Biomedical subjects

C Rae

Publications and source records attributed to C Rae.

35 records · Page 2Linked to original sources

PubMed1998

Brain biochemistry in Duchenne muscular dystrophy: a 1H magnetic resonance and neuropsychological study.

Duchenne muscular dystrophy (DMD) is a progressive muscle disorder associated with an intellectual deficit which is non-progressive. We obtained localised 1H magnetic resonance spectra from the left frontal lobe and left cerebellum of 15 boys with DMD (mean age 106 months+/-32) and 15 similarly aged control boys (mean age 115 months+/-31); all boys underwent a battery of neuropsychological tests. We found a significant (P<0.01) increase in the ratio of choline-containing compounds to N-acetylaspartate (Cho/NA) in the left cerebellum in boys with DMD compared with control boys. There was no change in the creatine/NA ratio and a significant increase (P=0.03) in the Cho/creatine ratio, suggesting that the change in Cho/NA ratio was due to an increase in choline-containing compounds; this increase has been previously observed in the brain of the murine model of DMD, the mdx mouse. No significant changes were observed in spectra obtained from left frontal lobe in DMD compared to controls. We also observed a significant association between Cho/NA in the left cerebellum, and the performance of DMD boys on the Matrix Analogies Test (MAT). The MAT is a test of visuo-spatial ability and non-verbal reasoning which requires neither manual dexterity nor a verbal response for an adequate performance. A comparison of DMD boys whose cerebellar Cho/NA fell within 2 standard deviations of the control norm (0.56+/-0.24) with DMD boys whose cerebellar Cho/NA was outside this range (i.e. >0.80) revealed a significant difference in ability on the MAT (P<0.05). DMD boys whose Cho/NA ratio is more than two standard deviations higher than controls perform significantly better on the MAT than DMD boys whose Cho/NA ratio is within the normal range. This finding suggests that the observed elevation in Cho/NA and Cho/creatine is not associated with intellectual deficit (as sampled by the MAT), and may represent a compensatory mechanism. The possible interpretations of these metabolic changes are discussed.

Age Factors↗

PubMed1998

Metabolic abnormalities in developmental dyslexia detected by 1H magnetic resonance spectroscopy.

BACKGROUND: Neurological and physiological deficits have been reported in the brain in developmental dyslexia. The temporoparietal cortex has been directly implicated in dyslexic dysfunction, and substantial indirect evidence suggests that the cerebellum is also implicated. We wanted to find out whether the neurological and physiological deficits manifested as biochemical changes in the brain. METHODS: We obtained localised proton magnetic resonance spectra bilaterally from the temporo-parietal cortex and cerebellum of 14 well-defined dyslexic men and 15 control men of similar age. FINDINGS: We found biochemical differences between dyslexic men and controls in the left temporo-parietal lobe (ratio of choline-containing compounds [Cho] to N-acetylaspartate [NA] p< or =0.01) and right cerebellum (Cho/NA, p< or = 0.01; creatine [Cre] to NA p< or =0.05; (not significant). We found lateral biochemical differences in dyslexic men in both these brain regions (Cho/NA in temporo-parietal lobe, left vs right, p< or =0.01; Cre/NA in cerebellum, left vs right, p< or =0.001). We found no such lateral differences in controls. There was no significant relation between the degree of contralateral chemical difference and handedness in dyslexic or control men. INTERPRETATION: We suggest that the observed differences reflect changes in cell density in the temporo-parietal lobe in developmental dyslexia and that the altered cerebral structural symmetry in dyslexia is associated with abnormal development of cells or intracellular connections or both. The cerebellum is biochemically asymmetric in dyslexic men, indicating altered development of this organ. These differences provide direct evidence of the involvement of the cerebellum in dyslexic dysfunction.

Adult↗

PubMed1998

Excitatory amino acid synthesis in hypoxic brain slices: does alanine act as a substrate for glutamate production in hypoxia?

Excitatory amino acids are an important cause of cell death in the hypoxic and ischaemic brain. Neuronal glutamate stores are depleted rapidly in hypoxia, but alanine production rises under such conditions and has been suggested to be a potential precursor of glutamate. To test this hypothesis, we have investigated amino acid metabolism using 13C NMR with superfused guinea pig cortical slices subjected to varying degrees of hypoxia. During severe hypoxia, brain slices metabolising 5 mM [2-(13)C]pyruvate exported [2-(13)C]alanine into the superfusion fluid. The metabolic fate of alanine during normoxia and hypoxia was tested by superfusion of brain slices with 10 mM glucose and 2 mM [2-(13)C,15N]alanine. Metabolism of exogenous alanine leads to the release of aspartate into the superfusion fluid. The pattern of labelling of aspartate indicated that it was synthesised via the glial-specific enzyme pyruvate carboxylase. 13C-labelled glutamate was produced with both normoxia and hypoxia, but concentrations were 30-fold lower than for labelled aspartate. Thus, although substantial amounts of glutamate are not synthesised from alanine in hypoxia, there is significant production of aspartate, which also may have deleterious effects as an excitatory amino acid.

Alanine↗

PubMed1998

Brain biochemistry in Williams syndrome: evidence for a role of the cerebellum in cognition?

OBJECTIVE: To determine what biochemical changes may occur in the brain in Williams syndrome (WS) and whether these changes may be related to the cognitive deficits. BACKGROUND: WS is a rare, congenital disorder with a characteristic physical, linguistic, and behavioral phenotype with known cognitive deficits. METHODS: We obtained 31P magnetic resonance spectra (MRS) from a region consisting of mostly frontal and parietal lobe of 14 patients with WS (age, 8 to 37 years) and 48 similarly-aged controls. 1H MRS (27 cm3) localized to the left cerebellum obtained from the WS cohort were compared with those from 16 chronological age- and sex-matched normal controls. A battery of cognitive tests were administered to all subjects undergoing 1H MRS. RESULTS: WS brains exhibited significant biochemical abnormalities. All 31P MRS ratios containing the phosphomonoester (PME) peak were significantly altered in WS, suggesting that PME is significantly decreased. Ratios of choline-containing compounds and creatine-containing compounds to N-acetylaspartate (Cho/NA and Cre/NA) were significantly elevated in the cerebellum in WS cf. controls, whereas the ratio of Cho/Cre was not altered. This suggests a decrease in the neuronal marker N-acetylaspartate in the cerebellum. Significant correlations were found between the cerebellar ratios Cho/NA and Cre/NA and the ability of all subjects at various neuropsychological tests, including Verbal and Performance IQ, British Picture Vocabulary Scale, Ravens Progressive Matrices, and Inspection Time. CONCLUSIONS: The correlations can be interpreted in two ways: 1) Our sampling of cerebellar biochemistry reflects a measure of "global" cerebral biochemistry and is unrelated to cerebellar function, or 2) The relations indicate that cerebellar neuronal integrity is a requirement (on a developmental time scale or in real-time) for ability on a variety of cognitive tests.

Adenosine Triphosphate↗

PubMed1996

Is pH a biochemical marker of IQ?

We have measured intracellular brain pH in vivo in 42 boys and found a significant correlation between this biochemical parameter and samples of intelligent behaviour. To the best of our knowledge this is the first reported relation between a biochemical marker which is within normal physiological values and intellectual ability. pH is one of the most accurate parameters that can be measured by 31P magnetic resonance spectroscopy and it reflects sensitively cellular ionic status and metabolic activity. The observed correlation, although not implying a causal relation, raises the possibility that intelligent behaviour may be influenced by the ionic status of brain tissue, or vice versa.

Brain↗

PubMed1996

Polychlorinated biphenyls in small mammals from contaminated landfill sites.

PCBs in small mammals living in a contaminated landfill site varied in both concentration and congener composition, reflecting both the feeding strategy of each species and the chlorination of the congeners. Body concentrations of SigmaPCB on a 1:1:1 Arochlor (1242:1254:1260) basis gave a rank order of Sorex araneus >> Apodemus sylvaticus > Microtus agrestis. Comparisons with data for other PCB-contaminated sites suggest that the SigmaPCB values in S. araneus from the landfill site may be of sufficient magnitude to inhibit reproductive capability and to cause other major organ dysfunction. There were also inter-specific differences in the whole body concentrations of the individually quantified congeners. For S. araneus, the rank order of concentrations of the selected ICES 7 marker PCB congeners was: No. 153>138>>180>118>>28>101>52. In M. agrestis the rank order was: No. 28>118>153>138>180, 52>101. The congener order for A. sylvaticus differed from both S. araneus and M. agrestis, but was more similar to the latter, being: No. 153>138>118>180>28>52, 101. The contribution of the different congeners to SigmaPCB in the mammals reflects the degree of chlorination of different congeners but varies with feeding strategy of the species. In the insectivorous S. araneus, heavily chlorinated congeners (Nos 153, 138 and 180) are most abundant, whereas the herbivore, M. agrestis showed congeners Nos 28 and 118 to be present in the highest amounts, whilst the equivalent pattern for the more omnivorous A. sylvaticus was intermediate between S. araneus and M. agrestis. The high concentrations of PCBs in S. araneus relative to the other species, suggests that it may serve as a useful biomarker of terrestrial environmental pollution by organochlorines.

Journal Article↗

PubMed1996

Human erythrocyte choline uptake in uraemia: the role of intracellular substrate and an investigation into the effects of haemodialysis.

1. Erythrocyte choline transport was studied in 10 haemodialysis patients immediately before and after a haemodialysis session and in 10 control subjects. Choline uptake was measured in erythrocytes from normal and uraemic patients after washing in vitro and subsequent incubation in autologous plasma. Amines present in uraemic plasma were examined for their effect on choline transport in normal erythrocytes. 2. NMR spectroscopy was used to measure choline, trimethylamine and dimethylamine in erythrocyte extracts from nine control subjects, 32 subjects with renal impairment and nine samples from haemodialysis patients. 3. The increased choline influx in uraemic erythrocytes is significantly decreased by prior haemodialysis (mean Vmax pre-dialysis 146 +/- 20 mumol h-1 l-1, post-dialysis 113 +/- 13 mumol h-1 l-1 (P < 0.005). After in vitro washing there is a fall in Vmax, and no longer any significant difference between pre- and post-dialysis samples. There remains a significant difference in the erythrocyte choline Vmax between samples from patients with chronic renal failure and from normal subjects (P < 0.005). 4. Human plasma was found to contain factors capable of increasing choline uptake. Trimethylamine and dimethylamine were found to inhibit choline uptake. Trimethylamine and trimethylamine-N-oxide trans-stimulated choline efflux, but the major transport substrate present in erythrocyte extracts from all groups was choline, which was higher in those with renal impairment (71 +/- 10 mumol/l) than in haemodialysis patients (47 +/- 10 mumol/l) and control subjects with normal renal function (40 +/- 9 mumol/l). 5. Our data suggest that erythrocyte choline transport is increased in uraemia as a consequence of increased transporter number or activity, rather than the presence of intracellular substrate.

Choline↗

PubMed1996

Anion channel blockers inhibit swelling-activated anion, cation, and nonelectrolyte transport in HeLa cells.

The effect of osmotic cell swelling on the permeability of HeLa cells to a range of structurally unrelated solutes including taurine, sorbitol, thymidine, choline, and K+ (96Rb+) was investigated. For each solute tested, reduction in the osmolality of the medium from 300 to 200 mosmol/kgH2O caused a significant increase in the unidirectional influx rate. In each case, the osmotically activated transport component was nonsaturable up to external substrate concentrations of 50 mM. Inhibitors of the swelling-activated anion channel of HeLa cells [quinine, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, niflumate, 1,9-dideoxyforskolin, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB), and tamoxifen] blocked the osmotically activated influx of each of the different substrates tested, as well as the osmotically activated efflux of taurine and I-. Tamoxifen and NPPB were similarly effective at blocking the osmotically activated efflux of 96Rb+. The simplest of several hypotheses consistent with the data is that the osmotically activated transport of the different solutes tested here is via a swelling-activated anion-selective channel that has a significant cation permeability and a minimum pore diameter of 8-9 A.

Anions↗

PubMed1994

Stereospecificity of substrate usage by glyoxalase 1: nuclear magnetic resonance studies of kinetics and hemithioacetal substrate conformation.

The specificity of glyoxalase 1 for the diastereomers of its hemithioacetal substrate [which forms spontaneously between an alpha-keto aldehyde and reduced glutathione (GSH)] was investigated by exploiting the differences between their 1H NMR spectra at pH* 4.4. The 1H NMR spectra of the hemithioacetals of glutathione with phenylglyoxal or methylglyoxal were assigned with the aid of conventional decoupling and two-dimensional NMR spectroscopic techniques. The rate of interconversion of the diastereomers was determined at 30 degrees C from the results of an inversion-transfer technique and found to be 0.30 +/- 0.04 s-1 (+/- sd) in the case of phenylglyoxal and 0.15 +/- 0.02 s-1 in the case of methylglyoxal. Stereopreference of the enzyme was tested by the addition of large amounts of yeast glyoxalase 1 to a reaction mixture; glyoxalase 1 preferentially operated on one diastereomer of the phenylglyoxal hemithioacetal but the diastereomers of methylglyoxal appeared to be operated upon indiscriminately. From computer models of the kinetics of possible reaction schemes, a mechanism involving glyoxalase 1 catalysis of both diastereomers of the hemithioacetals was shown to be the most consistent with the experimental data. Estimates of internuclear distances in the diastereomers, obtained from 2D NMR spectra were used in "dynamical simulated annealing" calculations to generate likely structures of the substrates. Relative ring-current shifts obtained from 1D NMR spectra were used, together with a ring-current shift algorithm, to select structures with compatible conformations. We conclude that the rate of conversion of substrate by the enzyme is dependent upon the overall conformation of the substrate molecule, rather than merely its stereochemical configuration (R or S).

Glutathione↗

PubMed1992

Aldolase-catalyzed diketone phosphate formation from oxoaldehydes. NMR studies and metabolic significance.

NMR spectroscopy showed fructose-1,6-bisphosphate aldolase from rabbit muscle accepts as substrates, in lieu of glyceraldehyde 3-phosphate, the oxoaldehydes methylglyoxal and phenylglyoxal but not hydroxymethylglyoxal. The enzyme catalyzed an aldol condensation between the oxoaldehyde and dihydroxyacetone phosphate to form a monophosphorylated diketone and was inactivated in the process. Circumvention of this reaction, by metabolism of oxoaldehydes to hydroxy acids, may be a metabolic role for the glyoxalase enzyme system. Transketolase and transaldolase were found not to accept oxoaldehydes as substrates in place of glyceraldehyde 3-phosphate.

Animals↗

PubMed1991

Glyoxalase 2 deficiency in the erythrocytes of a horse: 1H NMR studies of enzyme kinetics and transport of S-lactoylglutathione.

In mammalian red blood cells the metabolism of methylglyoxal, and some alpha-ketoaldehydes, takes place via two, generally, highly active enzymes, glyoxalase 1 and 2. The 1H NMR spin-echo spectra of horse erythrocytes, and the various reactants in the glyoxalase system, were characterized as a prelude to obtaining series of spectra in time courses of methylglyoxal metabolism. We characterized the kinetics of the enzyme system in red cells from a normal horse and also from one which had very low activity of glyoxylase 2. The kinetics of the reaction scheme, with methylglyoxal as the starting substrate, were obtained from 1H NMR spectra and analyzed with a computer model of the scheme. The most salient feature of the normal system was the very high feed-forward inhibition (KiHTA = 0.1 microM) of glyoxalase 2 by the hemithioacetal which is the substrate of glyoxalase 1. The glyoxalase-2-deficient red cells were used to test whether S-lactoylglutathione is transported from red cells via the glutathione-S-conjugate transporter; this transport appeared not to occur. Because methylglyoxal is extremely rapidly removed (half-life, approximately 5 min) from normal red cells, it is difficult to assess the effect of this compound on glycolysis but the slow decline evident in the deficient cells allowed a study of the effects on L-lactate production; no effects were apparent.

Animals↗

PubMed1991

Changes in hepatic glutathione metabolism in diabetes.

Glutathione is important in the regulation of the redox state, and a decline in its tissue level has often been considered to be indicative of increased oxidative stress in diabetes. In this study of diabetic rats, the level of hepatic glutathione was normal unless food intake was restricted. Thus, the previous report of a reduction in hepatic glutathione in diabetes is likely to be the result of food deprivation rather than diabetes alone. In contrast to changes characteristic of oxidative stress, the efflux of glutathione in bile from diabetic animals was significantly decreased, whereas hepatic mixed disulfides were unchanged, and the hepatic gamma-glutamyltransferase activity was considerably increased. These changes were not reproduced by food deprivation. The decrease in biliary excretion of glutathione in diabetes may reflect an attempt to conserve glutathione by activation of the hepatic gamma-glutamyl cycle. We conclude that the disturbances of glutathione metabolism in diabetes are not typical of those seen in oxidative stress or food restriction.

Animals↗

PubMed1990

Kinetic analysis of the human erythrocyte glyoxalase system using 1H NMR and a computer model.

1H NMR was used with methylglyoxal, purified by an HPLC technique, to study the kinetics of the human erythrocyte glyoxalase system. 1H NMR enabled the direct measurement of the time-dependent changes in concentrations of the two hydrates of methylglyoxal, which have not previously been directly measurable, as well as measurement of substrates and products of the glyoxalase enzyme system in the human red blood cell. A computer model of the reaction scheme was developed and NMR data numerically analyzed, thus allowing a complete kinetic description of the reactions. The rate constants describing the chemical equilibria between the hydrated species of methylglyoxal were determined by this numerical analysis or by a saturation-transfer technique, and found to be much slower (by several orders of magnitude) than previously determined by other methods. The kinetic parameters describing the enzyme-catalyzed reactions were also determined from experiments using a dilute haemolysate that was added to solutions of methylglyoxal and reduced glutathione (GSH). The maximal velocity of glyoxalase 1 is threefold greater (Vmax = 70.4 +/- 4.7 mmol.min-1.1 packed cells-1) than glyoxalase 2(Vmax = 24 +/- 5 mmol.min-1.1 packed cells-1) and it exhibits threefold-greater affinity for its substrate (Km = 0.46 +/- 0.04 mM) than the second enzyme (Km = 1.5 +/- 0.4 mM). Both enzymes are subject to competitive inhibition; glyoxalase 1 by reduced glutathione (KiGSH = 7.88 +/- 0.16 mM) and glyoxalase 2 by the hemithioacetal (HTA) of methylglyoxal and GSH (KiHTA = 0.29 +/- 0.04 mM).

Erythrocytes↗

PubMed1990

Diabetes, hyperinsulinemia, and hyperlipidemia in small aboriginal community in northern Australia.

A small rural Aboriginal community in northern Australia was surveyed for diabetes, impaired glucose tolerance (IGT), hyperinsulinemia, and lipid levels. Of the 122 adults greater than 17 yr of age who participated (95% response rate), 11.5% had diabetes, 7.4% had IGT, and the remaining 81.1% had normal glucose tolerance. Both diabetes and IGT were strongly age related. This high frequency of diabetes occurred, despite the population being relatively lean. Although the body mass index (BMI) increased with age in both men and women, only 25% of the population overall had BMI greater than 25 kg/m2. There were wide ranges of insulin responses to glucose, with the upper tertile of 2-h insulin levels being more than seven times higher than the lower tertile (144 +/- 13 vs. 19 +/- 1 mU/L). Hyperinsulinemia was associated with IGT, elevated triglycerides, and lower high-density lipoprotein cholesterol levels. Lipid abnormalities were much more frequent among men than women. Cholesterol levels were an average of 0.55 mM higher and triglycerides an average of 1.05 mM higher in men than in women, and both increased with age. In conclusion, this small isolated Aboriginal population from northern Australia had an unexpectedly high frequency of diabetes (in view of their relative leanness) in association with a high frequency of metabolic abnormalities indicative of insulin resistance (hyperinsulinemia, IGT, hypertriglyceridemia).

Adolescent↗

PubMed1989

High level expression in Escherichia coli of the DNA-binding domain of the glucocorticoid receptor in a functional form utilizing domain-specific cleavage of a fusion protein.

A fragment comprising the DNA-binding domain of the human glucocorticoid receptor has been expressed in a functional form in Escherichia coli as a fusion protein with protein A from Staphylococcus aureus. The DNA-binding domain was purified to apparent homogeneity by affinity chromatography on IgG-Sepharose and DNA-cellulose, a purification scheme which does not involve denaturation of the protein at any step. The DNA-binding domain was separated from the protein A part of the fusion protein by domain-specific enzymatic cleavage with chymotrypsin while immobilized on IgG-Sepharose. The recombinant protein has been characterized by amino acid analysis, NH2- and COOH-terminal sequence analysis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and reactivity to iodoacetate and was found to correspond to the primary structure derived from the cDNA sequence. DNase I footprinting showed that the purified recombinant protein bound to the same DNA sequences on the mouse mammary tumor virus long terminal repeat as glucocorticoid receptor purified from rat liver does. About 10 times more recombinant protein, on a molar basis, was needed to obtain the same level of protection. However, the protection of the three different footprints (1.3, 1.4, and 1.5') by the recombinant protein differed greatly from that of the natural receptor, with virtually no protection of footprint 1.4. This indicates cooperative binding of the natural receptor to adjacent footprints, dependent on other regions of the receptor than the DNA-binding domain.

Amino Acid Sequence↗

PubMed1988

Development of a simple, fluoroimmunoassay for paraquat.

High affinity populations of antibodies to paraquat were produced in high concentrations in the sera of immunised sheep. A simple fluoroimmunoassay was developed for rapid measurement of plasma paraquat. Fluorescein-labelled paraquat was added to sample or standard (20 microliter) followed by premixed first and second antisera. Equilibrium was reached within 2 min and results were available within 20 min. Correlation with weighed-in paraquat gave a regression line (r) of 0.997. All reagents proved stable and are suitable for general distribution.

Animals↗

PubMed1988

Development of a specific polarisation fluoroimmunoassay for paracetamol in serum.

A specific polarisation fluoroimmunoassay for the measurement of paracetamol levels in serum has been developed for use in emergency toxicological screening. It is based on the use of a fluorescein-labelled analogue and a sheep antiserum and exploits the rapid dissociation kinetics of the hapten-antibody complex to enable the label and antiserum to be combined as a single reagent. Paracetamol levels are determined by adding 5 microL of serum to 1.5 mL of the single reagent, incubation at ambient temperature for a few minutes and measurement of fluorescence polarisation. In addition to its speed and simplicity, the assay is both accurate and precise and the results obtained correlate closely with those from the commonly used chemical and enzymatic techniques.

Acetaminophen↗