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Biomedical subjects

J P Bonnefont

Publications and source records attributed to J P Bonnefont.

At least 19 recordsLinked to original sources
PubMed2006

Prenatal diagnosis of myopathy, encephalopathy, lactic acidosis, and stroke-like syndrome: contribution to understanding mitochondrial DNA segregation during human embryofetal development.

INTRODUCTION: Myopathy, encephalopathy, lactic acidosis, and stroke-like (MELAS) syndrome, a maternally inherited disorder that is among the most common mitochondrial DNA (mtDNA) diseases, is usually associated with the m.3242A>G mutation of the mitochondrial tRNA(leu) gene. Very few data are available with respect to prenatal diagnosis of this serious disease. The rate of mutant versus wild-type mtDNA (heteroplasmy) in fetal DNA is indeed considered to be a poor indicator of postnatal outcome. MATERIALS AND METHODS: Taking advantage of a novel semi-quantitative polymerase chain reaction test for m.3243A>G mutant load assessment, we carried out nine prenatal diagnoses in five unrelated women, using two different fetal tissues (chorionic villi v amniocytes) sampled at two or three different stages of pregnancy. RESULTS: Two of the five women, although not carrying m.3243A>G in blood or extra-blood tissues, were, however, considered at risk for transmission of the mutation, as they were closely related to MELAS-affected individuals. The absence of 3243A>G in the blood of first degree relatives was associated with no mutated mtDNA in the cardiovascular system (CVS) or amniocytes, and their three children are healthy, with a follow-up of 3 months-3 years. Among the six fetuses from the three carrier women, three were shown to be homoplasmic (0% mutant load), the remaining three being heteroplasmic, with a mutant load ranging from 23% to 63%. The fetal mutant load was fairly stable at two or three different stages of pregnancy in CVS and amniocytes. Although pregnancy was terminated in the case of the fetus with a 63% mutant load, all other children are healthy with a follow-up of 3 months-6 years. CONCLUSION: These data suggest that a prenatal diagnosis for MELAS syndrome might be helpful for at-risk families.

Acidosis, Lactic↗
PubMed2004

Novel CFTR mutations in black cystic fibrosis patients.

Cystic fibrosis (CF) is considered as a rare disease in black Africans. In fact, this disease is likely to be underestimated since clinical features consistent with CF diagnosis are often ascribed to environmental factors such as malnutrition. Very little is known about CFTR mutations in affected patients from Central Africa. We report here four novel mutations, i.e., IVS2 + 28 (intron 2), 459T > A (exon 4), EX17a_EX18del (exons 17-18), and IVS22 + IG > A (intron 22), in such patients. An update of CFTR mutations reported in black patients from various ethnies is included. These data might be helpful for genetic counselling regarding CF in black patients.

Black People↗
PubMed2003

Prenatal diagnosis of spinal muscular atrophy by genetic analysis of circulating fetal cells.

Spinal muscular atrophy (SMA) has a prevalence of one in 6000 births and a one in 40 heterozygote frequency. We aimed to develop a routine test for non-invasive prenatal diagnosis. We tested blood with ISET (isolation by size of epithelial tumour or trophoblastic cells) in 12 pregnant women whose babies were at risk of SMA. Using genetic analysis of fetal cells, we identified SMA in all nine isolated from the three mothers carrying an affected child. There was no mutation in any of the 26 fetal cells isolated from the nine women with an unaffected child. Our results show that non-invasive detection of genetic diseases by the analysis of maternal blood is feasible.

Female↗
PubMed2001

Survival of male patients with incontinentia pigmenti carrying a lethal mutation can be explained by somatic mosaicism or Klinefelter syndrome.

Incontinentia pigmenti (IP), or "Bloch-Sulzberger syndrome," is an X-linked dominant disorder characterized by abnormalities of skin, teeth, hair, and eyes; skewed X-inactivation; and recurrent miscarriages of male fetuses. IP results from mutations in the gene for NF-kappaB essential modulator (NEMO), with deletion of exons 4-10 of NEMO accounting for >80% of new mutations. Male fetuses inheriting this mutation and other "null" mutations of NEMO usually die in utero. Less deleterious mutations can result in survival of males subjects, but with ectodermal dysplasia and immunodeficiency. Male patients with skin, dental, and ocular abnormalities typical of those seen in female patients with IP (without immunodeficiency) are rare. We investigated four male patients with clinical hallmarks of IP. All four were found to carry the deletion normally associated with male lethality in utero. Survival in one patient is explained by a 47,XXY karyotype and skewed X inactivation. Three other patients possess a normal 46,XY karyotype. We demonstrate that these patients have both wild-type and deleted copies of the NEMO gene and are therefore mosaic for the common mutation. Therefore, the repeat-mediated rearrangement leading to the common deletion does not require meiotic division. Hypomorphic alleles, a 47,XXY karyotype, and somatic mosaicism therefore represent three mechanisms for survival of males carrying a NEMO mutation.

Alleles↗
PubMed2001

Prenatal diagnosis of respiratory chain deficiency by direct mutation screening.

Respiratory chain deficiency (RCD) is responsible for a clinically heterogeneous group of early-onset untreatable disorders. Enzymological prenatal diagnosis (PD) can only be offered to a fraction of families. Moreover, due to the two-fold genetic origin of the respiratory chain (nuclear and mitochondrial DNA) and owing to the large number of nuclear genes involved in the respiratory chain assembly, maintenance and functioning, the identification of the disease causing gene in a given family remains challenging. Here, we report on PD of RCD by direct screening of NDUFV1, SDH-Fp, SCO1 and SURF1 mutations in five unrelated families with complex I, II and IV deficiency, respectively. The identification of the disease-causing gene in a given family with RCD is a major issue to provide both adequate genetic counselling and early, reliable PD.

Electron Transport↗
PubMed2001

Denaturing high-performance liquid chromatography (DHPLC)-based prenatal diagnosis for tuberous sclerosis.

Tuberous sclerosis (TSC) is a frequent autosomal-dominant condition (affecting 1 in 6000 individuals) caused by various mutations in either the hamartin (TSC1) or the tuberin gene (TSC2). This allelic and non-allelic heterogeneity makes genetic counseling and prenatal diagnosis difficult, especially as a significant proportion of TSC cases are due to de novo mutations. For this reason the identification of the disease causing mutation is mandatory for accurate counseling, yet current mutation detection methods such as single-strand conformation polymorphism (SSCP) or denaturing gradient gel electrophoresis (DGGE) are labor intensive with limited detection efficiency. Denaturing high-performance liquid chromatography (DHPLC) is a high-throughput, semi-automated mutation detection system with a reported mutation detection rate close to 100% for PCR fragments of up to 800 bp. We used a recently described DHPLC assay allowing the efficient detection of mutations in TSC1 to analyze the DNA extracted from a chorion villus sample in order to perform a prenatal diagnosis for TSC. The fetus was found not to have inherited the deleterious mutation and the DHPLC diagnosis was confirmed by haplotype analysis. This represents the first DHPLC-based prenatal diagnosis of a genetic disease.

Chorionic Villi↗
PubMed2001

Hepatic carnitine palmitoyl transferase 1 (CPT1 A) deficiency in North American Hutterites (Canadian and American): evidence for a founder effect and results of a pilot study on a DNA-based newborn screening program.

We describe six patients with hepatic carnitine palmitoyl transferase (CPT1 A) deficiency who are members of a large extended Hutterite kindred living in widely scattered communities in the United States and Canadian Prairies. Two patients have significant neurological impairment due to severe recurrent hypoglycemic crises. The remaining four patients with earlier detection and treatment have near normal outcomes. The Canadian and American Hutterite families share two common ancestors who married in 1812, about 60 years before the Hutterites arrived in North America and prior to their subdivision into the three groups (Schmiedeleut, Dariusleut, and the Lehrerleut). These patients share a common haplotype on chromosome 11q13 and are all homozygous for a common CPT1 A G710E mutation, suggesting a founder effect. The clustering of such a rare disorder of fatty acid oxidation prompted us to initiate a pilot DNA-based neonatal screening program to determine the carrier frequency of this mutation in Hutterite newborns with the participation and support of the community. To date our carrier frequency is 1/16, close to the predicted frequency based on diagnosed patients and number of births. We believe our newborn screening program for CPT1 A deficiency in the Hutterite community will serve as a prototype model for delivery of targeted genetic services to other similar unique genetic isolates.

Adolescent↗
PubMed2001

Molecular and enzymatic characterization of a unique carnitine palmitoyltransferase 1A mutation in the Hutterite community.

Hepatic carnitine palmitoyltransferase 1 (CPT1A) deficiency is a rare disorder of mitochondrial fatty acid oxidation inherited as an autosomal recessive trait. Symptomatology comprises attacks of hypoketotic hypoglycemia with risk of sudden death or neurological sequelae. Only one CPT1A mutation has been reported so far. Identification of the disease-causing mutations allows both insights into the structure-function relationships of CPT1A and management of the patients and their relatives. The molecular analysis of CPT1A deficiency in a large Hutterite kindred illustrates this point. Both cDNA and genomic DNA analysis demonstrate that the affected patients are homozygous for a 2129G>A mutation predicting a G710E substitution. Studies in fibroblasts from one patient as well as heterologous expression of the mutagenized CPT1A in yeast show that the G710E mutation alters neither mitochondrial targeting nor stability of the CPT1A protein. By contrast, kinetic studies conclusively establish that the mutant CPT1A is totally inactive, indicating that the G710E mutation dramatically impairs the catalytic function of CPT1A. Finally, due to a strongly suspected founder effect for the origin of CPT1A deficiency in this Hutterite kindred, identification of this disease-causing mutation allows the setup of a targeted DNA-based newborn screening in this at-risk population.

Amino Acid Sequence↗
PubMed2001

MECP2 mutation in non-fatal, non-progressive encephalopathy in a male.

To study the clinical overlap between Rett (RTT) and Angelman syndromes (AS), we screened the MECP2 gene in a cohort of 78 patients diagnosed as possible AS but who showed a normal methylation pattern at the UBE3A locus. MECP2 missense (R106W, G428S), nonsense (R255X, R270X), and frameshift mutations (803 delG) were identified in 6/78 patients including 4/6 female cases consistent with RTT, one female case with progressive encephalopathy of neonatal onset, and one isolated male case with non-fatal, non-progressive encephalopathy of neonatal onset. This study shows that MECP2 mutations can account for a broad spectrum of clinical presentations and raises the difficult issue of the screening of the MECP2 gene in severe encephalopathy in both males and females.

Angelman Syndrome↗
PubMed2000

Mutations of the SCO1 gene in mitochondrial cytochrome c oxidase deficiency with neonatal-onset hepatic failure and encephalopathy.

Cytochrome c oxidase (COX) catalyzes both electron transfer from cytochrome c to molecular oxygen and the concomitant vectorial proton pumping across the inner mitochondrial membrane. Studying a large family with multiple cases of neonatal ketoacidotic comas and isolated COX deficiency, we have mapped the disease locus to chromosome 17p13.1, in a region encompassing two candidate genes involved in COX assembly-namely, SCO1 and COX10. Mutation screening revealed compound heterozygosity for SCO1 gene mutations in the patients. The mutated allele, inherited from the father, harbored a 2-bp frameshift deletion (DeltaGA; nt 363-364) resulting in both a premature stop codon and a highly unstable mRNA. The maternally inherited mutation (C520T) changed a highly conserved proline into a leucine in the protein (P174L). This proline, adjacent to the CxxxC copper-binding domain of SCO1, is likely to play a crucial role in the tridimentional structure of the domain. Interestingly, the clinical presentation of SCO1-deficient patients markedly differs from that of patients harboring mutations in other COX assembly and/or maturation genes.

Age of Onset↗
PubMed2000

First specific preimplantation genetic diagnosis for ornithine transcarbamylase deficiency.

Ornithine transcarbamylase (OTC) deficiency is an X-linked dominant metabolic disorder with partial penetrance in heterozygous females. Affected boys usually die from hyperammonemia in the first few days of life, while clinical expression in carrier females ranges from no symptoms to neonatal death. A young couple whose boy had died of OTC deficiency in the neonatal period was referred to our genetic department for their subsequent pregnancy. The fetus was found to be affected, and after genetic counseling the pregnancy was terminated. Prenatal diagnosis of the third pregnancy identified a heterozygous female, who died after a normal birth at age 11 days from hyperammonemia. After this, the couple asked for preimplantation genetic diagnosis (PGD). We have developed a duplex nested PCR assay allowing the amplification of both the mutation and an informative restriction fragment length polymorphism (RFLP) located in the 3' end of the OTC gene. After nested amplification, allele identification was carried out for both loci by double restriction digestion and electrophoresis gel analysis. The co-amplification of both loci provided a means of detecting potential allele dropout or incomplete digestion. Two PGD cycles were carried out, a total of 14 embryos were analysed and a diagnosis could be obtained in 13/14 embryos. There were four unaffected male embryos, four heterozygous females and four unaffected females; the final embryo was an affected one of undetermined gender. In both cycles, three unaffected embryos could be transferred early on Day 4 post-insemination. The second cycle resulted in the birth of a baby boy devoid of the OTC mutation. This constitutes the first birth following PGD carried out by a French team.

Base Sequence↗
PubMed2000

Spectrum of CFTR mutations in cystic fibrosis and in congenital absence of the vas deferens in France.

We have collated the results of cystic fibrosis (CF) mutation analysis conducted in 19 laboratories in France. We have analyzed 7, 420 CF alleles, demonstrating a total of 310 different mutations including 24 not reported previously, accounting for 93.56% of CF genes. The most common were F508del (67.18%; range 61-80), G542X (2.86%; range 1-6.7%), N1303K (2.10%; range 0.75-4.6%), and 1717-1G>A (1.31%; range 0-2.8%). Only 11 mutations had relative frequencies >0. 4%, 140 mutations were found on a small number of CF alleles (from 29 to two), and 154 were unique. These data show a clear geographical and/or ethnic variation in the distribution of the most common CF mutations. This spectrum of CF mutations, the largest ever reported in one country, has generated 481 different genotypes. We also investigated a cohort of 800 French men with congenital bilateral absence of the vas deferens (CBAVD) and identified a total of 137 different CFTR mutations. Screening for the most common CF defects in addition to assessment for IVS8-5T allowed us to detect two mutations in 47.63% and one in 24.63% of CBAVD patients. In a subset of 327 CBAVD men who were more extensively investigated through the scanning of coding/flanking sequences, 516 of 654 (78. 90%) alleles were identified, with 15.90% and 70.95% of patients carrying one or two mutations, respectively, and only 13.15% without any detectable CFTR abnormality. The distribution of genotypes, classified according to the expected effect of their mutations on CFTR protein, clearly differed between both populations. CF patients had two severe mutations (87.77%) or one severe and one mild/variable mutation (11.33%), whereas CBAVD men had either a severe and a mild/variable (87.89%) or two mild/variable (11.57%) mutations.

Adult↗
PubMed2000

Mutation analysis of the hamartin gene using denaturing high performance liquid chromatography.

Denaturing high performance liquid chromatography (DHPLC) is a novel high-capacity technique for gene mutation scanning. We have assessed the sensitivity and specificity of this method for analysis of the full coding sequence of the hamartin (TSC1) gene in 20 tuberous sclerosis patients, whose TSC1 genes previously had been studied by single strand conformation polymorphism analysis and protein truncation assay. All eight sequence variants previously identified were adequately detected by DHPLC. Additionally, this approach picked up three polymorphisms, one of which (IVS13-55 C>G) was hitherto unreported, therefore serving as proof of principle for this technique. Thus, DHPLC appears to be a highly sensitive method with advantages in terms of flexibility, fragments size analysis, cost and time and labor sparing, compared to classical approaches of mutation scanning.

Chromatography, High Pressure Liquid↗
PubMed2000

Isolated supravalvular aortic stenosis: functional haploinsufficiency of the elastin gene as a result of nonsense-mediated decay.

We have used single-strand conformation and heteroduplex analyses of genomic amplimers to identify point mutations within the elastin gene (ELN) in patients with non-syndromic supravalvular aortic stenosis (SVAS) from a total of eight unrelated families. Six novel point mutations were identified. We have collected detailed clinical information on mutation carriers and demonstrated significant non-penetrance in some of the families. Together with the new mutations described here, 14 point mutations have been reported in SVAS patients, and 10 of these result in premature stop codons (PTCs). We have analyzed the expression of ELN alleles in skin fibroblasts from one SVAS patient and shown that PTC mutations indeed result in selective elimination of mutant transcripts. Inhibition of the nonsense-mediated decay mechanism by cycloheximide resulted in the stabilization of mutant elastin mRNA. Allelic inactivation by the ELN mutation in this patient led to an overall decrease of the steady state levels of elastin mRNA. Finally, we have demonstrated reduced synthesis and secretion of tropoelastin by skin fibroblasts from the same SVAS patient. We conclude that PTC mutations in ELN result in nonsense-mediated decay of mutant mRNA in this patient. Given the predominance of PTC mutations in SVAS, we suggest that functional haploinsufficiency may be a pathomechanism underlying most cases of non-syndromic SVAS.

Adolescent↗