Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) as a cause of liver disease in infants in the UK.

Citrin deficiency is a disorder with two phenotypes: neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD), and adult-onset type II citrullinaemia (CTLN2). NICCD presents in the first few weeks of life with prolonged cholestasis and metabolic abnormalities including aminoacidaemia (notably citrulline, tyrosine, threonine, arginine and methionine) and galactosuria. Symptoms resolve within the first year of life, thus making a diagnosis difficult after this time. Although patients subsequently remain generally healthy, some may develop more severe symptoms of CTLN2, characterized by neurological changes, one or more decades later. To date more than 400 cases have been reported, almost all from East Asia (mainly Japan). Here we describe the first two cases of NICCD in infants from the UK, one of caucasian origin and one of Pakistani origin. Both showed typical clinical and biochemical changes with a diagnosis confirmed by the presence of previously unreported mutations in the SLC25A13 gene. The presence of citrin deficiency in other ethnic groups means that NICCD needs to be considered in the diagnosis of any neonate with an unexplained cholestasis. We discuss both the difficulties in diagnosing these patients in populations where very few DNA mutations have been identified and the problems faced in the management of these patients. These findings also raise the possibility of adults with CTLN2 in whom a diagnosis has yet to be made.

Variable phenotype including Leigh syndrome with a 9185T>C mutation in the MTATP6 gene.

We describe 15 members of a Caucasian family with an apparently homoplasmic T-->C mutation at nucleotide position 9185 (9185T>C) in the mtDNA encoded MTATP6 (ATPase 6) gene. The clinical phenotype is extremely variable and includes late-onset Leigh syndrome (LS), isolated demyelinating peripheral neuropathy and neurogenic muscle weakness, ataxia and retinitis pigmentosa (NARP). Following recent reports of this same mutation in a single case and in a family with late-onset LS and NARP-like features, our paper emphasises the role of MTATP6 in LS and expands the associated clinical phenotype further.

Assessment of the genetic causes of recessive childhood non-syndromic deafness in the UK - implications for genetic testing.

Approximately one in 2000 children is born with a genetic hearing impairment, mostly inherited as a non-syndromic, autosomal recessive trait, for which more than 30 different genes have been identified. Previous studies have shown that one of these genes, connexin 26 (GJB2), accounts for 30-60% of such deafness, but the relative contribution of the many other genes is not known, especially in the outbred UK population. This lack of knowledge hampers the development of diagnostic genetic services for deafness. In an effort to determine the molecular aetiology of deafness in the population, 142 sib pairs with early-onset, non-syndromic hearing impairment were recruited. Those in whom deafness could not be attributed to GJB2 mutations were investigated further for other mapped genes. The genetic basis of 55 cases (38.7%) was established, 33.1% being due to mutations in the GJB2 gene and 3.5% due to mutations in SLC26A4. None of the remaining 26 loci investigated made a significant contribution to deafness in a Caucasian population. We suggest that screening the GJB2 and SLC26A4 genes should form the basis of any genetic testing programme for childhood deafness and highlight a number of important issues for consideration and future work.

The contribution of USH1C mutations to syndromic and non-syndromic deafness in the UK.

Denaturing high-performance liquid chromatography (DHPLC) was used to screen 14 UK patients with Usher syndrome type 1, in order to assess the contribution of mutations in USH1C to type 1 Usher. In addition, 16 Caucasian sib pairs and two small consanguineous families with non-syndromic deafness, who were concordant for haplotypes around DFNB18, were also screened for mutations in the USH1C gene. Two Usher type 1 patients were found to have the 238-239insC mutation reported previously; one of Greek Cypriot origin was homozygous for the mutation and another Caucasian was heterozygous. This indicates that mutations in the USH1C gene make a greater contribution to Usher syndrome type 1 than originally thought, which has implications for the genetic testing of families with Usher syndrome in the UK. Analysis using intragenic single nucleotide polymorphisms (SNPs) revealed that the haplotypic background bearing this common mutation was not consistent across the gene in two families, and that there are either two haplotypes on which the mutation has arisen or that there has been a recombination on a single haplotype. We found no evidence of mutations in USH1C in the patients with non-syndromic deafness, suggesting that the gene is not a major contributor to autosomal-recessive non-syndromic deafness in the UK.

Antioxidant capacity after acute ischaemic stroke.

BACKGROUND: Experimental studies have reported a rapid increase in the production of markers of oxidative damage following acute stroke due to the reperfusion event following ischaemia, and that endogenous antioxidant defences are rapidly depleted, permitting further tissue damage. AIM: To measure changes in antioxidant capacity (individual and total) in stroke disease within a known time period post infarct. DESIGN: Observational cohort study. METHODS: We studied 31 acute ischaemic stroke patients; 26 hospitalized non-stroke patients and 23 community-based healthy controls. Non-fasting venous blood was obtained within 24 h, at 48-72 h and at 7 days after stroke onset (after hospitalization for non-stroke patients) and at baseline for community controls. Vitamins E and C, total plasma glutathione, total antioxidant capacity (TAC), uric acid, thiobarbituric-acid-reactive substances (TBARS), serum albumin, transferrin and C-reactive protein (CRP) were measured. RESULTS: Baseline glutathione concentrations were non-significantly lowest and TBARS significantly highest in ischaemic stroke patients compared with controls. Serum TAC strongly correlated with serum uric acid. Under multivariate analysis, serum uric acid explained most of the variance in TAC during the study period. Despite increased concentrations of uric acid, TAC was reduced in stroke patients compared with controls. Serum vitamin C concentrations deteriorated significantly in stroke patients, and differences between the cumulative changes between strokes and hospital controls were also statistically significant (p=0.013). DISCUSSION: There was some evidence of reduction in TAC, despite increased uric acid concentrations, and deterioration in serum vitamin C levels in ischaemic stroke patients compared with controls.

Mutations in a new gene encoding a protein of the hair bundle cause non-syndromic deafness at the DFNB16 locus.

Hearing impairment affects about 1 in 1,000 children at birth. Approximately 70 loci implicated in non-syndromic forms of deafness have been reported in humans and 24 causative genes have been identified (see also http://www.uia.ac.be/dnalab/hhh). We report a mouse transcript, isolated by a candidate deafness gene approach, that is expressed almost exclusively in the inner ear. Genomic analysis shows that the human ortholog STRC (so called owing to the name we have given its protein-stereocilin), which is located on chromosome 15q15, contains 29 exons encompassing approximately 19 kb. STRC is tandemly duplicated, with the coding sequence of the second copy interrupted by a stop codon in exon 20. We have identified two frameshift mutations and a large deletion in the copy containing 29 coding exons in two families affected by autosomal recessive non-syndromal sensorineural deafness linked to the DFNB16 locus. Stereocilin is made up of 1,809 amino acids, and contains a putative signal petide and several hydrophobic segments. Using immunohistolabeling, we demonstrate that, in the mouse inner ear, stereocilin is expressed only in the sensory hair cells and is associated with the stereocilia, the stiff microvilli forming the structure for mechanoreception of sound stimulation.

Multiple origins of the mtDNA 7472insC mutation associated with hearing loss and neurological dysfunction.

Several mtDNA mutations have been reported in families with both syndromic and non-syndromic hearing loss. One such mutation is the heteroplasmic 7472insC in the tRNA(Ser(UCN)) gene which has been found in six families, all from Western Europe. However, it was not clear if this distribution was due to a common founder effect or chance sampling of several unrelated families, the 7472insC mutation having occurred multiple times. Haplotype analysis of all six families supports the latter notion. This confirms the pathogenicity of the 7472insC mutation and suggests it may exist in other populations where it may prove to be a small but significant cause of hearing loss, particularly when neurological symptoms are also present.

Prevalence of mitochondrial DNA mutations in childhood/congenital onset non-syndromal sensorineural hearing impairment.

Genetic factors are the major causes of childhood hearing impairment. Whereas autosomal recessive mutations account for the majority of prelingual non-syndromic sensorineural hearing impairment (NSSHI), the relative contribution of mitochondrial DNA (mtDNA) mutations to childhood onset NSSHI has not been established. We screened 202 subjects with congenital/childhood onset NSSHI, consisting of 110 sporadic cases, 75 sib pairs, and 17 families with affected subjects in more than one generation, in order to determine the prevalence of mtDNA mutations associated with NSSHI.mtDNA mutations were found in three of 10 families (30%) in whom the affected members were related through the maternal lineage. One sporadic case (0.9%) was also found to have a known mtDNA mutation but none was found in the sib pairs. Although the prevalence of mtDNA mutations was low in the group as a whole (2%), we suggest that screening should be considered in cases of childhood hearing impairment when it is progressive and particularly in families where transmission is compatible with maternal inheritance.

Maternally inherited hearing impairment in a family with the mitochondrial DNA A7445G mutation.

Despite the increasing number of reports of families with hearing impairment and mitochondrial DNA (mtDNA) mutations, the frequency of these mutations as causes of non-syndromic sensorineural hearing impairment (NSSHI) remains unknown. Mutations such as A1555G, A7445G and 7472insC have been found in several unrelated families implying they are more frequent than initially thought. We describe a family with NSSHI due to the presence of the homoplasmic mtDNA A7445G mutation in the tRNASer(UCN) gene. This is the fourth such family described with this mutation, all of different genetic backgrounds. Our study also demonstrates the difficulties sometimes encountered in establishing mitochondrial inheritance of hearing impairment in some families.

A novel mutation in the mitochondrial tRNA(Ser(UCN)) gene in a family with non-syndromic sensorineural hearing impairment.

We describe a family with non-syndromic sensorineural hearing impairment inherited in a manner consistent with maternal transmission. Affected members were found to have a novel heteroplasmic mtDNA mutation, T7510C, in the tRNA(Ser(UCN)) gene. This mutation was not found in 661 controls, is well conserved between species, and disrupts base pairing in the acceptor stem of the tRNA, making it the probable cause of hearing impairment in this family. Sequencing of the other mitochondrial tRNA genes did not show any other pathogenic mutations. Four other mutations causing hearing impairment have been reported in the tRNA(Ser(UCN)) gene, two having been shown to affect tRNA(Ser(UCN)) levels. With increasing numbers of reports of mtDNA mutations causing hearing impairment, screening for such mutations should be considered in all cases unless mitochondrial inheritance can be excluded for certain.

Mitochondrial defects and hearing loss.

The techniques of human molecular genetics have been rapidly applied to the study of hearing loss. These studies have implicated more than 60 loci as causes of nonsyndromic hearing loss. Mutations at more than a dozen nuclear genes have been demonstrated to cause hearing loss, and these have been covered in recent reviews. However, a perhaps unexpected feature of the molecular characterization of human hearing loss has been the occurrence of mutations in the mitochondrial DNA (mtDNA). The importance of mitochondrial function in hearing is emphasized by the recent discovery of mutations in a nuclear-encoded mitochondrial protein which results in hearing loss. This article reviews the current status of our knowledge of mtDNA mutations that have been shown to cause hearing loss, and the suggestion of potential molecular, cellular and tissue-specific pathophysiological mechanisms by which dysfunction of mitochondria may lead to a loss of hearing.

Sensorineural hearing loss and the 1555G mitochondrial DNA mutation.

Recent studies have identified a mitochondrial DNA mutation (1555G) which causes sensorineural hearing loss (SNHL). In many cases deafness follows exposure to aminoglycoside antibiotics, the 1555G mutation sensitizing the inner ear to these drugs. The 50 cases reported to date are discussed, as are the possible mechanisms behind the pathogenesis of this mutation. This finding in families from a wide range of ethnic backgrounds suggests that the 1555G mutation is one of the more common genetic causes of SNHL and provides a fascinating example of how a genetic mutation interacts with an environmental factor with harmful effect.

Serum hyaluronic acid in Down's syndrome.

In agreement with previous studies we show an age-related increase in serum levels of hyaluronic acid in healthy individuals. Levels in Down's syndrome persons were slightly higher, especially in those with hypothyroidism. This provides further evidence that the normal ageing process is accelerated in Down's syndrome.

Familial streptomycin ototoxicity in a South African family: a mitochondrial disorder.

The vestibular and ototoxic effects of the aminoglycoside antibiotics (streptomycin, gentamycin, kanamycin, tobramycin, neomycin) are well known; streptomycin, in particular, has been found to cause irreversible, profound, high frequency sensorineural deafness in hypersensitive persons. Aminoglycoside ototoxicity occurs both sporadically and within families and has been associated with a mitochondrial DNA (mtDNA) 1555A to G point mutation in the 12S ribosomal RNA gene. We report on the molecular analysis of a South African family with streptomycin induced sensorineural deafness in which we have found transmission of this same predisposing mutation. It is now possible to identify people who are at risk of hearing loss if treated with aminoglycosides in the future and to counsel them accordingly. In view of the fact that aminoglycoside antibiotics remain in widespread use for the treatment of infections, in particular for tuberculosis, which is currently of epidemic proportions in South Africa, this finding has important implications for the family concerned. In addition, other South African families may potentially be at risk if they carry the same mutation.

Multiple origins of a mitochondrial mutation conferring deafness.

A point mutation (1555G) in the smaller ribosomal subunit of the mitochondrial DNA (mtDNA) has been associated with maternally inherited traits of hypersensitivity to streptomycin and sensorineural deafness in a number of families from China, Japan, Israel, and Africa. To determine whether this distribution was the result of a single or multiple mutational events, we carried out genetic distance analysis and phylogenetic analysis of 10 independent mtDNA D-loop sequences from Africa and Asia. The mtDNA sequence diversity was high (2.21%). Phylogenetic analysis assigned 1555G-bearing haplotypes at very divergent points in the human mtDNA evolutionary tree, and the 1555G mutations occur in many cases on race-specific mtDNA haplotypes, both facts are inconsistent with a recent introgression of the mutation into these races. The simplest interpretation of the available data is that there have been multiple origins of the 1555G mutation. The genetic distance among mtDNAs bearing the pathogenic 1555G mutation is much larger than among mtDNAs bearing either evolutionarily neutral or weakly deleterious nucleotide substitutions (such as the 4336G mutation). These results are consistent with the view that pathogenic mtDNA haplotypes such as 1555G arise on disparate mtDNA lineages which because of negative natural selection leave relatively few related descendants. The co-existence of the same mutation with deafness in individuals with very different nuclear and mitochondrial genetic backgrounds confirms the pathogenicity of the 1555G mutation.

Mitochondrial DNA mutations in Alzheimer's disease.

Several reports have indicated that point mutations of the mitochondrial DNA (mtDNA) contribute to the pathogenesis of Alzheimer's disease (AD). However, other groups have failed to find similar associations between these mutations and AD. A recent report described a set of mutations in the mtDNA encoded cytochrome oxidase genes which may account for 20% of all AD cases. We screened brain tissue from 65 AD patients for each of these previously reported mtDNA mutations but were unable to find an increased incidence of any of them in our AD sample. However, one patient with a mutation in the APP gene did harbour a novel mtDNA mutation (G to C at position 5705 in the tRNAAsn gene) that might have contributed to the very early onset of dementia in this individual. The role of mtDNA mutations in the pathogenesis of AD remains unclear, but they do not appear to be primary causes but may contribute to the onset of the disease.

Topical gentamicin-induced hearing loss: a mitochondrial ribosomal RNA study of genetic susceptibility.

Mitochondrial ribosomal RNA mutation has been shown to predispose affected individuals to aminoglycoside-induced hearing loss (AIHL). An A-to-G nucleotide substitution at the 1555 position within the 12S ribosomal RNA gene has been identified with a maternally inherited pattern in affected Asian pedigrees. The aim of this study was to identify the frequency of this DNA polymorphism in subjects who appear to show hypersensitivity to topical (middle ear) application of aminoglycoside. In this pilot study, 10 subjects with AIHL were recruited. Eight underwent vestibular ablative therapy with middle ear instillation of gentamicin for disabling vertigo, and two were treated with topical antibiotic drops for otitis media in the presence of a tympanic perforation. DNA samples were extracted, and polymerase chain reaction (PCR) technique was used for gene amplification and purification, searching for 1555 A-to-G substitution in the 12S ribosomal RNA gene. None of the subjects demonstrated this specific mutation.

Nicotinamide as a precursor for NAD+ prevents apoptosis in the mouse brain induced by tertiary-butylhydroperoxide.

The vitamin nicotinamide can protect against oxidative stress-induced apoptosis in the brain when used as a precursor for nicotinamide adenine dinucleotide (NAD+). The intracerebroventricular administration of tertiary-butylhydroperoxide (t-buOOH) to mice was used to simulate physiologic oxidative stress and apoptosis which may occur in some neurodegenerative conditions. t-buOOH produced characteristic apoptotic nuclear degeneration in neurons with extensive fragmentation of DNA. In this report we show that the elevation of NAD+ by nicotinamide prevents DNA fragmentation during apoptosis or necrosis in the brain as stimulated by t-buOOH administration. NAD+ levels can be increased by 50% in the brain. This may prevent the critical depletion of NAD+ by poly(ADP-ribose) polymerase (PARP) and provide additional substrate during the repair of DNA. Nicotinamide may be of particular interest in the treatment of neurodegeneration.

Degeneration of human oncogenes and mitochondrial genes occurs in cells that exhibit age-related pathology.

The development of a new class of assays to determine in vivo mutation frequencies has provided new perspectives on the timing, location, and distribution of somatic mutagenesis in mitochondrial genes and in oncogenes of the aging human body. This descriptive information has led to the inference of new models for age-related pathophysiology and oncogenesis. Mutations of mitochondrial genes rise rapidly with age to frequencies a thousand-fold higher than those of nuclear genes. Genotypic selection analysis has revealed that mitochondrial mutations accumulate predominantly in nonmitotic cells whose age-dependent loss is associated with pathology. Random mitochondrial mutation is most likely to inactivate Complex I, deficiency of which induces mitochondrial superoxide formation and cell death. Genotypic selection of oncogenic mutations at the BCL2 and p53 loci has revealed that the cell specificity of oncogenic mutations in persons without cancer correlates well with sites of tumor origin, indicating that cells bearing such mutations are the likely precursors of future tumors. Quantitative variation in human BCL2 mutation frequency is extensive, and BCL2 mutation frequency rises with age, concordant with increased risk for lymphoma. The clonality and persistence of BCL2 mutations suggests two specific testable mechanisms of lymphomagenesis. BCL2 mutation frequency rises in persons exposed to cigarette smoke, and more p53 mutations occur in skin exposed to sunlight than in unexposed skin. Thus, in addition to their likely relevance to future cancer risk, the dose-response relationship between exposure and oncogenic mutations indicates promise for their future use as in vivo biodosimeters of human exposure to carcinogens.