Biomedical subjects

J Pié

Publications and source records attributed to J Pié.

11 recordsLinked to original sources

PubMed1998

Two missense point mutations in different alleles in the 3-hydroxy-3-methylglutaryl coenzyme A lyase gene produce 3-hydroxy-3-methylglutaric aciduria in a French patient.

Two novel point mutations in the 3-hydroxy-3-methylglutaryl coenzyme A lyase gene were found in a French patient with double heterozygous 3-hydroxy-3-methylglutaric aciduria. Amplification by reverse transcriptase-polymerase chain reaction of the mRNA using five different pairs of oligonucleotides produced no differences in the fragments amplified with respect to the control. Single-strand conformation polymorphism analysis showed that only one amplified fragment was different in the patient vs. control. Sequencing of the amplified fragments showed two missense point mutations, A698G and T788C, each of them mixed with the wild-type sequence. These mutations produced the changes H233R and L263P, leading to changes in the enzyme activity, which was largely abolished. The father and one brother of the proband were heterozygous for the L263P mutation and the mother and one daughter were heterozygous for the H233R mutation, which confirms the double-heterozygous character of the patient. Another sibling was free of the mutations. An enzymatic restriction analysis has been proposed to screen the occurrence of these two mutations in future patients.

Alleles↗

PubMed1998

A nonsense mutation in the exon 2 of the 3-hydroxy-3-methylglutaryl coenzyme A lyase (HL) gene producing three mature mRNAs is the main cause of 3-hydroxy-3-methylglutaric aciduria in European Mediterranean patients.

3-Hydroxy-3-methylglutaric aciduria is a rare recessive monogenic disorder that affects ketogenesis and the catabolism of L-leucine. We report the biochemical and molecular characterization of a mutation in the 3-hydroxy-3-methylglutaryl coenzyme A lyase gene in four new probands, three Spanish and one Turkish, affected by 3-hydroxy-3-methylglutaric aciduria, all homozygous for the nonsense mutation Glu37Ter, which was reported by our group in two probands of Portuguese and Moroccan origin (15). In addition to the aberrant mRNAs found in the two previous probands, a novel species of mature HL mRNA was observed in the patients studied here, since a new cDNA, skipped in exons 2 and 3, was obtained from the mRNAs by reverse-transcription PCR (RT-PCR). Thus, three mRNA species were produced in aberrant splicings as a result of this nonsense mutation: (i) one of the expected size that contains the premature stop codon UAA, (ii) another with a deletion of 84 bp corresponding to the whole of exon 2, and (iii) a new species found now, with a deletion of 192 bp corresponding to skipping of the whole of exons 2 and 3, whose translation product led to the loss of seven amino acids in the leader peptide and 57 amino acids in the terminal domain of the mature enzyme. The association of a nonsense mutation with the skipping of the exon that contains it, plus the following exon, is an unusual finding not seen previously in HL deficiencies. The mutation described here shows the highest incidence (> 37%) of total HL deficiencies reported.

Child, Preschool↗

PubMed1997

A nonsense mutation in the 3-hydroxy-3-methylglutaryl-CoA lyase gene produces exon skipping in two patients of different origin with 3-hydroxy-3-methylglutaryl-CoA lyase deficiency.

A novel nonsense mutation associated with the skipping of constitutive exon 2 of the 3-hydroxy-3-methylglutaryl-CoA lyase gene was found in two patients, from Portugal and Morocco, with 3-hydroxy-3-methylglutaric acidemia. By reverse transcriptase PCR and single-strand conformational polymorphism a G-T transversion was located, at nucleotide 109, of the 3-hydroxy-3-methylglutaryl-CoA lyase cDNA, within exon 2. Two mRNAs were produced as a result of this nonsense mutation: one of the expected size that contains the premature stop codon UAA, and the other with a deletion of 84 bp corresponding to the whole of exon 2. This deletion produced the loss of the last seven amino acids of the leader peptide and the first 21 amino acids of the mature protein. The nonsense mutation was found in a purine-rich GGAAG sequence, which is equal to, or similar to, others reported to be exonic splicing enhancers (ESE). We suggest that the nonsense mutation may affect a possible ESE on exon 2, which would hinder the splice site selection and facilitate an aberrant splice with the skipping of this exon. Determination by quantitative PCR shows that the ratio of mRNA with the nonsense mutation to the mRNA with the deletion is approx. 3:1.

Amino Acid Sequence↗

PubMed1997

A two-base deletion in exon 6 of the 3-hydroxy-3-methylglutaryl coenzyme A lyase (HL) gene producing the skipping of exons 5 and 6 determines 3-hydroxy-3-methylglutaric aciduria.

A novel two-base deletion in the 3-hydroxy-3-methylglutaryl coenzyme A lyase (HL) gene was found in a Spanish patient with homozygous 3-hydroxy-3-methylglutaric aciduria. Amplification by RT-PCR of the mRNAs showed that the gene was transcribed into three different mRNAs. One showed the complete deletion of exons 5 and 6 located between nucleotides 348 and 561 of the HL cDNA. The second transcript showed deletion of exon 6 only, and the third contained a two-base deletion CT in exon 6, corresponding to nucleotides 504 and 505 of the HL cDNA. These aberrant mRNAs are predicted to encode three abnormal HMG-CoA lyase proteins; the first (from skipped exons 5 and 6) lacks 71 amino acids, which represents 24% of the mature protein; the second, (from the skipping of exon 6, producing a frameshift) contains only 192 amino acids, the last 26 of which are missense amino acids preceding a stop codon; the third contains only 175 amino acids, the last 7 of which are missense. Northern blot analysis showed that the HL mRNA levels of the patient were 4% of the control. PCR quantitative analysis indicated that the mRNA lacking exons 5 and 6 was the most abundant, representing 88% of the total. The other two mRNAs represented 8% and 4%, respectively. In the genomic DNA only one CT deletion was found at positions +7 and +8 at beginning of exon 6. No mutations were observed in the splice donor, splice acceptor, or pyrimidine-rich sequences of the intronic regions flanking exons 5 and 6. All three aberrant mRNAs resulted only from the deletion of nucleotides CT. We suggest that this deletion may affect the interaction between the small nuclear ribonucleoproteins (snRNPs) and exon 6, and that, as a result, the abnormal splicing of the pre-mRNA produces two different aberrant transcripts.

Adolescent↗

PubMed1996

Aberrantly spliced mRNAs of the 3-hydroxy-3-methylglutaryl coenzyme A lyase (HL) gene with a donor splice-site point mutation produce hereditary HL deficiency.

A novel point mutation in the 3-hydroxy-3methyl-glutaryl coenzyme A lyase gene was found in a Turkish patient with homozygous 3-hydroxy-3-methylglutaric acidemia. Amplification by RT-PCR of the mRNA using a six different pairs of oligonucleotides produced no differences in four of the fragments amplified with respect to the control, but generated two fragments of different size. One was representative of a deletion of 126 bp and the other of an insertion of 78 bp. These abnormal mRNAs resulted from a G-->C transversion at the nucleotide +1 of an intron, which changed the invariant GT dinucleotide of the 5' donor splice site. This was associated with the occurrence of an alternative splicing, which led to the skipping of the whole exon of 126 bp, and also with the activation of one cryptic donor splice site in the same intron. These aberrant spliced mRNAs are predicted to encode two abnormal HMG-CoA lyase proteins: the first results in a protein with an internal deletion of 42 amino acids, whose enzyme activity is largely abolished, as the catalytic site was completely removed; the second contains 17 missense amino acids that precede a stop codon. Northern blot analysis showed that the overall content of these aberrantly spliced mRNAs in proband fibroblasts was the same as that found in control fibroblasts. However, hardly any transcript was observed corresponding to the inserted mutated mRNA when it was examined by a specific probe. To quantify the relative proportion of the two mRNAs, a quantitative RT-PCR (the DNA-mimic PCR reaction) was carried out. Results show that the proportion of the inserted mRNAs with respect to the deleted mRNA is only 1.2%. The father, mother, and two brothers of the proband were heterozygous in the G-->C mutation in the +1 nucleotide of the intron considered, while the two alleles of another brother were free of the mutation.

Base Sequence↗

PubMed1993

[Hematic parameters in chronic hypoxemia].

The hematic effects of hypoxia have been studied in 184 patients divided in four groups: 1) 10 patients with pO2 values in capillary blood lower than 43 mmHg; 2) 40 patients with pO2 between 43 and 50 mmHg; 3) 134 subjects with pO2 between 51 and 62 mmHg; and 4) 39 normal subjects (control). Hypoxemic subjects show higher erythrocyte concentrations than normoxemic ones, but without reaching the point of polycythemia and without significant differences in the hemoglobin concentration. The values of the mean corpuscular volume, the mean corpuscular hemoglobin and the concentration of the mean corpuscular hemoglobin show no significant differences between the groups. The amplitude of the red corpuscle distribution has not shown statistically significant differences between the groups of more pronounced hypoxia and those of slight hypoxia and control. The hematocrit, determined in the autoanalyzer, does not show any significant differences between the groups, whereas the one obtained by centrifugation is higher in the hypoxemic groups. Differences are non-significant in platelet count, mean platelet volume, and platelet distribution among the groups. The leucocyte count yields progressively higher values as blood pO2 decreases, with an accompanying increase in the percentage of neutrophyle granulocytes and a decrease in lymphocytes. Leucocytes with peroxidasic activity are significantly higher when the oxygen offer is lower.

Adult↗

PubMed1989

[Differences of males and females regarding the binding capacity of insulin to erythrocyte receptors].

The analysis of insulin receptors in erythrocytes demands a relatively small blood sample, which justifies the interest in its use as an index of the cellular capacity for binding hormone. In order to establish criteria for normalcy, the capacity of erythrocytes for binding in vitro insulin labelled with 125I before increasing concentrations of cold insulin (from 0.5 to 10(3) ng/ml), was studied in a group of 41 healthy men and another of 35 women with normal menstrual cycles. In the female group the study was carried out in three different days of the same cycle (days 3, 12 and 21). The binding capacity in the male was higher than in the female (p less than 0.05) in the follicular phase (days 3 and 12) as well as in the luteal phase (day 21) and, among women, it was higher in the follicular phase than in the luteal one (p less than 0.05). The results indicate that progesterone, as well as prolactin and glucagon, may play an important role in the binding capacity of insulin to its receptor. To make the values comparable, it is suggested that blood extraction in women be carried out during the first five days of the cycle.

Adult↗

PubMed1988

Urinary triiodothyronine excretion.

The purpose of this work was to estimate the 24 h urinary excretion of free and conjugated triiodotironine (T3) using a direct radioimmunoassay and enzyme hydrolysis. Mean urinary values of free and total T3 (mean +/- 1 SD) in euthyroid controls were 2074 +/- 673 and 2819 +/- 809 pmol/24 h respectively. In patients with hyperthyroidism, values of free hormone were about 4.2 times higher than the mean value of the euthyroid controls, and about one-third in patients with hypothyroidism. These results show this measurement to be useful as an indicator of thyroid function. Mean renal clearance of free T3 was 211.6 +/- 62.8 ml/min (mean +/- 1 SD) in euthyroid controls, 260.8 +/- 87.5 ml/ml in hyperthyroid patients and 229 +/- 98.7 ml/min in hypothyroid patients. The data show that T3 renal clearance is, in all cases, greater than glomerular filtration rate, suggesting tubular secretion of T3.

Adult↗

PubMed1988

[Effects of hypoxia on plasma concentrations and daily elimination of sodium, potassium and chlorine ions].

Serum concentrations of Na+, K+ and Cl- are studied, as well as the elimination of these ions in urine, in patients suffering from chronic respiratory insufficiency, being classified in two groups according to the level of hypoxemia: group A (PO2 less than 6.66 KPa) and group B (PO2 less than 8 KPa). A third group C of healthy patients with analogous anthropological characteristics has served as a control group. The concentrations of serum of the three ions are noticeably similar in the three groups, but the daily elimination of Na+, K+ and Cl- is less in those suffering from respiratory insufficiency than in those of the control group, with significant statistical differences in all cases except with K+ in those suffering from pronounced hypoxemia. On analysing the correlation between the rates of elimination of ions in urine, with the plasmatic values of PO2, PCO2 and [H+] of all the patients studied, the highest values of the Pearson coefficient are found on correlating the elimination of ions with the partial pressures of oxygen, therefore suggesting that hypoxia could be the main motor inducing metabolic changes.

Aged↗

PubMed1986

Effects of chronic hypoxia on kidney function.

Renal disfunctions which appear in the chronic respiratory insufficient patient are analysed, as well as the participation of the arterial blood hypoxemia in their genesis. Renal clearances of Na, K, Cl, Ca, Mg and Pi, and those of urea and creatinine, were lower in 36 patients having chronic hypoxemia than in 15 normosemic controls, showing significant statistical differences for Na, K, Cl, Ca and urea. The correlations between the clearances of these substances and the pO2 arterial blood levels had a greater statistical significance than can be established with pCO2 or [H+] levels. Thus, the existence of a causal dependency between renal disfunction and hypoxemia may be deduced.

Blood Gas Analysis↗

PubMed1984

Valuation of protein metabolism and albumin in patients submitted to peripheral parenteral nutrition (PPN).

A protein breakdown study was carried out in ten post-operative patients, 24 hours after operation and after five days of peripheral parenteral nutrition; five of these patients were in a good nutritive condition (group A) and five were suffering from malnutrition and were critically ill (group B). Results indicate that the administration of amino-acids achieved a decrease in the degree of protein breakdown in the two groups under study, and also it was observed that better use was made of the nitrogen administered, particularly by the liver when the nitrogenous supply is completed with carbohydrates such as glucose. The difference obtained in values between the oxidized leucine and the leucine administered suggests that there is an endogenous donor pool, and the differences existing between breakdown and leucine oxidation values indicate that in group A, nourished solely with amino-acids, breakdown is lesser and oxidation greater than in group B, nourished with amino-acids and glucose, which suggests that there is more stimulation to re-use the amino-acids in this group.

Amino Acids↗