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Biomedical subjects

J Christodoulou

Publications and source records attributed to J Christodoulou.

At least 19 recordsLinked to original sources
PubMed1997

Automated quantitation of total protein in cultured skin fibroblasts.

An automated Coomassie Blue method of measuring total protein in cultured skin fibroblasts using a random access analyzer, the Roche MIRA, was compared to a manual dye binding method using bicinchoninic acid. The automated Coomassie Blue method is in common use in many routine laboratories for measurement of total protein in urine and CSF, and was found by us to be significantly faster than a manual protein method, with an average assay time of 2 min per sample. In addition, intra- and inter-run precision were comparable for the automated and manual methods, and compared favourable with other automated methods. We recommend that where the MIRA apparatus is available, consideration be given to its use for protein quantitation of cell or tissue extracts.

Fibroblasts↗
PubMed1997

Intragenic complementation at the human argininosuccinate lyase locus. Identification of the major complementing alleles.

To determine the molecular and biochemical basis of intragenic complementation observed at the human argininosuccinate lyase (ASL) locus, we identified the ASL alleles in ASL-deficient cell strains with two unique complementation phenotypes: (i) frequent complementers, strains that participated in the majority of complementation events, and (ii) high activity complementers, strains in which complementation was associated with a relatively high level of restoration of ASL activity. Four mutations (Q286R, D87G, A398D, and a deletion of exon 13) were identified in the four strains examined. One of the two frequent complementers was homozygous, and the other heterozygous, for the Q286R allele. Similarly, one of the two high activity complementers was homozygous, and the other heterozygous, for the D87G allele. When the Q286R and D87G mutations were introduced by site-directed mutagenesis into wild-type ASL cDNA, each conferred loss of ASL activity in COS cell transfection assays. To test directly the hypothesis that intragenic complementation occurs at the ASL locus, one of the major complementation events observed previously, between strains carrying the Q286R and D87G alleles, was reconstructed in COS cell transfection assays. A partial restoration of ASL activity, comparable with the increase seen in the fibroblast complementation analysis, was observed on joint cotransfection of these two alleles. The results provide molecular confirmation of the major features of the ASL mutant complementation map, identify the Q286R and D87D alleles as the frequent and high activity complementing alleles, respectively, and provide direct proof of intragenic complementation at the ASL locus.

Alleles↗
PubMed1997

Mitochondrial electron transport chain defect presenting as hypoglycemia.

A profoundly deaf female infant was found to have hypoglycemia and lactic acidemia after an episode of decreased oral intake and vomiting. Electron transport chain (ETC) enzyme studies revealed a combination defect of complexes I, III, and IV in liver but not in skeletal muscle. This case highlights the fact that defects of the ETC are clinically highly heterogeneous and should be considered with hypoglycemia and lactic acidosis in the absence of a glycogen storage disorder. Moreover, ETC defects can occur with a biochemical profile suggestive of a fatty acid oxidation disorder.

Acidosis, Lactic↗
PubMed1997

Mitochondrial myopathy with tRNA(Leu(UUR)) mutation and complex I deficiency responsive to riboflavin.

Deficiency of complex I (reduced nicotinamide adenine dinucleotide dehydrogenase-ubiquinone oxidoreductase) of the mitochondrial respiratory chain may be seen as a pure myopathy or as a neuromuscular disorder at presentation. Efficacy of long- term therapy for these disorders is yet to be established. We report the case of a female patient with complex I deficiency and skeletal myopathy, who has had a sustained clinical response to riboflavin during 3 years of therapy. Molecular studies found no mutations in the putative flavin mononucleotide binding site in the 51 kd subunit of complex I, but a T-to-C transition at nucleotide 3250 in the mitochondrial DNA tRNA(Leu(UUR)) gene was identified. This mutation has been reported in one other family in that five members had fatigue with or without muscle weakness. There were also five cases of unexplained infant deaths in that family and two cases in the family reported here. Riboflavin therapy should be attempted in all patients with complex I deficiency when the clinical presentation is one of isolated skeletal myopathy.

Carnitine↗
PubMed1997

Progressive myoclonic epilepsies: recent genetic advances.

The progressive myoclonic epilepsies are a rare group of debilitating epileptic encephalopathies characterized by myoclonic seizures, progressive neurological dysfunction and dementia. In the past year advances in gene mapping have isolated gene loci for the majority of progressive myoclonic disorders, paving the way for specific diagnosis, more accurate prognosis and risk calculation, as well as opening the potential for prenatal and pre-symptomatic diagnosis in at risk families.

Adolescent↗
PubMed1996

First prenatal diagnosis of the carnitine transporter defect.

We report the first attempt at prenatal diagnosis of the carnitine transporter defect in a fetus at high risk of having the disorder. Analysis of cultured CVS after prolonged culture predicted that the fetus was not affected but might be heterozygous for the carnitine transporter defect, but chromosome 15 satellite DNA markers showed no paternal contribution, suggesting that the CVS cells assayed were of predominantly maternal origin. Subsequent assay of cultured amniocytes predicted that the fetus would be affected, and this was confirmed in the newborn period. We conclude that prenatal diagnosis of the carnitine transporter defect is possible, but where results depend on extended culture of CVS, molecular studies should be performed to confirm genetic contributions from both parents.

Carnitine↗
PubMed1996

Leigh syndrome: clinical features and biochemical and DNA abnormalities.

We investigated the etiology of Leigh syndrome in 67 Australian cases from 56 pedigrees, 35 with a firm diagnosis and 32 with some atypical features. Biochemical or DNA defects were determined in both groups, ie, 80% in the tightly defined group and 41% in the "Leigh-like" group. Eleven patients had mitochondrial DNA point mutations (nucleotide [nt] 8993 T to G, nt 8993 T to C, or nt 8344 A to G) and 1 Leigh-like patient had a heteroplasmic deletion. Twenty-nine patients had enzyme defects, ie, 13 respiratory chain complex I, 9 complex IV, and 7 pyruvate dehydrogenase complex (PDHC). Complex I deficiency is more common than recognized previously. Six PDHC-deficient patients had mutations in the X-chromosomal gene encoding the E1alpha subunit of PDHC. Parental consanguinity suggested autosomal recessive inheritance in two complex IV-deficient sibships. We found no strong correlation between the clinical features and basic defects. An assumption of autosomal recessive inheritance (frequently made in the past) would have been wrong in nearly one-half (11 of 28 tightly defined and 18 of 41 total patients) of those in whom a cause was found. A specific defect must be identified if reliable genetic counseling is to be provided.

Brain↗
PubMed1996

Pregnancy and argininosuccinic aciduria.

We present the outcome of a pregnancy in a woman with mild argininosuccinic lyase deficiency to add to the collective experience of the maternal and fetal effects of urea cycle defects. In females affected with argininosuccinic lyase deficiency, careful clinical and biochemical monitoring of pregnancy will minimize the risk of metabolic decompensation in the perinatal period. Furthermore, it would appear that argininosuccinate is not teratogenic to the development of the human fetus.

Adult↗
PubMed1996

Repolarization abnormalities with prolonged hyperventilation in apparently healthy subjects: incidence, mechanisms and affecting factors.

Brief hyperventilation is occasionally accompanied by repolarization abnormalities in subjects without apparent heart disease. However, the effect of prolonged hyperventilation on repolarization abnormalities is not clearly defined. We analysed the repolarization abnormalities induced by prolonged hyperventilation, correlated with hyperventilation induced haemodynamic changes and exercise test results and assessed the effect on them of age, gender, smoking and hypertension. Prolonged hyperventilation (overbreathing at least 30 respirations/min for 5 min and 10 min recovery) was performed in 474 healthy volunteers (269 men, 205 women) 42.6 +/- 13.5 years old. The electrocardiogram was analysed for transient ST depression or T-wave inversion. Repolarization abnormalities were observed in 72 subjects, 47 men (17.5%) and 25 women (12.2%). Age, gender, smoking and hypertension did not influence the overall incidence of repolarization abnormalities. ST depression was more frequently observed in women (4.4 vs 0.7%, P < 0.01), while T wave inversion from negative to positive was more frequent in men (8.9 vs 2.4%, P < 0.006). All subjects with ST depression were non-smokers (4.1% of the non-smokers, P < 0.003 compared to smokers). Repolarization abnormalities occurred usually (80.5%), but not exclusively, within the first hyperventilation minute. Subjects with repolarization abnormalities developed higher heart rate change with hyperventilation than those without repolarization abnormalities (29.2 +/- 13.1 vs 24.2 +/- 12.7, P < 0.002). A positive exercise test was observed in 45.4% of subjects with hyperventilation induced ST depression but only in 13.1% of subjects with T-wave changes, P < 0.02. Repolarization abnormalities are not uncommon during prolonged hyperventilation. Hear rate change affects the occurrence of repolarization abnormalities, while gender and smoking influence the type of repolarization abnormalities. ST depression but not T-wave inversion during hyperventilation is associated with a positive exercise test. Increased understanding about hyperventilation-induced repolarization abnormalities could affect use of the method in the clinic, either as a provocation test or as a complement to the interpretation of the exercise test.

Adult↗
PubMed1996

Hyperventilation test in syndrome X.

The hyperventilation (HV) test has been extensively used in different forms of coronary artery disease. The purpose of this work was to investigate the response to HV in patients with syndrome X and compare HV with exercise (EX) test. The authors studied 20 patients with syndrome X (angina, a positive EX test, and normal coronary angiogram) and 20 healthy subjects who underwent HV and EX tests. In 7 patients, all women, angina and electrocardiographic (ECG) changes occurred during HV but in none of the controls. Patients with syndrome X and controls had a similar rate-pressure product (RPP) at rest and achieved a similar RPP with HV. The RPP achieved with HV in patients with syndrome X without ECG changes was significantly lower, 121.8 +/-29.1(mean +/-SD), than what was achieved by those with changes, 167.9 +/-42.9, P < 0.01. In patients with an abnormal response to HV the RPP at which angina and ECG changes occurred was similar to that where similar changes were observed during EX. There was a significant correlation of the RPPs between the two tests, r=0.82, P < 0.02. In contrast, in patients in whom no angina or ECG changes occurred, the RPP they achieved with HV was significantly lower than the anginal threshold during EX 204 +/- 47.4, P < 0.0001. In conclusion, a significant proportion of patients with syndrome X, mainly women, who achieve a high RPP with HV, develop angina and ECG changes during overbreathing. The close relation between EX and HV RPPs where these changes occur suggests an increased myocardial oxygen demand as the most likely underlying mechanism for this behavior.

Electrocardiography↗
PubMed1995

1H NMR of albumin in human blood plasma: drug binding and redox reactions at Cys34.

1H NMR methods are described which allow direct studies of the Cys34 binding site of albumin in intact human blood plasma in vitro. Antiarthritic gold drugs and the alcohol-aversive drug disulfiram induce a structural transition detectable via H epsilon 1 and H delta 2 resonances of His3 of albumin, and reactions of cystine, glutathione and captopril in plasma have also been investigated. Contrary to most assumptions, little of the albumin in normal plasma appears to be blocked at Cys34 as a cystine disulfide.

Alcohol Deterrents↗
PubMed1995

Hemodynamic response to hyperventilation test in healthy volunteers.

Hyperventilation is well known to affect the electrocardiogram (ECG) in subjects without heart disease and produce spasm in patients with variant angina. The autonomic nervous system is thought to play a significant role in these effects. However, the normal hemodynamic response to hyperventilation is not well defined. We subjected 369 healthy volunteers (200 men, 169 women) to prolonged hyperventilation (30 respirations for 5 min and 10 min recovery) under continuous ECG monitoring and to exercise testing. Heart rate (HR), systolic and diastolic blood pressures (SBP, DBP) and rate-pressure product were recorded. Hyperventilation resulted in an immediate (within the first min), significant increase in HR by 27.4%, a further small increase at min 2 of hyperventilation, and a subsequent small decrease in HR at mins 3-5. An immediate drop of HR by 20.1% was observed with discontinuation of hyperventilation. Apart from a slightly higher HR increase in men, a similar pattern of HR changes was found in both genders. On multivariate analysis, younger age, absence of smoking, and male gender were associated with a higher HR increase with hyperventilation (p < 0.0001, p < 0.0001, and p < 0.001, respectively). SBP and DBP increased with hyperventilation, with their highest value at min 5 of hyperventilation and a subsequent drop to baseline levels. Age and gender did not affect the degree and pattern of BP changes. Absence of smoking and the presence of hypertension were associated with a higher SBP with hyperventilation (p < 0.003 and p < 0.007). The rate-pressure product increased by 43.6% with hyperventilation, a change that was only 19.1% of the respective rate-pressure product observed with exercise. Hyperventilation results in significant HR and BP increases, changes that are influenced by age, gender, smoking, and hypertension. Our study could serve as a standard for comparison of the hyperventilation effects in different disease states.

Adult↗
PubMed1995

Peroxisomal assembly defects: clinical, pathologic, and biochemical findings in two patients in a newly identified complementation group.

We describe the clinical, pathologic, and biochemical findings for two peroxisome-deficient patients in a newly identified complementation group. Both patients had biochemical findings typical of patients with peroxisome biogenesis disorders. However, whereas one patient had the typical clinicopathologic features of Zellweger syndrome, the other patient's phenotype was atypical.

Catalase↗
PubMed1995

Neonatal onset of medium-chain acyl-coenzyme A dehydrogenase deficiency with confusing biochemical features.

A female neonate was seen because of shock, ketosis, and undetectable blood glucose. Initial urinary findings indicated the possibility of a defect of fatty acid beta-oxidation; subsequent studies showed that she had medium-chain acyl-coenzyme. A dehydrogenase deficiency. This case highlights the fact that the initial symptoms may occur in the first few days of life, and that the presence of ketosis does not exclude the possibility of a fatty acid oxidation defect; the profiles of urinary organic acids and acylglycines may not be characteristic at that time.

Acyl-CoA Dehydrogenase↗
PubMed1995

A mutation causing DHPR deficiency results in a frameshift and a secondary splicing defect.

In our analysis of mutations causing DHPR deficiency we identified a patient in whom there was an aberrant transcription pattern detected by PCR of DHPR cDNA. However, unlike the pattern observed as a result of most splicing mutations, there is some full length transcript. The mutation was located and is a single nucleotide deletion at position 570/571 of the DHPR cDNA sequence and results in a frameshift and premature termination after the addition of six amino acids. The mutation is present in a homozygous state in the patient and in a heterozygous state in both parents. The exon which is deleted at high frequency in the patient is the putative exon 4, which is remote from the mutation, and confirms our observation that exon 4 skipping is a relatively common event.

Base Sequence↗
PubMed1994

A new structural transition of serum albumin dependent on the state of Cys34. Detection by 1H-NMR spectroscopy.

1. Reactions of fatty-acid-free bovine serum albumin and recombinant human albumin with a range of antiarthritic gold(I) complexes [auranofin, deacetylated auranofin, triethylphosphinegold(I) chloride] and related thiols (thioglucose, tetraacetylthioglucose, glutathione, dithiothreitol) have been investigated using 1H-NMR spectroscopy. 2. In reactions of albumin with auranofin, tetraacetylthioglucose and dithiothreitol, release of cystine was detected, whereas for deacetylated auranofin, thioglucose and glutathione, mixed disulphides with cysteine were produced. It has been previously proposed that Cys34 of human and bovine serum albumins is partly blocked by disulphide formation with cysteine and glutathione. The above reactions lead to deblocking by thiol-disulphide interchange reactions. No release of glutathione from albumin was detected. 3. Changes in the His H epsilon 1 regions of the 1H-NMR spectra show that albumin exists in two structural forms dependent on whether the side-chain of Cys34 is a free thiolate, or blocked by gold(I)triethylphosphine, by disulphide formation with cysteine or by another form of oxidation. We propose that Cys34 is either in a buried or in an exposed environment; the possible molecular basis of the structural change is discussed. 4. The relationship between reactions at Cys34, cysteine release, and the observed structural transition are discussed in terms of chrysotherapy, albumin metabolism and the use of gold(I) as a heavy atom derivative in X-ray crystallographic studies of albumins.

Amino Acid Sequence↗