Increased incidence of obstetric complications in women carrying mitochondrial DNA mutations: a retrospective cohort study in a single tertiary centre.

OBJECTIVE: To investigate the obstetric outcome of women carriers of the oxidative phosphorylation (OXPHOS) disorder mutation. DESIGN: A retrospective cohort study in a single tertiary centre. SETTING: A review of the obstetric history of women referred for prenatal screening of a mitochondrial disorder was performed. POPULATION: Women were divided into three groups: (1) women carrying mitochondrial DNA (mtDNA) mutations; (2) healthy women with a family history of mtDNA-related OXPHOS disorder; and (3) healthy women carrying heterozygote nuclear DNA mutations. METHODS: Obstetric history and pregnancy complications were evaluated separately in the three groups and compared with the control group. MAIN OUTCOME MEASURES PREGNANCY COMPLICATIONS. RESULTS: Seventy-five women were included with 287 cumulative pregnancies. Groups 1 and 3 had a significantly greater proportion of terminations of pregnancy (20 and 13% versus 0.8%, P < 0.001), and a lower percentage of live births (52 and 72% versus 87%, P = 0.001), compared with controls. Apart from this, the rate of obstetric complications in group 3 did not differ from the controls. The obstetric history of women in group 1 was marked by higher rates of early miscarriages (26 versus 11%, P = 0.004), gestational diabetes (14 versus 3%, P = 0.02), intrauterine growth restriction (IUGR, 10 versus 1%, P = 0.008), and postpartum haemorrhage than were reported for controls (12 versus 2%, P = 0.01). CONCLUSION: Women who are heteroplasmic for OXPHOS mutations have a higher incidence of pregnancy losses, gestational diabetes, IUGR, and post postpartum haemorrhage. TWEETABLE ABSTRACT: Women heteroplasmic for mitochondrial DNA mutations have a higher incidence of obstetric complications, compared with the control group.

Brain imaging in mitochondrial respiratory chain deficiency: combination of brain MRI features as a useful tool for genotype/phenotype correlations.

Mitochondrial diseases are characterised by a broad clinical and genetic heterogeneity that makes diagnosis difficult. Owing to the wide pattern of symptoms in mitochondrial disorders and the constantly growing number of disease genes, their genetic diagnosis is difficult and genotype/phenotype correlations remain elusive. Brain MRI appears as a useful tool for genotype/phenotype correlations. Here, we summarise the various combinations of MRI lesions observed in the most frequent mitochondrial respiratory chain deficiencies so as to direct molecular genetic test in patients at risk of such diseases. We believe that the combination of brain MRI features is of value to support respiratory chain deficiency and direct molecular genetic tests.

Phenotype and genotype in 101 males with X-linked creatine transporter deficiency.

BACKGROUND: Creatine transporter deficiency is a monogenic cause of X-linked intellectual disability. Since its first description in 2001 several case reports have been published but an overview of phenotype, genotype and phenotype--genotype correlation has been lacking. METHODS: We performed a retrospective study of clinical, biochemical and molecular genetic data of 101 males with X-linked creatine transporter deficiency from 85 families with a pathogenic mutation in the creatine transporter gene (SLC6A8). RESULTS AND CONCLUSIONS: Most patients developed moderate to severe intellectual disability; mild intellectual disability was rare in adult patients. Speech language development was especially delayed but almost a third of the patients were able to speak in sentences. Besides behavioural problems and seizures, mild to moderate motor dysfunction, including extrapyramidal movement abnormalities, and gastrointestinal problems were frequent clinical features. Urinary creatine to creatinine ratio proved to be a reliable screening method besides MR spectroscopy, molecular genetic testing and creatine uptake studies, allowing definition of diagnostic guidelines. A third of patients had a de novo mutation in the SLC6A8 gene. Mothers with an affected son with a de novo mutation should be counselled about a recurrence risk in further pregnancies due to the possibility of low level somatic or germline mosaicism. Missense mutations with residual activity might be associated with a milder phenotype and large deletions extending beyond the 3' end of the SLC6A8 gene with a more severe phenotype. Evaluation of the biochemical phenotype revealed unexpected high creatine levels in cerebrospinal fluid suggesting that the brain is able to synthesise creatine and that the cerebral creatine deficiency is caused by a defect in the reuptake of creatine within the neurones.

Causes, diagnosis and therapy of common diseases in neonatal puppies in the first days of life: cornerstones of practical approach.

Neonatal diseases and losses are a common and often unavoidable problem within breeding kennels. Altogether, morbidity and mortality ranges, according to the literature, from 5 to 35%. Among non-infectious causes besides hypoxia during birth, hypothermia, hypoglycaemia and dehydration are mostly responsible for puppy diseases and losses. Approximately 90% of all deaths in hypoxaemic pups occur during the first 2 days. Of 183 pups with hypoxia, 63 died, 92.7% of them within 48 h after birth. Among infectious causes, bacterial infection is the most common cause of neonatal mortality. Escherichia coli, streptococci, staphylococci, Pseudomonas sp., Klebsiella sp., Enterobacter sp. and some other micro-organisms are regularly involved in neonatal infections. Post-mortem findings especially document E. coli, Staphylococcus sp. and Streptococcus sp. as responsible bacteria. The dam and the environment are suspected as sources of neonatal infections as it was shown by genetic relatedness of responsible bacterial strains isolated in both puppies and their dams. From a total of 517 puppies with bacterial infections, the treatment results documented that parenteral administration of amoxicillin/clavulanic acid in 308 neonates showed the best result. Diagnosis of diseases is often made difficult by the absence of variability in clinical signs contrary to adult dogs. Findings during a physical examination in pups differ from those in adults. Furthermore, treatment recommendations have to meet the special conditions in neonates concerning drug metabolism and excretion.

[Aspect of brain MRI in mitochondrial respiratory chain deficiency. A diagnostic algorithm of the most common mitochondrial genetic mutations]. / Aspect en IRM cérébrale des maladies mitochondriales. Algorithme décisionnel des maladies mitochondriales les plus fréquentes.

Mitochondrial diseases are due to deficiency of the respiratory chain and are characterized by a broad clinical and genetic heterogeneity that makes diagnosis difficult. Some clinical presentations are highly suggestive of given gene mutations, allowing rapid genetic diagnosis. However, owing to the wide pattern of symptoms in mitochondrial disorders and the constantly growing number of disease genes, their genetic diagnosis is frequently difficult and genotype/phenotype correlations remain elusive. For this reason, brain MRI appears as a useful tool for genotype/phenotype correlations. Here, we report the most frequent neuroradiological signs in mitochondrial respiratory chain deficiency and we propose a diagnostic algorithm based on neuroimaging features, so as to direct molecular genetic tests in patients at risk of mitochondrial respiratory chain deficiency. This algorithm is based on the careful analysis of five areas on brain MRI: (1) basal ganglia (hyperintensities on T2 or calcifications); (2) cerebellum (hyperintensities on T2 or atrophy); (3) brainstem (hyperintensities on T2 or atrophy); (4) white matter (leukoencephalopathy); (5) cortex (sub-tentorial atrophy); (6) stroke-like episodes. We believe that the combination of brain MRI features is of value to support respiratory chain deficiency and direct molecular genetic tests.

A constant and similar assembly defect of mitochondrial respiratory chain complex I allows rapid identification of NDUFS4 mutations in patients with Leigh syndrome.

Isolated complex I deficiency is a frequent cause of respiratory chain defects in childhood. In this study, we report our systematic approach with blue native PAGE (BN-PAGE) to study mitochondrial respiratory chain assembly in skin fibroblasts from patients with Leigh syndrome and CI deficiency. We describe five new NDUFS4 patients with a similar and constant abnormal BN-PAGE profile and present a meta-analysis of the literature. All NDUFS4 mutations that have been tested with BN-PAGE result in a constant and similar abnormal assembly profile with a complete loss of the fully assembled complex I usually due to a truncated protein and the loss of its canonical cAMP dependent protein kinase phosphorylation consensus site. We also report the association of abnormal brain MRI images with this characteristic BN-PAGE profile as the hallmarks of NDUFS4 mutations and the first founder NDUFS4 mutations in the North-African population.

Monozygotic twins discordant for submicroscopic chromosomal anomalies in 2p25.3 region detected by array CGH.

Although discordant phenotypes in monozygotic twins with developmental disorder are not an exception, underlying genetic discordance is rarely reported. Here, we report on the clinical and cytogenetic details of 4-year-old female monozygotic twins with discordant phenotypes. Twin 1 exhibited global developmental delay, overweight and hyperactivity. Twin 2 had an autistic spectrum disorder. Molecular karyotyping in twin 1 identified a 2p25.3 deletion, further confirmed by Fluorescence in situ hybridization (FISH) analysis on leukocytes. Interestingly, array comparative genomic hybridization was normal in twin 2 but FISH analysis using the same probe as twin 1 showed mosaicism with one-third of cells with a 2p25.3 deletion, one-third of cells with a 2p25.3 duplication, and one-third of normal cells. Genotyping with microsatellite markers confirmed the monozygosity of the twins. We propose that the chromosome imbalance may be due to a mitotic non-allelic recombination occurring during blastomeric divisions of a normal zygote. Such event will result in three distinct cell populations, whose proportion in each embryo formed after separation from the zygote may differ, leading to discordant chromosomal anomalies between twins. We also discuss that the MYTL1L and the SNTG2 genes within the reported region could probably relate to the phenotypic discordance of the monozygotic twins.

A novel microdeletion syndrome at 3q13.31 characterised by developmental delay, postnatal overgrowth, hypoplastic male genitals, and characteristic facial features.

BACKGROUND: Congenital deletions affecting 3q11q23 have rarely been reported and only five cases have been molecularly characterised. Genotype-phenotype correlation has been hampered by the variable sizes and breakpoints of the deletions. In this study, 14 novel patients with deletions in 3q11q23 were investigated and compared with 13 previously reported patients. METHODS: Clinical data were collected from 14 novel patients that had been investigated by high resolution microarray techniques. Molecular investigation and updated clinical information of one cytogenetically previously reported patient were also included. RESULTS: The molecular investigation identified deletions in the region 3q12.3q21.3 with different boundaries and variable sizes. The smallest studied deletion was 580 kb, located in 3q13.31. Genotype-phenotype comparison in 24 patients sharing this shortest region of overlapping deletion revealed several common major characteristics including significant developmental delay, muscular hypotonia, a high arched palate, and recognisable facial features including a short philtrum and protruding lips. Abnormal genitalia were found in the majority of males, several having micropenis. Finally, a postnatal growth pattern above the mean was apparent. The 580 kb deleted region includes five RefSeq genes and two of them are strong candidate genes for the developmental delay: DRD3 and ZBTB20. CONCLUSION: A newly recognised 3q13.31 microdeletion syndrome is delineated which is of diagnostic and prognostic value. Furthermore, two genes are suggested to be responsible for the main phenotype.

Correlation between severity and SMN protein level in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a common autosomal recessive neuromuscular disorder characterized by degeneration of motor neurons of the spinal cord. Three different forms of childhood SMA have been recognized on the basis of age at onset and clinical course: Werdnig-Hoffmann disease (type-1), the intermediate form (type-II) and Kugelberg-Welander disease (type-III). A gene termed 'survival of motor neuron' (SMN) has been recognized as the disease-causing gene in SMA. SMN encodes a protein located within a novel nuclear structure and interacts with RNA-binding proteins. To elucidate the molecular mechanism underlying the pathogenesis of the disease, we examined the expression of the SMN gene in both controls and SMA patients by western blot and immunohistochemical analyses using antibodies raised against the SMN protein. The present study shows a marked deficiency of the SMN protein in SMA.

Autosomal dominant familial spastic paraplegia is genetically heterogeneous and one locus maps to chromosome 14q.

Autosomal dominant familial spastic paraplegia (FSP) is a degenerative disorder of unknown aetiology characterized by a progressive spasticity of the legs. Three families with autosomal dominant FSP of early onset were analysed in linkage studies using highly polymorphic microsatellite markers. Close linkage to a group of markers on chromosome 14q (maximum multipoint lodscore z = 10) was observed in one family. This chromosome 14q candidate region was entirely excluded in the two other families, providing evidence of genetic heterogeneity within a homogeneous clinical form of FSP.

Mutation of the endothelin-3 gene in the Waardenburg-Hirschsprung disease (Shah-Waardenburg syndrome).

Hirschsprung disease (HSCR) and Waardenburg sundrome (WS) are congenital malformations regarded as neurocristopathies since both disorders involve neural crest-derived cells. The WS-HSCR association (Shah-Waardenburg syndrome) is a rare autosomal recessive condition that occasionally has been ascribed to mutations of the endothelin-receptor B (EDNRB) gene. WS-HSCR mimicks the megacolon and white coat-spotting observed in Ednrb mouse mutants. Since mouse mutants for the EDNRB ligand, endothelin-3 (EDN3), displayed a similar phenotype, the EDN3 gene was regarded as an alternative candidate gene in WS-HSCR. Here, we report a homozygous substitution/deletion mutation of the EDN3 gene in a WS-HSCR patient. EDN3 thus becomes the third known gene (after RET and EDNRB) predisposing to HSCR, supporting the view that the endothelin-signaling pathways play a major role in the development of neural crests.

SHOX mutations in dyschondrosteosis (Leri-Weill syndrome).

Dyschondrosteosis (DCS) is an autosomal dominant form of mesomelic dysplasia with deformity of the forearm (Madelung deformity; ref. 3). Based on the observation of XY translocations (p22,q12; refs 4-6) in DCS patients, we tested the pseudoautosomal region in eight families with DCS and showed linkage of the DCS gene to a microsatellite DNA marker at the DXYS233 locus (Zmax=6.26 at theta=0). The short stature homeobox-containing gene (SHOX), involved in idiopathic growth retardation and possibly Turner short stature, maps to this region and was therefore regarded as a strong candidate gene in DCS. Here, we report large-scale deletions (in seven families) and a nonsense mutation (in one family) of SHOX in patients with DCS and show that Langer mesomelic dwarfism results from homozygous mutations at the DCS locus.

Aconitase and mitochondrial iron-sulphur protein deficiency in Friedreich ataxia.

Friedreich ataxia (FRDA) is a common autosomal recessive degenerative disease (1/50,000 live births) characterized by a progressive-gait and limb ataxia with lack of tendon reflexes in the legs, dysarthria and pyramidal weakness of the inferior limbs. Hypertrophic cardiomyopathy is observed in most FRDA patients. The gene associated with the disease has been mapped to chromosome 9q13 (ref. 3) and encodes a 210-amino-acid protein, frataxin. FRDA is caused primarily by a GAA repeat expansion within the first intron of the frataxin gene, which accounts for 98% of mutant alleles. The function of the protein is unknown, but an increased iron content has been reported in hearts of FRDA patients and in mitochondria of yeast strains carrying a deleted frataxin gene counterpart (YFH1), suggesting that frataxin plays a major role in regulating mitochondrial iron transport. Here, we report a deficient activity of the iron-sulphur (Fe-S) cluster-containing subunits of mitochondrial respiratory complexes I, II and III in the endomyocardial biopsy of two unrelated FRDA patients. Aconitase, an iron-sulphur protein involved in iron homeostasis, was found to be deficient as well. Moreover, disruption of the YFH1 gene resulted in multiple Fe-S-dependent enzyme deficiencies in yeast. The deficiency of Fe-S-dependent enzyme activities in both FRDA patients and yeast should be related to mitochondrial iron accumulation, especially as Fe-S proteins are remarkably sensitive to free radicals. Mutated frataxin triggers aconitase and mitochondrial Fe-S respiratory enzyme deficiency in FRDA, which should therefore be regarded as a mitochondrial disorder.

X-linked spastic paraplegia and Pelizaeus-Merzbacher disease are allelic disorders at the proteolipid protein locus.

Three forms of X-linked spastic paraplegia (SPG) have been defined. One locus (SPG 1) maps to Xq28 while two clinically distinct forms map to Xq22 (SPG2). A rare X-linked dysmyelinating disorder of the central nervous system, Pelizaeus-Merzbacher disease (PMD), has also been mapped to Xq21-q22, and is caused by mutations in the proteolipid protein gene (PLP) which encodes two myelin proteins, PLP and DM20. While narrowing the genetic interval containing SPG2 in a large pedigree, we found that PLP was the closest marker to the disease locus, implicating PLP as a possible candidate gene. We have found that a point mutation (His139Tyr) in exon 3B of an affected male produces a mutant PLP but a normal DM20, and segregates with the disease (Zmax = 6.63, theta = 0.00). It appears, therefore, that SPG2 and PMD are allelic disorders.

A gene for achondroplasia-hypochondroplasia maps to chromosome 4p.

Achondroplasia (ACH) is a frequent condition of unknown origin characterized by short-limbed dwarfism and macrocephaly. Milder forms, termed hypochondroplasias (HCH) result in short stature with radiological features similar to those observed in ACH. We report on the mapping of a gene causing ACH/HCH to human chromosome 4p16.3, by linkage to the iduronidase A (IDUA) locus, in 15 informative families (Z max = 3.01 at theta = 0 for ACH; Z max = 4.71 at theta = 0 for ACH/HCH). Multipoint linkage analysis provides evidence for mapping the disease locus telomeric to D4S412 (location score in log 10 = 4.60). Moreover, this study supports the view that ACH and HCH are genetically homogeneous in our series.

A frame-shift deletion in the survival motor neuron gene in Spanish spinal muscular atrophy patients.

Spinal muscular atrophy (SMA) is a frequent autosomal recessive disease characterized by degeneration of the motor neurons of the spinal cord causing proximal paralysis with muscle atrophy. The region on chromosome 5q13 encompassing the disease gene is particularly unstable and prone to large-scale deletions whose characterization recently led to the identification of the survival motor neuron (SMN) gene. We now present a genetic analysis of 54 unrelated Spanish SMA families that has revealed a 4-basepair (bp) deletion (AGAG) in exon 3 of SMN in four unrelated patients. This deletion, which results in a frameshift and a premature stop codon, occurs on the same haplotype background, suggesting that a single mutational event is involved in the four families. The other patients showed either deletions of the SMN gene (49/54) or a gene conversion event changing SMN exon 7 into its highly homologous copy (cBCD541, 1/54). This observation gives strong support to the view that mutations of the SMN gene are responsible for the SMA phenotype as it is the first frameshift mutation reported in SMA.

Mutation of a nuclear succinate dehydrogenase gene results in mitochondrial respiratory chain deficiency.

We now report a mutation in the nuclear-encoded flavoprotein (Fp) subunit gene of the succinate dehydrogenase (SDH) in two siblings with complex II deficiency presenting as Leigh syndrome. Both patients were homozygous for an Arg554Trp substitution in the Fp subunit. Their parents (first cousins) were heterozygous for the mutation that occurred in a conserved domain of the protein and was absent from 120 controls. The deleterious effect of the Arg to Trp substitution on the catalytic activity of SDH was observed in a SDH- yeast strain transformed with mutant Fp cDNA. The Fp subunit gene is duplicated in the human genome (3q29; 5p15), with only the gene on chromosome 5 expressed in human-hamster somatic cell hybrids. This is the first report of a nuclear gene mutation causing a mitochondrial respiratory chain deficiency in humans.

A duplication in the L1CAM gene associated with X-linked hydrocephalus.

Recently, a mutation in the gene for the neural cell adhesion molecule L1CAM, located at chromosome Xq28, was found in a family with X-linked hydrocephalus (HSAS). However, as the L1CAM mutation could only be identified in one HSAS family, it remained unclear whether or not L1CAM was the gene responsible for HSAS. We have conducted a mutation analysis of L1CAM in 25 HSAS families. The mutation reported previously was not found in any of these families. In one family, however, a 1.3 kilobases (kb) genomic duplication was identified, cosegregating with HSAS and significantly changing the intracellular domain of the L1CAM protein. These results confirm that L1CAM is the HSAS gene.

Mutations of the TWIST gene in the Saethre-Chotzen syndrome.

Saethre-Chotzen syndrome (acrocephalo-syndactyly type III, ACS III) is an autosomal dominant craniosynostosis with brachydactyly, soft tissue syndactyly and facial dysmorphism including ptosis, facial asymmetry and prominent ear crura. ACS III has been mapped to chromosome 7p21-22. Of interest, TWIST, the human counterpart of the murine Twist gene, has been localized on chromosome 7p21 as well. The Twist gene product is a transcription factor containing a basic helix-loop-helix (b-HLH) domain, required in head mesenchyme for cranial neural tube morphogenesis in mice. The co-localisation of ACS III and TWIST prompted us to screen ACS III patients for TWIST gene mutations especially as mice heterozygous for Twist null mutations displayed skull defects and duplication of hind leg digits. Here, we report 21-bp insertions and nonsense mutations of the TWIST gene (S127X, E130X) in seven ACS III probands and describe impairment of head mesenchyme induction by TWIST as a novel pathophysiological mechanism in human craniosynostoses.