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K L Allen

Publications and source records attributed to K L Allen.

At least 19 recordsLinked to original sources
PubMed2003

Tremorgenic mycotoxins increase gastric smooth muscle activity of sheep reticulum and rumen in vitro.

Reticulum and rumen strips (consisting of both muscle layers and the myenteric plexus) were superfused with Tyrode Ringer and their contractions recorded isometrically. The strips were subjected to exogenous acetylcholine and electrical field stimulation (EFS) resulting in contractions that could be blocked by atropine. Responses to the tremorgenic mycotoxin penitrem A and others thought to be involved in ryegrass staggers, paxilline and lolitrem B (10(-10)-10(-6)M), were compared with those of control vehicle (0.1% DMSO). The tremorgens were without effect on quiescent preparations. Penitrem A and paxilline enhanced spontaneously active preparations and the amplitude of contractions in response to EFS. Responses to paxilline had a shorter latency than to penitrem A. Responses of spontaneously active preparations were resistant to atropine. Penitrem A, but not paxilline, increased responses to exogenous acetylcholine. Lolitrem B (10(-6)M) increased responses to EFS, but many responses were equivocal, possibly due to the lower solubility of lolitrem B in aqueous solutions compared to the other tremorgens. The results show that these mycotoxins have peripheral excitatory effects on the reticulorumen and it is suggested that such activity in vivo may reflexly affect centrally derived cyclical contractions.

Animals↗
PubMed1997

A hematopoietic cell L-selectin ligand exhibits sulfate-independent binding activity.

L-selectin is a leukocyte cell-surface glycoprotein that mediates adhesive interactions between circulating cells and vascular endothelium. All endothelial ligands of L-selectin characterized to date are glycoproteins that require sulfation for activity and share reactivity with MECA 79, a monoclonal antibody that recognizes a sulfate-dependent epitope involved in L-selectin attachment. We have recently identified by functional assay a glycoprotein L-selectin ligand expressed on the human hematopoietic cell line KG1a. We report here that this ligand is not recognized by MECA 79 and that it retains binding activity after metabolic inhibition of sulfation by chlorate. A native membrane L-selectin ligand exhibiting sulfate-independent function has not been described previously. Identification of this novel ligand on a nonendothelial cell type suggests that structural determinants conferring L-selectin binding may vary in a cell- and tissue-specific manner.

Animals↗
PubMed1997

Free radical oxidation of brain proteins in accelerated senescence and its modulation by N-tert-butyl-alpha-phenylnitrone.

According to the free radical theory of aging, reactive oxygen species cause oxidative damage, proposed to be an underlying factor of the aging process. In the current study, we have used electron paramagnetic resonance spin labeling, measurements of protein carbonyl content, an index of protein oxidation, and determination of the activity of glutamine synthetase (an oxidatively sensitive enzyme) to report that cortical synaptosomal membranes from the senescence accelerated-prone (SAMP8) mouse showed structural characteristics of free radical oxidative stress relative to the senescence accelerated-resistant (SAMR1) mouse. The SAMP8 mouse exhibited a decrease in the relevant EPR parameter consistent with oxidative stress (P < 0.002), a decreased glutamine synthetase activity (P < 0.05), and an increased protein carbonyl content (P < 0.01) compared with these parameters in the SAMR1 mouse. Further, because free radical trapping compounds have been demonstrated to extend maximum life span and improve cognition in SAMP8 mice, we investigated the protective nature of the known free radical scavenger, N-tert-butyl-alpha-phenylnitrone (PBN), on the physical state of cortical synaptosomal membrane proteins. For 14 days, SAMR1 and SAMP8 mice were injected with 30 mg/kg PBN while the controls were injected with the corresponding volume of saline. Characteristic of less oxidized systems, cortical synaptosomal membranes from the PBN-injected SAMP8 mouse exhibited a return toward normal values of the relevant EPR parameter [the M1 = +1 low-field weakly immobilized line/M1 = +1 low-field strongly immobilized line (W/S) ratio of a protein-specific spin label] (P < 0.001) compared with that from saline-injected SAMP8 mice. In SAMR1 mice, in contrast to SAMP8, there was no significant change in the conformation of membrane proteins or protein carbonyl content of cortical synaptosomal membranes from the PBN-injected and saline-injected SAMR1 mice, showing that PBN itself did not induce conformational changes in cortical synaptosomal membrane proteins. The results are discussed with reference to the use of free radical scavengers as potential anti-aging agents.

Aging↗
PubMed1997

The formation of desethyl-piperacillin from piperacillin by human liver S9 in vitro.

Piperacillin (PIPC) has been used as one of the most useful beta-lactam antibiotics over the past 10 years. The metabolism of PIPC has been thoroughly investigated and it has been recognized that PIPC gives few metabolites in laboratory species or humans. Recently, an active metabolite, desethyl-piperacillin (DEt-PIPC), was detected in human plasma and urine after PIPC administration. In the current study, human tissues were obtained from organ donors (n = 3) and subcellular fractions (S9) were prepared. The time course of metabolism by S9 mix from liver, kidney cortex, and kidney medulla was then determined using 0.5 mM PIPC. For comparative purposes, rat liver S9 were also prepared and incubated with PIPC under the same conditions. DEt-PIPC was formed by human liver S9 mix from all three specimens studied, with the rate varying approximately eightfold. No DEt-PIPC was detected in any of the incubations with rat liver S9 mix (n = 3) and kidney S9 mix (n = 3) prepared from either the cortex or medulla. In summary, these data suggest that the formation of the unique human metabolite, DEt-PIPC, can be predicted by in vitro studies with human tissues and that this metabolite is formed predominantly by the liver.

Adult↗
PubMed1997

Acute stroke: usefulness of early CT findings before thrombolytic therapy.

PURPOSE: To determine whether the extent of subtle parenchymal hypoattenuation detected on computed tomographic (CT) scans obtained within 6 hours of ischemic stroke is a factor in predicting patients' response to thrombolytic treatment. MATERIALS AND METHODS: The baseline CT scans of 620 patients, who received either recombinant tissue plasminogen activator (rt-PA) or a placebo, in a double-blind, randomized multicenter trial were prospectively evaluated and assigned to one of three categories according to the extent of parenchymal hypoattenuation: none, 33% or less (small), or more than 33% (large) of the middle cerebral artery territory. The association between the extent of hypoattenuation on the baseline CT scans and the clinical outcome in the placebo-treated and the rt-PA-treated groups after 3 months was analyzed. RESULTS: In 215 patients with a small hypoattenuating area, treatment increased the chance of good outcome. In 336 patients with a normal CT scan and in 52 patients with a large hypoattenuating area, rt-PA had no beneficial effect but increased the risk for fatal brain hemorrhage. CONCLUSION: The response to rt-PA in patients with ischemic stroke can be predicted on the basis of initial CT findings of the extent of parenchymal hypoattenuation in the territory of the middle cerebral artery.

Acute Disease↗
PubMed1996

Prevention of hyperoxia-induced alterations in synaptosomal membrane-associated proteins by N-tert-butyl-alpha-phenylnitrone and 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl (Tempol).

Hyperoxia has been considered a model of free radical reactive oxygen species production in aging and age-related disorders. Previously, we studied the membrane protein alterations that occur during hyperoxia; we found that exposure of young animals to 24 h of hyperoxia provided the greatest degree of oxidation of cortical synaptosomal membrane proteins. We reasoned that free radical oxidation was involved in this protein oxidation. In accordance, in the current study we investigated the protective nature of two known free radical scavengers, N-tert-butyl-alpha-phenylnitrone (PBN) and 4-hydroxy-2,2,6,6-tetramethylpiperidin-1-oxyl (Tempol), against 24-h hyperoxia damage. The three techniques used in this study were electron paramagnetic resonance (EPR) protein-specific spin labeling, assay of the activity of the oxidatively sensitive enzyme glutamine synthetase (GS), and measurement of protein carbonyl content. Before hyperoxia, gerbils received intraperitoneal injections of varying concentrations of either of the two free radical scavengers. After 30 min, the gerbils were exposed to 90-100% O2 for 24 h. For the spin labeling experiments, cortical synaptosomes were isolated from gerbils. The membrane proteins were spin labeled with the thiol-specific label MAL-6 (2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl). As in our earlier study, the EPR spectral parameter of MAL-6-labeled membranes, the W/S ratio, decreased with hyperoxia (p < 0.00001). This effect was lessened significantly with administration of PBN (p < 0.0003) or Tempol (p < 0.00003). For the GS and protein carbonyl assays, cortical proteins were used. The activity of the GS decreased with hyperoxia (p < 0.000005), and this effect likewise was lessened with administration of PBN (p < 0.004) or Tempol (p < 0.002). The protein carbonyl content increased with hyperoxia (p < 0.0002), and there was a protective effect found with Tempol (p < 0.000001). The optimum doses for PBN and Tempol were 20 and 5 mg/kg, respectively. The results are discussed with reference to the use of free radical scavengers as potential antiaging agents.

Animals↗
PubMed1996

The effect of gamma-irradiation on the antibacterial activity of honey.

There is increasing usage of honey as a dressing on infected wounds, burns and ulcers, but there is some concern that there may be a risk of wound botulism from the clostridial spores sometimes found in honey. It is well-established that the antibacterial activity is heat-labile so would be destroyed if honey were sterilized by autoclaving, but the effect of gamma-irradiation on the antibacterial activity of honey is not known. Therefore an investigation was carried out to assess the effect on the antibacterial activity of honey when the honey was subjected to a commercial sterilization procedure using gamma-irradiation (25 kGy). Two honeys with antibacterial activity due to enzymically-generated hydrogen peroxide and three manuka honeys with non-peroxide antibacterial activity were investigated. The honeys were tested against Staphylococcus aureus in an agar well diffusion assay. There was no significant change found in either type of antibacterial activity resulting from this form of sterilization of honey, even when the radiation was doubled (to 50 kGy). Testing of honey seeded with spores of Clostridium perfringens and C. tetani (10000 and 1000 spores g-1 of honey, respectively) showed that 25 kGy of gamma-irradiation was sufficient to achieve sterility.

Honey↗
PubMed1995

Changes of respiratory chain activity in mitochondrial and synaptosomal fractions isolated from the gerbil brain after graded ischaemia.

In this study we have examined (1) the integrated function of the mitochondrial respiratory chain by polarographic measurements and (2) the activities of the respiratory chain complexes I, II-III, and IV as well as the ATP synthase (complex V) in free mitochondria and synaptosomes isolated from gerbil brain, after a 30-min period of graded cerebral ischaemia. These data have been correlated with cerebral blood flow (CBF) values as measured by the hydrogen clearance technique. Integrated functioning of the mitochondrial respiratory chain, using both NAD-linked and FAD-linked substrates, was initially affected at CBF values of approximately 35 ml 100 g-1 min-1, and declined further as the CBF was reduced. The individual mitochondrial respiratory chain complexes, however, showed differences in sensitivity to graded cerebral ischaemia. Complex I activities decreased sharply at blood flows below approximately 30 ml 100 g-1 min-1 (mitochondria and synaptosomes) and complex II-III activities decreased at blood flows below 20 ml 100 g-1 min-1 (mitochondria) and 35-30 ml 100 g-1 min-1 (synaptosomes). Activities declined further as CBF was reduced below these levels. Complex V activity was significantly affected only when the blood flow was reduced below 15-10 ml 100 g-1 min-1 (mitochondria and synaptosomes). In contrast, complex IV activity was unaffected by graded cerebral ischaemia, even at very low CBF levels.

Animals↗
PubMed1995

Effect of reperfusion following cerebral ischaemia on the activity of the mitochondrial respiratory chain in the gerbil brain.

The effect of reperfusion following 30 min of cerebral ischaemia on brain mitochondrial respiratory chain activity has been studied in the gerbil. The state 3 respiration rates with both FAD- and NAD-linked substrates were reduced after ischaemia. After 5 min of reperfusion, state 3 respiration with FAD-linked substrates was restored, but levels of NAD-linked substrates did not return to control values until 30 min of reperfusion. By 120 min of reperfusion state 3 respiration decreased relative to control values with all substrates studied. Measurement of the individual respiratory chain complexes showed that complex I, complex II-III, and complex V activities were reduced after ischaemia. By 5 min of reperfusion complex II-III activity was restored, but the activities of complexes I and V did not return to control values until 30 min of reperfusion. In contrast, complex IV activity was unaffected by ischaemia or 5 and 30 min of reperfusion but was significantly reduced after 120 min of reperfusion, possibly owing to free radical production and lipid peroxidation.

Animals↗
PubMed1994

Identification of ethanolamine in rat and gerbil brain tissue extracts by NMR spectroscopy.

Some NMR resonances which have previously been observed but not identified in mammalian brain tissue extracts have been shown to arise from ethanolamine. This conclusion is drawn from a systematic study of the perchloric acid extracts of rodent brain tissue in which several NMR experiments were used to assign the peaks unambiguously. The extraction procedure used in this work gave samples with highly reproducible spectra, and ethanolamine was observed in all our extract samples. A localized increase in the concentration of ethanolamine was seen in the spectra of extracts produced from a cerebral infarct induced by occlusion of the middle cerebral artery in the rat.

Animals↗
PubMed1994

A sodium magnetic resonance imaging study of acute cerebral ischaemia in the gerbil.

23Na magnetic resonance imaging has been used to investigate sodium changes during and after cerebral ischaemia in a gerbil model. The sodium signal decreased within 4 minutes of the onset of ischaemia, and subsequently increased between 4 and 8 minutes after the onset of reperfusion. These observations may be reflecting the redistribution of tissue sodium resulting from energy failure and recovery.

Animals↗
PubMed1994

Early changes in cerebral sodium distribution following ischaemia monitored by 23Na magnetic resonance imaging.

23Na magnetic resonance imaging has been used in this preliminary study to investigate early changes in brain sodium signal intensity during and after cerebral ischaemia in a gerbil model. The total sodium signal in selected brain regions decreased between 15 and 30% within 4 min of the onset of ischaemia, and then remained constant throughout the ischaemic period. The same pattern was observed in the eyes. On reperfusion, there was no significant change in the sodium signal over the first 4 min, but by 8 min the signal intensity had returned to or passed through control levels in all regions measured, with the exception of the eyes. These observations are consistent with the loss and resynthesis of ATP as seen in this model, and may be reflecting the redistribution of tissue sodium resulting from energy failure and recovery.

Acute Disease↗
PubMed1992

Diffusion-weighted imaging studies of cerebral ischemia in gerbils. Potential relevance to energy failure.

BACKGROUND AND PURPOSE: Diffusion-weighted magnetic resonance imaging has been shown to be particularly suited to the study of the acute phase of cerebral ischemia in animal models. The studies reported in this paper were undertaken to determine whether this technique is sensitive to the known ischemic thresholds for cerebral tissue energy failure and disturbance of membrane ion gradients. METHODS: Diffusion-weighted images of the gerbil brain were acquired under two sets of experimental conditions: as a function of cerebral blood flow after controlled graded occlusion of the common carotid arteries (partial ischemia), as a function of time following complete bilateral carotid artery occlusion (severe global ischemia), and on deocclusion after 60 minutes of ischemia. RESULTS: During partial cerebral ischemia, the diffusion-weighted images remained unchanged until the cerebral blood flow was reduced to 15-20 ml.100 g-1.min-1 and below, when image intensity increased as the cerebral blood flow was lowered further. This is similar to the critical flow threshold for maintenance of tissue high-energy metabolites and ion homeostasis. After the onset of severe global cerebral ischemia, diffusion-weighted image intensity increased gradually after a delay of approximately 2.5 minutes, consistent with complete loss of tissue adenosine triphosphate and with the time course of increase in extracellular potassium. This hyperintensity decreased on deocclusion following 60 minutes of ischemia. CONCLUSIONS: The data suggest that diffusion-weighted imaging is sensitive to the disruption of tissue energy metabolism or a consequence of this disruption. This raises the possibility of imaging energy failure noninvasively. In humans, this could have potential in visualizing brain regions where energy metabolism is impaired, particularly during the acute phase following stroke.

Animals↗
PubMed1991

A survey of the antibacterial activity of some New Zealand honeys.

To assess the variation in antibacterial activity of honey a survey was carried out on 345 samples of unpasteurized honey obtained from commercial apiarists throughout New Zealand. Most of the honeys were considered to be monofloral, from 26 different floral sources. The honeys were tested against Staphylococcus aureus in an agar well diffusion assay, with reference to phenol as a standard. Antibacterial activity was found to range from the equivalent of less than 2% (w/v) phenol to 58% (w/v) phenol, with a median of 13.6 and a standard deviation of 12.5. Neither the age of the honey samples nor whether they had been processed by the apiarist was associated with lower activity. However, the difference between floral sources in the antibacterial activity was very highly significant. Kanuka (Kunzea ericoides (A. Rich.) J. Thompson. Family: Myrtaceae), manuka (Leptospermum scoparium J. R. et G. Forst. Family: Myrtaceae), ling heather (Calluna vulgaris (L.) Hull. Family: Ericaceae) and kamahi (Weinmannia racemosa Linn. f. Family: Cunoniaceae) were shown to be sources likely to give honey with high antibacterial activity. When antibacterial activity was assayed with catalase added to remove hydrogen peroxide, most of the honeys showed no detectable antibacterial activity. Only manuka and vipers bugloss (Echium vulgare L. Family: Boraginaceae) honeys showed this type of activity in a significant proportion of the samples. The high antibacterial activity of manuka honey was in many cases due entirely to this non-peroxide component.

Anti-Bacterial Agents↗
PubMed1990

Residues flanking the COOH-terminal C-region of a model eukaryotic signal peptide influence the site of its cleavage by signal peptidase and the extent of coupling of its co-translational translocation and proteolytic processing in vitro.

The polar, COOH-terminal c-region of signal peptides has been considered to be most important for influencing the efficiency and fidelity of signal peptidase cleavage while the hydrophobic core or h-region appears indispensable for initiating translocation. To identify structural features of residues flanking the c-region that influence the fidelity and efficiency of signal peptidase cleavage as well as co-translational translocation, we introduced six amino acid substitutions into the COOH terminus of the hydrophobic core and seven substitutions at the NH2 terminus of the mature region (the +1 position) of a model eukaryotic preprotein-human pre(delta pro)apoA-II. This preprotein contains several potential sites for signal peptidase cleavage. The functional consequences of these mutations were assayed using an in vitro co-translational translocation/processing system and by post-translational cleavage with purified, detergent-solubilized, hen oviduct signal peptidase. The efficiency of translocation could be correlated with the hydrophobic character of the residue introduced at the COOH terminus of the h-region. Some h/c boundary mutants underwent co-translational translocation across the microsomal membrane with only minimal cleavage yet they were cleaved post-translationally by hen oviduct signal peptidase more efficiently than other mutants which exhibited a high degree of coupling of co-translational translocation and cleavage. These data suggest that features at the COOH terminus of the h-domain can influence "presentation" of the cleavage site to signal peptidase. The +1 residue substitutions had minor effects on the extent of co-translational translocation and processing. However, these +1, as well as h/c boundary mutations, had dramatic effects on the site of cleavage chosen by signal peptidase, indicating that residues flanking the c-region of this prototypic eukaryotic signal peptide can affect the fidelity of its proteolytic processing. The site(s) selected by canine microsomal and purified hen oviduct signal peptidase were very similar, suggesting that "intrinsic" structural features of this prepeptide can influence the selectivity of eukaryotic signal peptidase cleavage, independent of the microsomal membrane and associated translocation apparatus.

Amino Acid Sequence↗
PubMed1990

Restoration of energy metabolism and resolution of oedema following profound ischaemia.

Cerebral ischaemia was produced in 2 groups of gerbils by occlusion of the common carotid arteries for 30 minutes, resulting in cerebral oedema. In group 1 cerebral oedema was measured by specific gravity microgravimetry, and in group 2 brain metabolism and blood flow were measured by 31P and 1H NMR spectroscopy and hydrogen clearance respectively. In group 1 the brain water content did not return to control levels by 180 minutes of reperfusion. Energy metabolism, determined by 31P NMR spectroscopy returned to control by 12 minutes, intracellular pH (pHi) by 20 minutes, and lactate, determined by 1H NMR spectroscopy, by 50 minutes. There was a lag of about 10 minutes before lactate began to be cleared from the brain. We suggest that while pHi is low, Na+/H+ exchange will negate the Na+ extrusion driven by the Na+/K+ ATPase. When pHi approaches normal there will be a net extrusion of Na+, taking osmotic water with it, and presumably with passive washout of lactate. This may be the cause of the initial delay in lactate clearance.

Animals↗