NDMA metabolism.

Acetone↗

PubMed1987

Effect of food and various antacids on the absorption of tenoxicam.

1 Twelve healthy volunteers received a single oral dose of tenoxicam 20 mg on six occasions separated by 3 weeks. 2 The six occasions were: fasted overnight; postprandial; fasting and 15 ml aluminium hydroxide gel; postprandial and 15 ml aluminium hydroxide gel; fasting and 15 ml aluminium and magnesium hydroxide gel; postprandial and 15 ml aluminium and magnesium hydroxide gel. 3 Twenty plasma samples were collected over 15 days following dosing with tenoxicam. 4 The following kinetic parameters for plasma tenoxicam were compared: peak concentrations, time taken to reach peak concentrations, area under the plasma concentration-time curve (AUC) and half-life of elimination. 5 Food lengthened the time taken to reach peak tenoxicam concentrations (5.82 +/- 4.6 vs 1.84 +/- 1.0 h in the fasting state; P less than 0.02) and marginally reduced the peak concentrations achieved. AUC was not affected by any of the different regimens. 6 These effects of food on tenoxicam bioavailability are unlikely to be of clinical significance during chronic dosing with the drug.

Adult↗

PubMed1987

Kinetic isotope effect on the demethylation and denitrosation of N-nitrosodimethylamine in vitro.

Deuteration of N-nitrosodimethylamine (NDMA) has been shown to decrease the carcinogenicity of this compound. This result is believed to be due to a kinetic isotope effect on the metabolic activation of this carcinogen, but conflicting views exist concerning whether the isotope substitution affects the Km or Vmax of the reaction. In order to elucidate the molecular basis of these observations, as well as the mechanisms of the demethylation and denitrosation reactions, the metabolism of NDMA and deuterated NDMA (NDMA-d6) was studied using acetone-induced rat-liver microsomes. The demethylation of NDMA displayed a Km of 0.06 mM and a Vmax of 7.9 nmol/min per mg protein. Deuteration of NDMA increased the Km value by five fold but did not appreciably affect the Vmax. The denitrosation of NDMA also displayed a Km of 0.06 mM, but the Vmax was 0.83 nmol/min per mg; deuteration again increased the Kmax several fold but had no effect on the Vmax. The results indicate that deuteration inhibits the metabolism of NDMA by increasing the Km but not the Vmax and suggest that there is a close relationship between the demethylation and denitrosation reactions.

Acetone↗

PubMed1984

Binding of ethidium and bis(methidium)spermine to Z DNA by intercalation.

The interaction of ethidium bromide, a DNA intercalating drug, and bis( methidium )spermine, a DNA bis-intercalating compound, with the left-handed Z form of poly(dG-dC) has been studied in 4.4 M NaCl. Spectrophotometric analysis using absorption, fluorescence and circular dichroism indicates that the complex formed between ethidium and Z DNA resembles very closely that formed with B DNA. This suggests that ethidium binds to Z DNA by intercalation. 31P NMR spectra are presented showing both the conversion of the Z form to the B form with increasing amounts of drug and the typical Z form spectrum at low binding densities. Data are also presented which show that the bifunctional intercalator bis( methidium )spermine binds to Z DNA in a manner similar to its binding to B DNA, i.e., by bis-intercalation. These results are important for our understanding the behavior of Z DNA and its biological significance.

Animals↗

PubMed1983

Clonidine raises blood pressure in severe idiopathic orthostatic hypotension.

The hemodynamic effects of clonidine were studied in four patients with sever idiopathic orthostatic hypotension and one patient with baroreceptor dysfunction. No depressor response to clonidine was observed in any patient with idiopathic orthostatic hypotension at any dosage. Rather, two patients responded to 0.4 mg of oral clonidine with a 40 mm Hg increment in systolic blood pressure lasting several hours. Each has been receiving clonidine, 0.4 mg twice daily, for one year with greatly increased functional capacity. The other two patients with idiopathic orthostatic hypotension had an even greater pressor response to 0.8 mg of oral clonidine, but adverse effects prevented continued therapeutic use. In marked contrast, the patient with baroreceptor dysfunction had a profound depressor response to 0.2 mg of clonidine. In the treatment of idiopathic orthostatic hypotension, the major advantage of clonidine over other pressor agents is its longer duration of action. The major adverse effects of the drug in these patients are sedation, dry mouth, altered mentation, and excessive hypertension. The drug should not be given to patients with mild idiopathic orthostatic hypotension or selective baroreceptor dysfunction, since severe hypotension may result.

Aged↗

PubMed1983

Effects of phenytoin, phenobarbital, and ascorbic acid on misonidazole elimination.

The kinetics of an oral dose (1.0 gm/m2) of the 2-nitroimidazole radiosensitizer misonidazole were studied in three groups of six healthy subjects before and after a 1-wk course of phenytoin, phenobarbital, or ascorbic acid. Phenytoin and phenobarbital decreased mean misonidazole half-life by 27% and 23% and the decrease was associated with the respective increases in mean clearance of 42% and 31%. The area under the plasma concentration-time curve for the metabolite O-desmethylmisonidazole increased correspondingly. Volume of distribution of misonidazole was unchanged. After treatment with ascorbic acid there was a very small increase in the mean clearance of misonidazole, but there was no significant change in other kinetic parameters. Induction by phenytoin and phenobarbital of the oxidative metabolism of misonidazole is the most likely mechanism responsible. Deliberate induction of a patient's metabolism may help to reduce the neurotoxicity associated with the use of the drug. The efficacy of the radiosensitizing action of the drug is unlikely to be compromised under these conditions since peak plasma concentrations of misonidazole were not affected by treatment with either phenytoin or phenobarbital. The potentiation of the cytotoxic effects of misonidazole by ascorbic acid is unlikely to be related to a direct effect on the oxidative metabolism of misonidazole.

Adult↗

PubMed1981

Tolerance to the humoral and hemodynamic effects of caffeine in man.

Acute caffeine in subjects who do not normally ingest methylxanthines leads to increases in blood pressure, heart rate, plasma epinephrine, plasma norepinephrine, plasma renin activity, and urinary catecholamines. Using a double-blind design, the effects of chronic caffeine administration on these same variables were assessed. Near complete tolerance, in terms of both humoral and hemodynamic variables, developed over the first 1-4 d of caffeine. No long-term effects of caffeine on blood pressure, heart rate, plasma renin activity, plasma catecholamines, or urinary catecholamines could be demonstrated. Discontinuation of caffeine ingestion after 7 d of administration did not result in a detectable withdrawal phenomenon relating to any of the variables assessed.

Adult↗

PubMed1977

Pigmentation and musculoskeletal abnormalities in an aged state hospital population.

In a State hospital population, 686 patients (mean age, 65 years) were examined for patterned pigmentation on unexposed areas of the body. Of these patients, 285 (41.5 percent) met the criteria for pigmentation. There were no sex or age differences. Psychosis had been diagnosed in 96 percent of the pigmented group. All of the 239 patients examined orthopedically had vertebral malalignment greater than 15 degrees and often had other bony deformities. Nearly all of the pigmented group displayed dyskinetic activity. A possible mechanism for the production of the pigmentation-psychosis-skeletal deformity triad could be an alteration in the concentration of available melatonin. Continued research along these lines may lead to an effective means of pharmacologic treatment as well as better psychologic and physical care.

Adult↗