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Biomedical subjects

D Cooke

Publications and source records attributed to D Cooke.

At least 37 records · Page 2Linked to original sources
PubMed1994

Use of ion-exchange and hydrophobic-interaction chromatography for the rapid purification of lysozyme-estrone glucuronide conjugates.

Estrone glucuronide conjugates of hen egg white lysozyme were prepared by both the mixed-anhydride and active-ester coupling procedures. Both methods gave good yields of conjugate but the active-ester procedure gave a more diverse range of products consistent with a greater acylating ability. Unreacted lysozyme which was present in all cases was removed by a combination of cation-exchange chromatography on a Pharmacia Mono-S column and hydrophobic-interaction chromatography on an Alkyl Superose column. The conjugate families were more hydrophobic than native lysozyme. The chromatographic behaviour of the reaction mixtures on Mono S columns under non-denaturing conditions was complex as a result of hydrophobic effects and only at pH values above 7.0 did the conjugates elute in the order of their overall charges. At pH values below 6.0 the conjugates, although less charged than lysozyme, eluted last on salt gradients. In contrast when denaturing 7 M urea buffers were used the conjugates eluted in the order of their electrostatic charges and reproducible patterns were obtained which served as an excellent analytical system for lysozyme-steroid glucuronide conjugates. The purified conjugate material from the active-ester reaction gave over 90% inhibition of the lytic activity in the presence of an estrone glucuronide antibody. When used in a homogeneous enzyme immunoassay system the levels of urinary estrone glucuronide encountered in a normal menstrual cycle were easily measured.

Chemical Phenomena↗
PubMed1994

Role of residue Lys315 in the mechanism of action of the Enterobacter cloacae 908R beta-lactamase.

The role of the highly conserved Lys315 residue in the catalytic mechanism of a class C beta-lactamase has been probed by site-directed mutagenesis. Lys315 has been replaced by a histidine in the Enterobacter cloacae 908R beta-lactamase, thus introducing a tritratable group to probe the role of the positive charge, and by a glutamine. The effects of these mutations have been studied on the kinetics of penicillin G and cephalothin turnover and on the pre-steady-state kinetics with carbenicillin at different pH. Results showed that substrate binding was not impaired by the mutations, so that an interaction with the substrate-free carboxylate in the Henri-Michaelis complex could be ruled out. Lys315 must have a catalytic role as shown by the decreased acylation and deacylation rates observed with the mutant enzymes. The mutants exhibited a lower activity at acidic pH, and this observation could be correlated with a decreased affinity for (3-aminophenyl)boronate, a compound devoid of free carboxylate which binds to the active site and forms an adduct mimicking the tetrahedral intermediate. This suggested that Lys315 was somehow involved in accelerating the nucleophilic substitutions along the reaction pathway. The study was extended to modified substrates where the free carboxylate had been esterified. Neither acylation nor deacylation seemed severely impaired with these compounds, showing that the interaction between the enzyme and the substrate-free carboxylate did not play a major role in catalysis.

Amino Acid Sequence↗
PubMed1993

Assessing the diagnostic value of an ECG containing leads V4R, V8, and V9: the 15-lead ECG.

STUDY OBJECTIVES: To assess sensitivity, specificity, and odds ratios of ECG findings on leads V4R, V8, and V9 for acute myocardial infarction. DESIGN: Prospective, two-stage cohort study. SETTING: A 660-bed university-affiliated community hospital. TYPE OF PARTICIPANTS: One hundred forty-nine admitted patients with suspected myocardial infarction or unstable angina. INTERVENTIONS: Standard 12-lead ECG followed immediately by V4R, V8, and V9. MEASUREMENTS: Initial ECG findings of ST-segment displacement, Q waves, T-wave inversion, and eligibility for thrombolytic therapy. RESULTS: Major abnormalities (ST-segment deviation, T-wave inversion, Q waves) were found on the extra three leads in 28.9% (43 of 149) of patients. Sensitivity of ST-segment elevation for acute myocardial infarction on 12 versus 15 leads increased from 47.1% to 58.8%, respectively, with no decrease in specificity. McNemar's pair-matched analysis for ST-segment elevation on myocardial infarction subgroup showed an association of ST elevation with the 15-lead ECG (P < .05). An eightfold increase in the odds of detecting ST elevation was found (90% confidence interval, 1.42 to 14.58); 22% of patients negative for ST elevation on 12 leads were positive on 15 leads. Analysis of ECG criteria for thrombolytic therapy presenting uniquely on extra leads showed an increased sensitivity from 35.3% to 44.1% on 12 versus 15 leads, respectively; there was a sixfold increase in the odds of meeting ECG thrombolytic therapy criteria (90% confidence interval, 0.34 to 11.66); 13.5% of patients not meeting criteria on 12 leads did so on 15 leads. CONCLUSION: The 15-lead ECG provides increased sensitivity and odds of detecting ST-segment elevation in acute myocardial infarction patients with no loss of specificity; its use may expand the selection of thrombolytic therapy candidates and provide a fuller ECG description of the extent of myocardial injury and necrosis.

Aged↗
PubMed1992

Protease composition of exocytosed human skin mast cell protease-proteoglycan complexes. Tryptase resides in a complex distinct from chymase and carboxypeptidase.

We characterized the release and the protease composition of high m.w. complexes released from dispersed human skin mast cells, under conditions that did not disrupt the binding of proteases to proteoglycan. The net percent release ratio of tryptase to histamine, after anti-IgE and calcium ionophore A23187 stimulation was higher than those for chymase or carboxypeptidase. This was explained by the greater cell association of carboxypeptidase and chymase, compared with tryptase, after mast cell degranulation and/or differential cosedimentation of the proteases with mast cells, because treatment of activated mast cells with 1 M NaCl increased the release ratios of chymase and carboxypeptidase more than that of tryptase. Tryptase, after release, was stable in 0.12 M NaCl and had a molecular mass of approximately 200 to 250 kDa, suggesting that it was bound to proteoglycan. We demonstrated that complexes containing chymase and carboxypeptidase were separable from tryptase-containing complexes by gel filtration and by affinity chromatography. First, on fast protein liquid chromatography, released tryptase filtered at a molecular mass of approximately 200 to 250 kDa, compared with chymase and carboxypeptidase at 400 to 560 kDa. Second, by using affinity chromatography with immobilized antitryptase mAb in 0.15 M NaCl, carboxypeptidase and chymase activities were recovered primarily in the effluent and washes of an antitryptase antibody affinity column and cofiltered at 400 to 560 kDa. Tryptase was recovered only in the eluate. Finally, by using potato tuber carboxypeptidase inhibitor-Sepharose affinity chromatography, tryptase activity was found primarily in the effluent and washes, filtered at a molecular mass of 200 kDa on fast protein liquid chromatography, and was stable in 0.12 M NaCl buffer at 37 degrees C. Carboxypeptidase and chymase activities were found primarily in the eluate. These findings suggest that tryptase and carboxypeptidase/chymase reside in distinct macromolecular complexes. Separate complexes containing these proteases may help explain previous ultrastructural observations in which the distributions of chymase and tryptase within a single granule did not always coincide.

Carboxypeptidases↗
PubMed1991

Effects of conventional sucrose-based, fructose-based and isomalt-based chocolates on postprandial metabolism in non-insulin-dependent diabetics.

Milk chocolate is rich in both sucrose and fat, and is therefore considered unsuitable for diabetics. Nevertheless there is little information on the metabolic effects of conventional chocolate or specialized formulations with reduced sucrose content. In the present study six male non-insulin-dependent diabetes mellitus patients (age range 35-60 years; body-mass index less than 28) consumed test meals of chocolate (75 g) on three separate occasions. The control chocolate contained sucrose (45.5% w/w); the test chocolates contained either fructose (45.5% w/w) or isomalt (45.1% w/w). The latter is a sweet disaccharide alcohol which has no glycaemic effect when consumed as a pure compound. Venous blood samples were obtained at 30 min intervals for 5 h, and analysed for glucose, insulin, lactate and triglycerides. All three chocolates provoked a sustained rise in blood glucose, which reached a maximum at 90 min after ingestion and returned to baseline values by 5 h. The highest blood glucose levels occurred after conventional chocolate, and differences were statistically significant at 60 and 90 min (P less than 0.05). The area under the glycaemic curve for isomalt chocolate was 36% smaller than that for conventional chocolate (P less than 0.05), and there were differences in insulin and lactate levels, consistent with the lower glycaemic effect. The glycaemic response to the fructose-based chocolate was also lower than that to control chocolate but the difference was not significant. All three chocolates led to a similar sustained rise in serum triglyceride levels. Isomalt appears to be a palatable alternative sweetener capable of reducing the glycaemic effect of diabetic confectionary.

Adult↗
PubMed1991

Interactions of immunoglobulins and chondrocytes.

Our study relates to damage of articular cartilage by deposition of immune complexes. Addition of IgG to cultured chondrocytes affected their metabolism--boosting metalloprotease synthesis, generating superoxide anion (O2-), and reducing the synthesis of sulfated glycosaminoglycans (S-GAG). Heat aggregation of the IgG enhanced its effect on O2- and S-GAG. Some interleukin 1 beta (38 pg/ml) was detected by ELISA after exposure to heat aggregated IgG. There was no cytotoxic effect due to IgG alone, but addition of complement (5% bovine serum) caused significant loss of intracellular 51Cr, which was further increased by addition of heat aggregated IgG. Similar responses of chondrocytes in vivo might account for immune complex mediated damage of cartilage.

Animals↗
PubMed1989

Modification and calibration of the Bigliano-Webb Tonometer for improved accuracy of tonometry in rabbits.

The Bigliano-Webb Tonometer (Pneumatonograph) was calibrated for adult and young rabbit eyes using open stopcock and closed stopcock methods. The performance of an old and new instrument were compared illustrating the importance of recent calibration of the instrument for most accurate use. The fact that this tonometer is a high displacement tonometer which requires specific calibration, not only for species but for age of the animal (or size of the eye) is illustrated. A minor modification of the instrument--reducing the setting of a pressure regulator which supplies gas to the tip from its nominal 9 PSI to 2.5 PSI--is illustrated as a method of reducing the difference between intraocular pressure before tonometry and during tonometry, Pt-Po.

Animals↗
PubMed1989

A large multicentre, parallel group, double-blind study comparing tenoxicam and piroxicam in the treatment of osteoarthritis and rheumatoid arthritis.

A total of 1,328 patients with osteoarthritis or rheumatoid arthritis were entered into this double-blind, parallel group study of tenoxicam and piroxicam. The patient populations were well matched. An improvement was seen in pain on moving and at night in both groups and in both indications. Stiffness was also improved by both drugs, being most marked in the rheumatoid arthritis group. The primary efficacy variable was global assessment, and this showed tenoxicam to have slightly greater effect in osteoarthritis and the reverse in rheumatoid arthritis. There were no statistically significant differences in any of these findings. There were no significant differences in tolerance ratings, although the more serious gastrointestinal events occurred in the piroxicam group.

Adult↗
PubMed1988

The anatomy and pathology of the brachial plexus as demonstrated by computed tomography.

Computed tomography (CT) scanning can demonstrate a wide range of abnormalities affecting both the brachial plexus and the surrounding structures. Narrow section (4 mm) CT was used with bolus intravenous enhancement to examine the root of the neck and axilla in 62 patients with cancer, many of whom had symptoms of brachial plexus neuropathy. The normal anatomy of the plexus and its relations are described and illustrated. Examples of pathological changes caused by tumour and irradiation are also presented. The narrow scanning width (4 mm) is needed as the details sought are small and will be missed on thicker slice widths, particularly the changes of fibrosis in the upper axilla. Injection of intravenous contrast medium is essential for identifying the vascular structures which are used to locate the brachial plexus, especially when disease processes and post-irradiation fibrosis have destroyed the tissue planes.

Axilla↗
PubMed1988

The emergency department ECG and immediately life-threatening complications in initially uncomplicated suspected myocardial ischemia.

The emergency physician's disposition of patients with suspected myocardial ischemia is currently debated; some physicians believe that a subgroup of patients can be managed safely outside the coronary care unit. Clinical predictors are needed in assessing the patient with suspected myocardial ischemia to help identify this subgroup. Through a retrospective cohort study, we investigated the value of the initial emergency department ECG in discriminating between chest pain patients with low and high risk for immediately life-threatening complications. Two hundred eleven initially uncomplicated consecutive coronary care unit admissions with suspected unstable angina or myocardial infarction were studied. Patient outcome, including the incidence of myocardial infarction, complications, and mechanical and pharmacologic interventions, was reviewed. Immediately life-threatening complications included ventricular fibrillation, ventricular tachycardia, shock, 2 degrees and 3 degrees block, and death. Mechanical interventions included electrocardioversion or defibrillation, endotracheal intubation, intra-aortic balloon pump, Swan-Ganz catheter, or pacemaker insertion. Pressors, antiarrhythmics, and vasodilators were the reviewed pharmacologic interventions. A positive ECG was defined by the presence of ST elevation or depression, T wave inversion, left ventricular hypertrophy, left bundle branch block, paced rhythm, or new Q waves. All other ECG interpretations were considered negative. Patients were divided into two groups based on this initial emergency physician ECG interpretation and their complication incidences compared. Of the 211 patients, 96 had a positive ECG; 115 had negative ECGs. Patients with positive ECGs were older, had a greater history and concurrent incidence of myocardial infarction, and more complications and intensive interventions.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1986

Use of the CAPE Pm test with disabled patients.

The problem of estimating a patient's CAS total score on CAPE when the Pm (maze) test cannot be administered because of sensory or physical--rather than cognitive--disability is considered. Analysis of the scores of 250 elderly subjects who attempted all three CAS tests suggests that 'proration' of total CAS scores from I/O and MA scores, while preferable to alternative methods, results in error in a significant proportion of cases and should be used only with caution.

Aged↗
PubMed1981

A prospective study of the radiological changes in the cervical spine in early rheumatoid disease.

The cervical spine radiographs of 100 patients with early rheumatoid disease were studied annually, on a prospective basis, for a mean follow-up period of 7 years 2 months. Atlantoaxial subluxation developed in 12 patients. The subluxation was more frequent in females, more severe in patients with progressive, seropositive, erosive rheumatoid disease, and more marked in patients treated with oral corticosteroids. Subaxial subluxation, affecting upper cervical disc levels, occurred in a further 20 patients. Three patients developed vertical subluxation. The mobility of the cervical spine affects the degree of subluxation achieved, and when assessing serial films for subluxation it may be necessary to measure the cervical spine flexion before deciding whether subluxation has progressed or not. Over 80% of the patients with subluxation developed the first evidence of subluxation within 2 years of disease onset. Subluxation in the cervical spine is not, therefore, a late complication of rheumatoid disease. During the follow-up period none of the patients developed neurological signs.

Arthritis, Rheumatoid↗