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R M Allen

Publications and source records attributed to R M Allen.

At least 19 recordsLinked to original sources
PubMed2001

A district general hospital's method of post-operative infection surveillance including post-discharge follow-up, developed over a five-year period.

We report on a post-operative infection surveillance system which includes post-discharge follow-up, developed over five years in a district general hospital in the West Midlands, UK. The methods used for following up 667 patients undergoing one of five representative surgical procedures are described. Emergency, elective and day-case procedures are included. A combination of healthcare worker questionnaire, telephone calls and patient questionnaire gave a follow-up rate of 92.7%. The system took infection control staff an average of 40 min per patient (30 min inpatient assessment, 10 min post-discharge). Almost half (48%) of surgical site infections were diagnosed after discharge from hospital. The system worked equally well when conducted as part of the UK Nosocomial Infection National Surveillance Scheme (NINSS), or as in-house projects. It is likely that the system could be used in other areas with similar population characteristics and support from local general practitioners working in the community.

Aftercare↗
PubMed2001

N-methyl-D-aspartate receptor antagonists potentiate the antinociceptive effects of morphine in squirrel monkeys.

Data from rodent antinociception models indicate that N-methyl-D-aspartate (NMDA) receptor antagonists do not produce antinociception alone or potentiate morphine antinociception, but do attenuate the development of morphine tolerance. This study examined the antinociceptive effects of the noncompetitive NMDA receptor antagonist dizocilpine, the competitive NMDA receptor antagonist (-)-6-phosphonomethyl-decahydroisoquinoline-3-carboxylic acid (LY235959), and the glycine-site antagonist (+)-(1-hydroxy-3-aminopyrrolidine-2-one) [(+)-HA-966], alone and in combination with morphine in a squirrel monkey titration procedure. In this procedure, shock (delivered to the tail) increased in intensity every 15 s from 0.01 to 2.0 mA in 30 increments. Five lever presses during any given 15-s shock period produced a 15-s shock-free period after which shock resumed at the next lower intensity. Morphine (0.3-3.0 mg/kg i.m.) dose-dependently increased the intensity below which monkeys maintained shock 50% of the time (median shock level; MSL). In contrast, dizocilpine (0.003-0.1 mg/kg i.m.) produced only modest increases in MSL in some monkeys (three of five) at the highest dose tested. Neither LY235959 (0.1-1.0 mg/kg i.m.) or (+)-HA-966 (10-56 mg/kg i.m.) increased MSL in any monkey tested. Dizocilpine, LY235959, and (+)-HA-966, when administered in combination with doses of morphine (1.0 mg/kg, 1.7 mg/kg) that either produced no antinociception or produced very little antinociception, were all found to dose-dependently potentiate the antinociceptive effect of morphine. Importantly, although these NMDA antagonists in combination with morphine produced marked increases in MSL, these combinations did not alter response rate, demonstrating that the potentiation was not due to nonspecific motor effects.

Analgesics↗
PubMed2000

Role of morphine maintenance dose in the development of tolerance and its attenuation by an NMDA receptor antagonist.

RATIONALE: Current research shows that N-methyl-d-aspartate (NMDA) receptor antagonists attenuate the development of morphine tolerance in rodent antinociceptive assays. OBJECTIVE: The purpose of this study was to determine the role of morphine maintenance dose in the attenuation of morphine tolerance by the competitive NMDA receptor antagonist, LY235959. METHODS: A rat warm-water tail-withdrawal procedure was used to measure the antinociceptive effects of morphine and LY235959. In this procedure, the distal 8 cm of each rat's tail is immersed in 40 degrees (non-noxious) and 55 degrees C (noxious) water, and the latency to remove the tail is recorded. RESULTS: Morphine (0.3-10 mg/kg, SC) produced dose-dependent increases in tail-withdrawal latencies from the 55 degrees C water. Following determination of the morphine dose-effect curves, rats were administered chronically one of three doses of morphine (10, 20, or 40 mg/kg) either alone or in combination with LY235959 (1.0, 3.0, or 5.6 mg/kg, SC) twice daily for 7 days. Chronic administration of 10, 20, and 40 mg/kg morphine produced rightward shifts in the morphine dose-effect curves of approximately 3-, 6-, and 12-fold, respectively. When LY235959 (1.0-5.6 mg/kg) was co-administered with 10 mg/kg morphine, the development of morphine tolerance was attenuated in a dose-dependent manner, with complete prevention observed following 3.0 mg/kg LY235959. LY235959 (1.0, 3.0 mg/kg) also attenuated the development of tolerance to 20 and 40 mg/kg morphine; however, tolerance was not completely prevented. Administering 3.0 mg/kg LY235959 along with 20 and 40 mg/kg morphine was functionally equivalent to treating rats with half the amount of morphine. CONCLUSION: These data suggest that the maintenance dose of morphine, and thus the magnitude of tolerance, can determine the effectiveness of an NMDA receptor antagonist to attenuate morphine tolerance.

Animals↗
PubMed2000

A reliable and valid asthma general knowledge questionnaire useful in the training of asthma educators.

Using the responses of 115 adults attending an asthma educator training course, the Asthma General Knowledge Questionnaire for Adults (AGKQA) was found to be an acceptably valid and reliable measure for assessing knowledge related to the management of asthma by adults. Content and face validity: expert assessors considered the AGKQA to be a relevant and plausible test of the asthma general knowledge content of the programme. Criterion-related validity: the pretraining scores of educators were significantly higher (P < 0.001) than those of adults with no experience of asthma; total scores for the AGKQA and an asthma knowledge questionnaire developed for parents of children with asthma correlated strongly, 0.72. Test-retest reproducibility: the Spearman rank correlation for the test-retest score was 0.72 (P < 0.02), kappas for concordance of item responses were moderate to very good for two thirds of the items. Internal consistency for the total scale was also acceptable, KR20 0.66.

Adolescent↗
PubMed2000

Attenuation of mu-opioid tolerance and cross-tolerance by the competitive N-methyl-D-aspartate receptor antagonist LY235959 is related to tolerance and cross-tolerance magnitude.

Although NMDA receptor antagonists attenuate the development of morphine tolerance, it is not clear whether NMDA receptor antagonists also prevent tolerance and cross-tolerance to other mu-opioid agonists and, if so, whether prevention is related to the efficacy of the agonist used to examine tolerance. A rat tail-withdrawal procedure was used to test the antinociceptive effects of the mu-opioids etorphine, morphine, and dezocine before and after twice-daily subcutaneous injections with either 0. 003 mg/kg etorphine, 10 mg/kg morphine, or 3.0 mg/kg dezocine, each administered alone or in combination with 3.0 mg/kg of the competitive NMDA antagonist LY235959. After chronic etorphine, the etorphine, morphine, and dezocine curves were shifted rightward 1.0-, 2.2-, and 3.4-fold, respectively. LY235959 prevented cross-tolerance to morphine and dezocine. After chronic morphine, the etorphine and morphine curves were shifted rightward 2.5- and 2. 9-fold, respectively, and the dezocine curve was flattened. LY235959 prevented morphine tolerance and cross-tolerance to etorphine and reduced the magnitude of cross-tolerance to dezocine. After chronic dezocine, the etorphine, morphine, and dezocine curves were shifted rightward 4.1-, 3.5-, and 9.6-fold, respectively. LY235959 did not prevent but reduced the magnitude of tolerance and cross-tolerance. In a separate experiment, the following rank order of efficacy was determined from the magnitudes of rightward shift in each dose-effect curve after administration of 1.0 mg/kg of the irreversible antagonist clocinnamox: etorphine > morphine > dezocine. These data show that differences in tolerance magnitude are related to opioid efficacy and that attenuation of mu-opioid tolerance and cross-tolerance by LY235959 depends upon the magnitude of opioid tolerance.

Animals↗
PubMed1999

Incorporation of molybdenum into the iron-molybdenum cofactor of nitrogenase.

The biosynthesis of the iron-molybdenum cofactor (FeMo-co) of dinitrogenase was investigated using 99Mo to follow the incorporation of Mo into precursors. 99Mo label accumulates on dinitrogenase only when all known components of the FeMo-co synthesis system, NifH, NifNE, NifB-cofactor, homocitrate, MgATP, and reductant, are present. Furthermore, 99Mo label accumulates only on the gamma protein, which has been shown to serve as a chaperone/insertase for the maturation of apodinitrogenase when all known components are present. It appears that only completed FeMo-co can accumulate on the gamma protein. Very little FeMo-co synthesis was observed when all known components are used in purified forms, indicating that additional factors are required for optimal FeMo-co synthesis. 99Mo did not accumulate on NifNE under any conditions tested, suggesting that Mo enters the pathway at some other step, although it remains possible that a Mo-containing precursor of FeMo-co that is not sufficiently stable to persist during gel electrophoresis occurs but is not observed. 99Mo accumulates on several unidentified species, which may be the additional components required for FeMo-co synthesis. The molybdenum storage protein was observed and the accumulation of 99Mo on this protein required nucleotide.

Adenosine Triphosphate↗
PubMed1999

The competitive NMDA receptor antagonist LY235959 modulates the progression of morphine tolerance in rats.

A rat warm-water tail-withdrawal procedure was used to examine the effects of chronic administration of the competitive NMDA receptor antagonist LY235959 in morphine tolerant rats. Morphine dose-dependently increased tail-withdrawal latencies from 55 degree C water. When morphine (10 mg/kg) was administered twice-daily for 7 days, the morphine dose-effect curves shifted 0.3-0.5 log unit to the right. When morphine was administered for an additional 7 days, the morphine dose-effect curve shifted 0.4 log unit further to the right. Co-administration of LY235959 (1, 3, 10 mg/kg) along with morphine prevented the development of tolerance observed during the second week of chronic morphine administration. Although the highest dose of LY235959 (10 mg/kg) partially reversed tolerance in five of seven rats, tolerance was not reversed by lower doses of LY235959. These data suggest that NMDA receptor antagonists may effectively prevent the progressive development of morphine tolerance at doses that are not sufficient to reverse pre-established morphine tolerance.

Analgesics, Opioid↗
PubMed1999

Effects of neuropeptide Y on the discriminative stimulus and antinociceptive properties of morphine.

Previous research indicates that opioid receptor blockade diminishes the effects of neuropeptide Y (NPY) on feeding and memory. Conversely, NPY attenuates naloxone-precipitated morphine withdrawal. The present study evaluated the effects of NPY on the discriminative stimulus and antinociceptive effects produced by the prototypical mu opioid, morphine. Rats were trained to discriminate 5.6 mg/kg morphine (IP) from saline using a standard two-lever, food-reinforced, drug discrimination procedure. Across a range of doses (3.0, 5.0, and 10 microg), intracerebroventricular (ICV) injection of NPY failed to substitute for, antagonize, or potentiate the discriminative stimulus effects of morphine. A warm-water tail-withdrawal procedure was used to examine the antinociceptive effects of morphine and NPY, alone and in combination. NPY (3.0 and 10 microg, ICV) failed to alter tail-withdrawal latencies from 52 degrees and 56 degrees C water, whereas morphine (1.0-30 mg/kg, IP) produced a dose-related increase in latencies at both water temperatures. A 10-microg dose of NPY also failed to alter the antinociceptive effects of morphine. This study does not support the idea that the discriminative stimulus and antinociceptive effects of morphine are dependent on an NPYergic pathway.

Analgesics, Opioid↗
PubMed1999

Requirement of NifX and other nif proteins for in vitro biosynthesis of the iron-molybdenum cofactor of nitrogenase.

The iron-molybdenum cofactor (FeMo-co) of nitrogenase contains molybdenum, iron, sulfur, and homocitrate in a ratio of 1:7:9:1. In vitro synthesis of FeMo-co has been established, and the reaction requires an ATP-regenerating system, dithionite, molybdate, homocitrate, and at least NifB-co (the metabolic product of NifB), NifNE, and dinitrogenase reductase (NifH). The typical in vitro FeMo-co synthesis reaction involves mixing extracts from two different mutant strains of Azotobacter vinelandii defective in the biosynthesis of cofactor or an extract of a mutant strain complemented with the purified missing component. Surprisingly, the in vitro synthesis of FeMo-co with only purified components failed to generate significant FeMo-co, suggesting the requirement for one or more other components. Complementation of these assays with extracts of various mutant strains demonstrated that NifX has a role in synthesis of FeMo-co. In vitro synthesis of FeMo-co with purified components is stimulated approximately threefold by purified NifX. Complementation of these assays with extracts of A. vinelandii DJ42. 48 (DeltanifENX DeltavnfE) results in a 12- to 15-fold stimulation of in vitro FeMo-co synthesis activity. These data also demonstrate that apart from the NifX some other component(s) is required for the cofactor synthesis. The in vitro synthesis of FeMo-co with purified components has allowed the detection, purification, and identification of an additional component(s) required for the synthesis of cofactor.

Azotobacter vinelandii↗
PubMed1998

The validity and reliability of an asthma knowledge questionnaire used in the evaluation of a group asthma education self-management program for adults with asthma.

This paper reports aspects of the validity and reliability of an asthma general knowledge questionnaire for adults (AGKQA), developed as one of the outcome measures for a randomized controlled effectiveness trial of an asthma education program for adults with asthma. It also illustrates how study data can provide a valuable, generally neglected opportunity to assess the validity and reliability of a measure where resources are not available for the extensive investigation of these properties prior to administration of the measure in a study. The AGKQA was demonstrated to have good content and face validity. Construct validity assessed using the principal components method for factor analysis suggested the scale was unidimensional. Criterion-related validity, assessed using the contrasted groups method, demonstrated a significant difference (p < 0.0001) in total score and for 68% of item responses for the adults with and without direct experience of asthma. The Kuder-Richardson 20 reliability coefficient for internal consistency calculated using responses at baseline, immediately, and 12 months post-intervention were, respectively, 0.56, 0.80, and 0.75, indicating excellent reliability. The AGKQA is an acceptably valid and reliable measure for the assessment of program content mastery that it was designed to test.

Adolescent↗
PubMed1998

Clocinnamox antagonism of the antinociceptive effects of mu opioids in squirrel monkeys.

The opioid agonists morphine, etorphine, buprenorphine and U50,488 were examined alone and in combination with the insurmountable opioid antagonist clocinnamox (C-CAM) in squirrel monkeys responding under a schedule of shock titration. In this procedure, shock intensity increased every 15 sec from 0.01 to 2.0 mA in 30 increments. Five lever presses during any given 15-sec shock period produced a 15-sec timeout, after which shock resumed at the next lower intensity. When given alone, each of these agonists increased the median intensity at which the monkeys maintained shock [median shock level (MSL)]. At the highest dose examined alone, each agonist produced maximal increases in MSL and, except buprenorphine, decreased response rates. C-CAM dose-dependently antagonized the effects of morphine, etorphine and buprenorphine on MSL. In the presence of the higher C-CAM doses, etorphine, morphine and buprenorphine did not produce maximal effects on MSL. The effects of U50,488 were not systematically altered when tested in combination with the highest C-CAM dose. In general, C-CAM was more potent and the duration of antagonism was slightly longer against buprenorphine than against morphine and etorphine. Quantitative analysis of these data according to an extended model of yielded the following apparent affinity and efficacy estimates, respectively: etorphine (0. 085 mg/kg, 117); morphine (49 mg/kg, 24) and buprenorphine (0.62 mg/kg, 7.1). Determination of the individual q values over time indicated that the receptor population recovers more quickly after C-CAM antagonism of etorphine than from C-CAM antagonism of either morphine or buprenorphine. These data suggest that C-CAM functions as a long-lasting antagonist of mu opioid agonist actions in a shock titration procedure and yields estimates of relative intrinsic efficacy with the rank order of etorphine > morphine > buprenorphine.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗
PubMed1997

[2Fe-2S] to [4Fe-4S] cluster conversion in Escherichia coli biotin synthase.

The type and properties of the Fe-S cluster in recombinant Escherichia coli biotin synthase have been investigated in as-prepared and dithionite-reduced samples using the combination of UV-visible absorption and variable-temperature magnetic circular dichroism (VTMCD), EPR, and resonance Raman spectroscopies. The results confirm the presence of one S = 0 [2Fe-2S]2+ cluster in each subunit of the homodimer in aerobically purified samples, and the Fe-S stretching frequencies suggest incomplete cysteinyl-S coordination. However, absorption and resonance Raman studies show that anaerobic reduction with dithionite in the presence of 60% (v/v) ethylene glycol or glycerol results in near-stoichiometric conversion of two [2Fe-2S]2+ clusters to form one S = 0 [4Fe-4S]2+ cluster with complete cysteinyl-S coordination. The stoichiometry and ability to effect reductive cluster conversion without the addition of iron or sulfide suggest that the [4Fe-4S]2+ cluster is formed at the subunit interface via reductive dimerization of [2Fe-2S]2+ clusters. EPR and VTMCD studies indicate that more than 50% of the Fe is present as [4Fe-4S]+ clusters in samples treated with 60% (v/v) glycerol after prolonged dithionite reduction. The [4Fe-4S]+ cluster exists as a mixed spin system with S = 1/2 (g = 2. 044, 1.944, 1.914) and S = 3/2 (g = 5.6 resonance) ground states. Subunit-bridging [4Fe-4S]2+,+ clusters, that can undergo oxidative degradation to [2Fe-2S]2+ clusters during purification, are proposed to be a common feature of Fe-S enzymes that require S-adenosylmethionine and function by radical mechanisms involving the homolytic cleavage of C-H or C-C bonds, i.e., biotin synthase, anaerobic ribonucleotide reductase, pyruvate formate lyase, lysine 2, 3-aminomutase, and lipoic acid synthase. The most likely role for the [4Fe-4S]2+,+ cluster lies in initiating the radical mechanism by directly or indirectly facilitating reductive one-electron cleavage of S-adenosylmethionine to form methionine and the 5'-deoxyadenosyl radical. It is further suggested that oxidative cluster conversion to [2Fe-2S]2+ clusters may play a physiological role in these radical enzymes, by providing a method of regulating enzyme activity in response to oxidative stress, without irreversible cluster degradation.

Dithionite↗
PubMed1997

In vitro synthesis of the iron-molybdenum cofactor and maturation of the nif-encoded apodinitrogenase. Effect of substitution of VNFH for NIFH.

NIFH (the nifH gene product) has several functions in the nitrogenase enzyme system. In addition to reducing dinitrogenase during nitrogenase turnover, NIFH functions in the biosynthesis of the iron-molybdenum cofactor (FeMo-co), and in the processing of alpha2beta2 apodinitrogenase 1 (a catalytically inactive form of dinitrogenase 1 that lacks the FeMo-co) to the FeMo-co-activatable alpha2beta2gamma2 form. The molybdenum-independent nitrogenase 2 (vnf-encoded) has a distinct dinitrogenase reductase protein, VNFH. We investigated the ability of VNFH to function in the in vitro biosynthesis of FeMo-co and in the maturation of apodinitrogenase 1. VNFH can replace NIFH in both the biosynthesis of FeMo-co and in the maturation of apodinitrogenase 1. These results suggest that the dinitrogenase reductase proteins do not specify the heterometal incorporated into the cofactors of the respective nitrogenase enzymes. The specificity for the incorporation of molybdenum into FeMo-co was also examined using the in vitro FeMo-co synthesis assay system.

Apoproteins↗
PubMed1996

Purification and characterization of the vnf-encoded apodinitrogenase from Azotobacter vinelandii.

The vnf-encoded apodinitrogenase (apodinitrogenase 2) has been purified from Azotobacter vinelandii strain CA117.30 (DeltanifKDB), and is an alpha2beta2delta2 hexamer. Apodinitrogenase 2 can be activated in vitro by the addition of the iron-vanadium cofactor (FeV-co) to form holodinitrogenase 2, which functions in C2H2, H+, and N2 reduction. Under certain conditions, the alpha2beta2delta2 hexamer dissociates to yield the free delta subunit (the VNFG protein) and a form of apodinitrogenase 2 that exhibits no C2H2, H+, or N2 reduction activities in the in vitro FeV-co activation assay; however, these activities can be restored upon addition of VNFG to the FeV-co activation assay system. No other vnf-, nif-, or non-nif-encoded proteins were able to replace the function of VNFG in the in vitro processing of alpha2beta2 apodinitrogenase 2 (in the presence of FeV-co) to a form capable of substrate reduction. Apodinitrogenase 2 is also activable in vitro by the iron-molybdenum cofactor to form a hybrid enzyme with unique properties, most notably the inability to reduce N2 and insensitivity to CO inhibition of C2H2 reduction.

Azotobacter vinelandii↗
PubMed1996

The requirement of reductant for in vitro biosynthesis of the iron-molybdenum cofactor of nitrogenase.

A source of reductant is routinely added to the in vitro iron-molybdenum cofactor (FeMo-co) synthesis assay, although a requirement for reductant has not been established. This report demonstrates that the addition of reductant to the in vitro FeMo-co synthesis system is not required when Azotobacter vinelandii cell-free extract is prepared in buffer that lacks added reductant. The addition of reductant is required, however, if the A. vinelandii cell-free extract is chemically oxidized prior to addition to the assay. These results might suggest that extracts of A. vinelandii contain a physiological source of reductant that functions in the in vitro synthesis of FeMo-co. It is possible that the proteins required for FeMo-co biosynthesis (e.g. NIFNE and dinitrogenase reductase) are at the appropriate redox state to function in the in vitro reaction in the extract that is free of added reductant but not in the chemically oxidized extract. It is also possible that dinitrogenase reductase and/or NIFNE (both Fe-S proteins required for FeMo-co synthesis) might catalyze the reductant-dependent reaction for FeMo-co synthesis. Dithionite, Ti(III) citrate, and NADH are able to serve as the source of reductant for in vitro FeMo-co biosynthesis.

Azotobacter vinelandii↗
PubMed1995

Incorporation of iron and sulfur from NifB cofactor into the iron-molybdenum cofactor of dinitrogenase.

NifB-co is an iron- and sulfur-containing precursor to the iron-molybdenum cofactor (FeMo-co) of dinitrogenase. The synthesis of NifB-co requires at least the product of the nifB gene. Incorporation of 55Fe and 35S from NifB-co into FeMo-co was observed only when all components of the in vitro FeMo-co synthesis system were present. Incorporation of iron and sulfur from NifB-co into dinitrogenase was not observed in control experiments in which the apodinitrogenase (lacking FeMo-co) was initially activated with purified, unlabeled FeMo-co or in assays where NifB-co was oxygen-inactivated prior to addition to the synthesis system. These data clearly demonstrate that iron and sulfur from active NifB-co are specifically incorporated into FeMo-co of dinitrogenase and provide direct biochemical identification of an iron-sulfur precursor of FeMo-co. Under different in vitro FeMo-co synthesis conditions, iron and sulfur from NifB-co were associated with at least two other proteins (NIFNE and gamma) that are involved in the formation of active dinitrogenase. The results presented here suggest that multiple FeMo-co processing steps might occur on NIFNE and that FeMo-co synthesis is most likely completed prior to the association of FeMo-co with gamma.

Azotobacter vinelandii↗