Hyperhomocysteinemia, a risk factor for atherosclerosis in chronic uremic patients.

Hyperhomocysteinemia has been shown to constitute an independent risk factor for premature occlusive arterial disease (N Engl J Med 324:1149), a frequent complication in chronic uremic patients in whom homocysteine (Hcy) accumulation has been reported to occur. We prospectively determined fasting plasma level of total, protein-bound Hcy in 118 adult chronic uremic patients, either dialyzed or not. In 79 non-dialyzed patients (47 male) with various degrees of chronic renal failure (RF) assessed by creatinine clearance (CCr), none receiving folate, B6 or B12 vitamin supplementation, mean (+/- 1 SD) plasma Hcy level was 16.2 +/- 8.1 mumol/liter in 28 patients with mild RF (CCr 30 to 75 ml/min), 23.3 +/- 14.7 in 29 patients with moderate RF (CCr 10 to 29.9), and 29.5 +/- 14.4 in 22 patients with advanced RF (CCr < 10), a significant difference (P < 0.01 for all groups) compared to 45 healthy controls (8.2 +/- 2.2 mumol/liter). Linear regression analysis showed a significant correlation between plasma creatinine and Hcy concentrations (r = 0.49, P < 0.0001). Hcy was significantly higher in 20 patients (16 males) who had past histories of occlusive arterial disease than in the 59 (31 males) who did not (30.9 +/- 19.1 vs. 19.6 +/- 9.7 mumol/liter, P < 0.001) and all of the former had Hcy level > 14.1 mumol/liter (the upper limit in healthy controls) versus 35 of 59 in the latter.(ABSTRACT TRUNCATED AT 250 WORDS)

Organic acids in post-mortem cerebrospinal fluid.

Organic acids of cerebrospinal fluid (CSF) have been determined by gas chromatography-mass spectrometry in 29 post-mortem samples obtained from infants and 10 samples obtained from hospitalized children, as controls. Though the organic acids profile was similar in the two groups, eight organic acids not observed in CSF from live infants were inconstantly found in post-mortem CSF and for three of them, malic acid, lactyllactic acid and uracile, concentrations were correlated with the delay in sampling.

Early amniocentesis and amniotic fluid organic acid levels in the prenatal diagnosis of organic acidemias.

Organic acids were studied in amniotic fluids taken by early amniocentesis between 6 to 12 weeks in control pregnancies. The prenatal diagnosis of methylmalonic acidemia and propionic acidemia could be made by the measurement of methylmalonic and methylcitric acid respectively in the amniotic fluid taken simultaneously with chorionic villi at the 11th week of gestation.

Potentiation by piridoxilate of the synthesis of hippurate from benzoate in isolated rat hepatocytes. An approach to the determination of new pathways of nitrogen excretion in inborn errors of urea synthesis.

The synthesis of hippurate from benzoate as compared to ureagenesis has been investigated in isolated rat hepatocytes. Half-maximal synthesis of hippurate was observed at 0.3 mmol/l benzoate. In the presence of 1 mmol/l benzoate, hippurate synthesis proceeded linearly with time at a rate of 40 +/- 10 mumol X h-1 X g-1 dry weight. This provided less than 10% of nitrogen epuration supported by concomitant urea synthesis (350 +/- 82 mumol X h-1 X g-1 dry weight). The incorporation of benzoate to hippurate was markedly limited by the availability of glycine. Half-maximal hippurate synthesis was observed at 2 mmol/l glycine. In the absence of glycine, piridoxilate, a glyoxylate derivative, markedly potentiated hippurate synthesis. Half-maximal stimulation was observed at 10 mmol/l piridoxilate. In the presence of 10 mmol/l piridoxilate, hippurate synthesis (420 +/- 35 mumol X h-1 X g-1 dry weight) provided more than 50% of nitrogen epuration supported by urea synthesis. It is concluded that supplementation with nitrogen-free analogues of glycine such as piridoxilate are required to potentiate hippurate synthesis in an attempt to replace ureagenesis as an alternative pathway of waste nitrogen excretion in inborn errors of urea synthesis.

[Radioisotopic assay of total L-homocysteine in plasma and urine: application to serial determinations]. / Dosage radioisotopique de la L-homocystéine totale dans le plasma et l'urine: application à des dosages en séries.

The authors report here a simple radioenzymatic determination of total L-homocysteine in plasma and urine. L-homocysteine-containing disulfides were reduced with dithioerythritol. L-homocysteine was then condensed by S-adenosyl L-homocysteine hydrolase to 14C-adenosine to form S-14C adenosyl L-homocysteine that was separated from 14C-adenosine by paper descendant chromatography. The concentration of the total plasma L-homocysteine of 45 normal subjects was 8.04 +/- 0.26 (mean +/- SEM) mumol/l and total L-homocysteine concentration in urine was 0.59 +/- 0.06 mumol/mmol of creatinine (25 subjects). This method is as sensitive than the other methods described in the literature but more rapid and less expensive.

[Molecular genetics: pre- and post-analytic "best practices"]. / Génétique moléculaire: les << bonnes pratiques>> pré-et postanalytieques.

Medical prescriptions for molecular genetic analyses are not yet very common in general practice, neverless they are becoming more and more frequent, and therefore it is more difficult to deal with them in part because of the recent french rules. Laboratory managers are supposed to be able to deal with such requests. This document, describing good laboratory practices, has been elaborated by members of the group "molecular genetics" from the "College National de Biochimie des Hôpitaux", providing details about: assessment of the prescriptions, including patient's consent, choice of the executing laboratory, specimen transmission, assessment and control of clinical and biological data, results transmission, confidentiality, archiving system. Such recommendations should facilitate exchanges with specialized laboratories, being specifically approved for practicing such analyses. The authors draw the attention of laboratory managers to the specificities of such requests.

[RNA isolation and purification methods]. / Les méthodes d'extraction et de purification des ARN.

Recent advances in human, bacterial and viral genome projects and the development of quantitative real-time reverse transcription-polymerase chain reaction methods offer the possibility of analysing a large number of gene transcripts. These molecular developments represent an important advancein the field of genetics, cancer, virology, bacteriology and hematology. A limiting step remains the isolation of high quality mRNA purified from biological samples. This review describes the different methods used to isolate mRNA from biological samples and to verify RNA integrity and gives precise details about RNA storage conditions.

Changing of hepatitis C virus genotype patterns in France at the beginning of the third millenium: The GEMHEP GenoCII Study.

This cross-sectional study aimed to investigate, during a short period between 2000 and 2001, in a large population of patients with chronic hepatitis C, the epidemiological characteristics of hepatitis C virus (HCV) genotypes in France. Data from 26 referral centres, corresponding to 1769 patients with chronic hepatitis C were collected consecutively during a 6-month period. HCV genotyping in the 5'-non-coding region (NCR) was performed in each center using the line probe assay (LiPA, in 63% of cases), sequencing (25%) or primer-specific polymerase chain reaction (PCR) (12%). HCV genotypes 1a, 1b, 2, 3, 4, 5, non-subtyped 1 and mixed infection were found in 18, 27, 9, 21, 9, 3, 11 and 1% of our population, respectively. HCV genotype distribution was associated with gender, age, source and duration of infection, alanine aminotransferase (ALT) levels, cirrhosis, alcohol consumption, hepatitis B virus (HBV) and human immunodeficiency virus (HIV) coinfection. In multivariate analysis, only the source of infection was the independent factor significantly associated with genotype (P = 0.0001). In conclusion, this study shows a changing pattern of HCV genotypes in France, with i.v. drug abuse as the major risk factor, an increase of genotype 4, and to a lesser extent 1a and 5, and a decrease of genotypes 1b and 2. The modification of the HCV genotype pattern in France in the next 10 years may require new therapeutic strategies, and further survey studies.

Long-term folic acid (but not pyridoxine) supplementation lowers elevated plasma homocysteine level in chronic renal failure.

Moderate hyperhomocysteinemia, a risk factor for premature atherosclerosis, is present in chronic uremic patients. We prospectively evaluated the effects of sequential supplementation with pyridoxine (70 mg/day) and folic acid (10 mg/day) for two 3-month periods in 37 nondialyzed patients (29 males) with creatinine clearance (CCr) ranging from 10 to 80 ml/min, whose plasma vitamin B12 and folate level was in the normal range. Mean (+/- SD) baseline plasma total homocysteine (Hcy) was 14.9 +/- 5.2, 16.5 +/- 5.1 and 26.1 +/- 12.1 mumol/l (upper limit in 45 healthy controls 14.1 mumol/l) in patients with CCr 40-80, 20-40 and < 20 ml/min, respectively. Following pyridoxine Hcy did not significantly decrease whereas following folic acid Hcy decreased significantly to 9.9 +/- 2.9 (-33% vs. baseline), 10.3 +/- 3.4 (-37%) and 15.4 +/- 5.5 (-40%), respectively (Student's paired t test, p < 0.001) in the 3 groups. We conclude that folate (but not pyridoxine) pharmacologic supplementation is effective in lowering elevated plasma Hcy in chronic renal failure patients, thus suggesting that enhancing the Hcy remethylation pathway may overcome hyperhomocysteinemia in such patients. In view of the potential atherogenic effects of hyperhomocysteinemia, long-term folate supplementation should be considered in uremic patients.

Increased plasma homocysteine concentration in patients with chronic renal failure.

Since moderate hyperhomocysteinemia is associated with premature occlusive arterial disease, a frequent complication in uremic patients, we prospectively determined fasting plasma concentration of total homocysteine (Hcy) in 79 nondialyzed patients (47 males) with chronic renal failure. None received folate supplementation. Mean (+/- SD) Hcy concentrations were 16.2 +/- 8.1, 23.3 +/- 14.7 and 29.5 +/- 14.4 mumol/l in patient groups with creatinine clearance (Ccr) of, respectively, 30-75, 10-30 and < 10 ml/min, and significantly higher (p < 0.01) than in 45 healthy controls (8.2 +/- 2.2 mumol/l). Linear regression analysis showed a significant negative correlation between Ccr and Hcy (r = 0.40, p < 0.01). Among 37 male patients aged > or = 50 years, Hcy was significantly higher in 15 who had clinical evidence of occlusive arterial disease than in 22 who did not (28.9 +/- 13.3 vs. 17.8 +/- 8.9 mumol/l, p < 0.01). We conclude that hyperhomocysteinemia is present from the early stage of chronic renal failure and may constitute a risk factor for premature arteriosclerosis in uremic patients.

Four novel mutations at the cystathionine beta-synthase locus causing homocystinuria.

We describe four new mutations in the cystathionine beta-synthase gene: three point mutations localized in exons 3, 9 and 10 and one mutation in exon 12 which results in stop codon.

Organic acids in aqueous humour and plasma: post mortem study in infants and diagnosis of enzymopathies.

Organic acids have been determined in aqueous humour and plasma by gas chromatography-mass spectrometry in 38 cases of infant death and 4 cases of inherited metabolic disease: one had a complex fatty-acid oxidation disorder with a large urinary excretion of adipic acid, the others had a disorder of propionate catabolism with a large urinary excretion of methylmalonic acid. In each case we found in aqueous humour the abnormal metabolite present in urine. Thus aqueous humour could be a suitable material for retrospective diagnosis of inherited metabolic diseases at autopsy in sudden infant death syndrome.

aku, a mutation of the mouse homologous to human alkaptonuria, maps to chromosome 16.

Alkaptonuria is a human hereditary metabolic disease characterized by a very high urinary excretion of homogentisic acid, an intermediary product in the metabolism of tyrosine, in association with ochronosis and arthritis. This disease is due to a deficiency in the enzyme homogentisic acid oxidase and is inherited as an autosomal recessive condition. We have found a new recessive mutation (aku) in the mouse that is homologous to human alkaptonuria, during a mutagenesis program with ethylnitrosourea. Affected mice show high levels of urinary homogentisic acid without signs of ochronosis or arthritis. This mutation has been mapped to Chr 16 close to the D16Mit4 locus, in a region of synteny with human 3q.

Antithrombin III activity is not related to plasma homocysteine concentrations.

Arterial and venous thromboembolic events represent frequent and life-threatening complications in homocystinuric patients and are responsible for their early deaths. Reduced levels of antithrombin III activity in homocystinuric patients have recently been reported. So, high plasma L-homocysteine concentration could play a role in the low antithrombin III activity level. In the present study, we have studied the relationship between total plasma homocysteine and inhibitors of blood coagulation levels in 16 patients with malignancies who received bone marrow grafts. There were no correlations between homocysteine values and inhibitors of blood coagulation levels. So, while the defect in amino acid transsulfuration that is responsible for homocystinuria can directly affect the synthesis or activity of some clotting factors, homocysteine concentration is not responsible for this effect.

[Respiratory chain diseases in infancy. Clinical presentation and diagnosis]. / Las enfermedades de la cadena respiratoria en la infancia. Presentación clínica y diagnóstico.

Nineteen children with defects of the mitochondrial respiratory chain are described. First symptoms appeared during the first two years of their lives. Four types of clinical pictures were identified: 1/neonatal hypotonia. 2/cardiomyopathy. 3/progressive neurological deterioration. 4/multisystem disease. A study of pyruvate and fatty acids metabolism and a skeletal muscle biopsy were performed in all cases. Elevation of beta-hydroxybutyrate/acetoacetate and lactate were the most frequent biochemical abnormalities. Muscular biopsy in light microscopy showed in most cases abnormal lipid storage.