Mitochondrial DNA deletions in human brain: regional variability and increase with advanced age.

We have examined the role of somatic mitochondrial DNA (mtDNA) mutations in human ageing by quantitating the accumulation of the common 4977 nucleotide pair (np) deletion (mtDNA4977) in the cortex, putamen and cerebellum. A significant increase in the mtDNA4977 deletion was seen in elderly individuals. In the cortex, the deleted to total mtDNA ratio ranged from 0.00023 to 0.012 in 67-77 year old brains and up to 0.034 in subjects over 80. In the putamen, the deletion level ranged from 0.0016 to 0.010 in 67 to 77 years old up to 0.12 in individuals over the age of 80. The cerebellum remained relatively devoid of mtDNA deletions. Similar changes were observed with a different 7436 np deletion. These changes suggest that somatic mtDNA deletions might contribute to the neurological impairment often associated with ageing.

Mitochondrial DNA mutation associated with Leber's hereditary optic neuropathy.

Leber's hereditary optic neuropathy is a maternally inherited disease resulting in optic nerve degeneration and cardiac dysrhythmia. A mitochondrial DNA replacement mutation was identified that correlated with this disease in multiple families. This mutation converted a highly conserved arginine to a histidine at codon 340 in the NADH dehydrogenase subunit 4 gene and eliminated an Sfa NI site, thus providing a simple diagnostic test. This finding demonstrated that a nucleotide change in a mitochondrial DNA energy production gene can result in a neurological disease.

Mitochondrial DNA mutations in human degenerative diseases and aging.

A wide variety of mitochondrial DNA (mtDNA) mutations have recently been identified in degenerative diseases of the brain, heart, skeletal muscle, kidney and endocrine system. Generally, individuals inheriting these mitochondrial diseases are relatively normal in early life, develop symptoms during childhood, mid-life, or old age depending on the severity of the maternally-inherited mtDNA mutation; and then undergo a progressive decline. These novel features of mtDNA disease are proposed to be the product of the high dependence of the target organs on mitochondrial bioenergetics, and the cumulative oxidative phosphorylation (OXPHOS) defect caused by the inherited mtDNA mutation together with the age-related accumulation mtDNA mutations in post-mitotic tissues.

Mitochondrial DNA complex I and III mutations associated with Leber's hereditary optic neuropathy.

Four new missense mutations have been identified through restriction analysis and sequencing of the mitochondrial DNAs (mtDNA) from Leber's hereditary optic neuropathy (LHON) patients who lacked the previously identified 11778 mutation. Each altered a conserved amino acid and correlated with the LHON phenotype in population and phylogenetic analyses. The nucleotide pair (np) 13708 mutation (G to A, ND5 gene) changed an alanine to a threonine and was found in 6/25 (24%) of non-11778 LHON pedigrees and in 5.0% of controls, the np 15257 mutation (G to A, cytochrome b gene) changed an aspartate to an asparagine and was found in 4 of the 13708-positive pedigrees and 0.3% of controls, the np 15812 mutation (G to A, cytochrome b gene) changed a valine to a methionine and was detected in two of the 15257-positive pedigrees and 0.1% of controls and the np 5244 mutation (G to A, ND2 gene) changed a glycine to a serine and was found in one of the 15812-positive patients and none of 2103 controls. The 15257 mutation altered a highly conserved amino acid in an extramembrane domain of cytochrome b that is associated with the ligation of the low potential b566 heme and the 5244 mutation altered a strongly evolutionarily conserved region of the ND2 polypeptide. The 13708 and 15812 mutations changed moderately conserved amino acids. Haplotype and phylogenetic analysis of the four np 15257 mtDNAs revealed that all harbored the same rare Caucasian haplotype and that the np 13708, np 15257, np 15812 and np 5244 mutations were added sequentially along this mtDNA lineage. Since the percentage of sighted controls decreases as these mutations accumulate, it appears that they interact synergistically, each increasing the probability of blindness. The involvement of both mitochondrial complex I (np 5244, 11778, 13708) and complex III (np 15257, 15812) mutations in LHON indicates that the clinical manifestations of this disease are the product of an overall decrease in mitochondrial energy production rather than a defect in a specific mitochondrial enzyme.

Mitochondrial DNA variation in human evolution and disease.

Analysis of mitochondrial DNA (mtDNA) variation has permitted the reconstruction of the ancient migrations of women. This has provided evidence that our species arose in Africa about 150000 years before present (YBP), migrated out of Africa into Asia about 60000 to 70000 YBP and into Europe about 40000 to 50000 YBP, and migrated from Asia and possibly Europe to the Americas about 20000 to 30000 YBP. Although much of the mtDNA variation that exists in modern populations may be selectively neutral, studies of the mildly deleterious mtDNA mutations causing Leber's hereditary optic neuropathy (LHON) have demonstrated that some continent-specific mtDNA lineages are more prone to manifest the clinical symptoms of LHON than others. Hence, all mtDNA lineages are not equal, which may provide insights into the extreme environments that were encountered by our ancient ancestor, and which may be of great importance in understanding the pathophysiology of mitochondrial disease.

Atypical Leber's hereditary optic neuropathy with molecular confirmation.

OBJECTIVE: Leber's hereditary optic neuropathy (LHON) is typically a familial disease of primarily young, male adults. Analysis of mitochondrial DNA has identified point mutations associated with LHON and allowed us to identify cases of LHON not consistent with traditional descriptions of the disease. DATA SOURCES: The collective experience of three tertiary referral centers contributed to this report. STUDY SELECTION: Patients with bilateral optic neuropathies who were positive for the 11778 LHON mutation were included in this study if they were female and there was no family history of visual loss. DATA EXTRACTION: Six case histories are presented. DATA SYNTHESIS: The diagnosis of LHON remained unknown in six female patients with bilateral optic neuropathies until molecular analysis revealed the 11778 mitochondrial DNA mutation. None of the patients had a family history of visual loss, and five were initially diagnosed as having factitious visual loss. Other individual features atypical for LHON included lack of the characteristic LHON funduscopic appearance, bitemporal hemianopia, optic disc cupping, and premonitory episodes of transient visual loss. In one patient the correct diagnosis was delayed 17 years. CONCLUSIONS: The diagnosis of LHON should be considered in all cases of unexplained optic neuropathy, including those with negative family history, late or early age at onset, female gender, or normal funduscopic appearance.

Evidence in a lethal infantile mitochondrial disease for a nuclear mutation affecting respiratory complexes I and IV.

A child died at 4 months of age of a lethal infantile mitochondrial disease associated with cardiomyopathy. Detailed pathologic evaluation of this patient revealed abnormalities in the striated muscle, smooth muscle, heart, and liver, but not the central nervous system. Biochemical analysis revealed a combined complex I and IV deficiency in skeletal muscle, heart, and liver, but not in kidney and brain. Analysis of mitochondrial translation products and mitochondrial DNA failed to detect any abnormality. Parallel studies on both parents were uniformly normal. These data support the hypothesis that this disease was the result of a nuclear DNA mutation in a developmental stage-specific and tissue-specific oxidative phosphorylation-gene.

The clinical characteristics of pedigrees of Leber's hereditary optic neuropathy with the 11778 mutation.

In a study of the phenotypic characteristics of pedigrees of Leber's hereditary optic neuropathy positive for the mitochondrial DNA mutation at position 11778, 28 of 49 pedigrees were represented by singleton cases. Seven families, including six singleton pedigrees, had maternal family members with a mixture of mutant and normal mitochondrial DNA (heteroplasmy). Seventy-two affected individuals from 43 families showed a male predominance of 81.9% (59/72) and ages of onset of visual loss ranging from 8 to 60 years. The time interval between affected eyes averaged 1.8 months; the duration of progression of visual loss in each eye averaged 3.7 months. Visual acuity was 20/200 or worse in 107 of 109 (98.2%) eyes. Telangiectatic microangiopathy, disk pseudoedema, or vascular tortuosity, ophthalmoscopic features believed to be classic of Leber's hereditary optic neuropathy, were noted in 30 of 52 patients. Visual-evoked responses were typically absent or abnormal. Electrocardiograms, fluorescein angiograms, cerebrospinal fluid analyses, brain computed tomography, and magnetic resonance imaging were usually normal. There were no consistent neurologic or systemic illnesses associated with these Leber's pedigrees. In many cases, the diagnosis would not have been suspected because of the absence of a compatible family history, typical clinical profile, or ophthalmoscopic appearance. Genetic analysis showed the mitochondrial DNA mutation at position 11778, which established the diagnosis of Leber's hereditary optic neuropathy and has allowed for a broader view of the clinical features of this disease.

Variable genotype of Leber's hereditary optic neuropathy patients.

Leber's hereditary optic neuropathy is caused by a single nucleotide change in the mitochondrial deoxyribonucleic acid (mtDNA). Each cell contains thousands of mitochondrial DNA molecules. We demonstrated that in certain isolated instances, the proband and close maternal lineage relatives can have mixtures of mutant and normal mitochondrial DNA molecules (heteroplasmy). The proportion of mutant mitochondrial DNA molecules was found to shift markedly across generations and within the tissues of an individual. One unaffected mother had 65% mutant mitochondrial DNA molecules whereas her affected son had essentially 100% mutant mitochondrial DNA molecules. Two affected individuals had predominantly mutant mitochondrial DNA in their blood, but significant normal mitochondrial DNA in their hair. The demonstration of heteroplasmy within maternal lineages and affected individuals means that the successful determination of the mitochondrial DNA genotype of a family or patient with Leber's hereditary optic neuropathy requires testing of more than one family member and more than one tissue from each individual.

Epidemic neuropathy in Cuba not associated with mitochondrial DNA mutations found in Leber's hereditary optic neuropathy patients. Cuba Neuropathy Field Investigation Team.

An epidemic neuropathy in Cuba has caused bilateral optic neuropathies in more than 26,000 people during the past three years. Various pathogenetic factors have been proposed, including toxins, nutritional deficiencies, and an underlying genetic predisposition involving mitochondrial DNA. As part of a case-control collaborative investigation, 135 Cuban blood samples were analyzed for the most common mitochondrial DNA mutations associated with Leber's hereditary optic neuropathy. None of the participants tested were found to have the mitochondrial DNA mutations at nucleotide positions 11778, 3460, 14484, 7444, or 9804. Of 57 definite case subjects and 69 normal control subjects, three case and three control subjects had the mutation at nucleotide position 9438, three different case and three different control subjects had the mutation at position 13708, and one case and one control subject had the mutation at position 15257 in association with the mutation at position 13708. The most common mitochondrial DNA mutations associated with Leber's hereditary optic neuropathy do not appear to be contributing factors in the epidemic neuropathy in Cuba. We also identified a large Cuban family with maternally related members who experienced visual loss consistent with the diagnosis of Leber's hereditary optic neuropathy. Maternal family members harbored the highly pathogenetic mutation at nucleotide position 11778.

Optic disk cupping and electrocardiographic abnormalities in an American pedigree with Leber's hereditary optic neuropathy.

We examined the clinical characteristics of affected and unaffected members of an American black family with the 11778 mitochondrial DNA mutation associated with Leber's hereditary optic neuropathy. Thirty-six individuals from four generations were included. All maternally related subjects were shown to be essentially homoplasmic for the 11778 mitochondrial DNA mutation in blood. Paternally related subjects did not carry this mutation. Patients affected with optic neuropathy had optic nerve head cupping. Loss of unmyelinated axons from the prelaminar optic nerve may be responsible for cupping in these patients. Electrocardiographic analysis of subjects carrying the 11778 mitochondrial DNA mutation disclosed statistically significant (P = .02) prolongation of the corrected OT interval as compared to control subjects. While the clinical significance of this magnitude of corrected QT prolongation is unknown, it may represent a systemic manifestation of the 11778 mutation. No electrocardiographic evidence of preexcitation syndromes was seen.

Association of mitochondrial DNA damage with aging and coronary atherosclerotic heart disease.

The role of somatic mitochondrial DNA (mtDNA) damage in human aging and progressive diseases of oxidative phosphorylation (OXPHOS) was examined by quantitating the accumulation of mtDNA deletions in normal hearts and hearts with coronary atherosclerotic disease. In normal hearts, mtDNA deletions appeared after 40 and subsequently accumulated with age. The common 4977 nucleotide pair (np) deletion (mtDNA4977) reached a maximum of 0.007%, with the mtDNA7436 and mtDNA10,422 deletions appearing at the same time. In hearts deprived of mitochondrial substrates due to coronary artery disease, the level of the mtDNA4977 deletion was elevated 7-220-fold over age-matched controls, with the mtDNA7436 and mtDNA10,422 deletions increasing in parallel. This cumulative mtDNA damage was associated with a compensatory 3.5-fold induction of nuclear OXPHOS gene mRNA and regions of ischemic hearts subjected to the greatest work load (left ventricle) showed the greatest accumulation of mtDNA damage and OXPHOS gene induction. These observations support the hypothesis that mtDNA damage does accumulate with age and indicates that respiratory stress greatly elevates mitochondrial damage.

MERRF: a model disease for understanding the principles of mitochondrial genetics.

The principles of mitochondrial genetics have evolved over the past 20 years. Careful identification of large pedigrees that were consistent with maternal inheritance has permitted detailed clinical and genetic investigations. Myoclonic epilepsy and ragged-red fiber (MERRF) disease has been a model disease for the application of these principles. MERRF is caused by an A to G mutation of the mitochondrial tRNA(Lys) at position 8344. The mutation is maternally inherited and heteroplasmic. Disease manifestations are dependent on replicative segregation of mutant and wild type mitochondrial DNAs and on the threshold effect. Characterization of the clinical, physiological, biochemical, and genetic manifestations of this disease has provided a better understanding of how to diagnose and manage oxidative phosphorylation diseases which are caused by mutations in the mitochondrial DNA.

A mitochondrial DNA mutation as a cause of Leber's hereditary optic neuropathy.

Leber's hereditary optic neuropathy is a maternally inherited disease associated with the late onset of bilateral loss of central vision and cardiac dysrhythmias. The maternal inheritance is explained by the mitochondrial origin of the disease. Analysis of the sequence of a mitochondrial DNA has indicated that a single nucleotide change at position 11778 is associated with this disease. This mutation converts the 340th amino acid of NADH dehydrogenase subunit 4 from an arginine to a histidine and eliminates an SfaNI endonuclease restriction site. A survey of restriction-fragment-length polymorphisms in the mitochondrial DNA of three independent families with this disease (an American black and two white European families) and 10 controls confirmed that this SfaNI site is associated with the disease. A phylogenetic tree for mitochondrial DNA polymorphism and sequence variants from three probands with Leber's disease and four controls was constructed, and the mutation at position 11778 was found to be associated with two mitochondrial DNA backgrounds--an American black mitochondrial DNA and a European mitochondrial DNA. Thus, this mutation must have arisen twice independently. Since the mutation correlated with symptoms of Leber's disease in both cases, these findings indicate that the mutation is a cause of the disease. This genetic analysis has identified the specific point mutation in the mitochondrial DNA that results in Leber's hereditary optic neuropathy.

African, Native American, and European mitochondrial DNAs in Cubans from Pinar del Rio Province and implications for the recent epidemic neuropathy in Cuba. Cuba Neuropathy Field Investigation Team.

Genetic predisposition, particularly specific mitochondrial DNA (mtDNA) backgrounds, has been proposed as a contributing factor in the expression of an epidemic of bilateral optic neuropathy that has affected residents of Cuba since 1991. To substantiate or refute the possibility that specific subsets of mtDNAs could participate in disease expression, we took advantage of the heterogeneous ethnic origin of the Cuban population and the recent identification of a number of mtDNA polymorphisms that appear to be specific for Africans, Native Americans, and Europeans. The screening of both carefully selected people with epidemic neuropathy and control subjects from the Pinar del Rio Province for these polymorphisms revealed that African, Native American, and European mtDNA haplotypes were equally represented among case and control subjects, and suggested that approximately 50% of Cuban mtDNAs originated from Europeans, 46% from Africans, and 4% from Native Americans. These findings demonstrate that mutations arising in specific mtDNAs are unlikely to play a role in the epidemic neuropathy and indicate that analysis of mtDNA haplotypes can be a valuable tool for assessing the relative maternal contribution of Africans, Native Americans, and Europeans in a mixed population.

Mitochondrial DNA mutations in epilepsy and neurological disease.

Recent discoveries in mitochondrial clinical genetics have revealed that a broad spectrum of clinical phenotypes are associated with mutations in mitochondrial DNA. Diseases caused by mutations in mitochondrial DNA are by nature quantitative. Myoclonic epilepsy and ragged-red fiber disease are caused by a mutation in the transfer RNA gene lysine. Although everyone in a maternal lineage will harbor the same mutation, the nature and severity of the symptoms vary markedly among individuals. This variability correlates with the inherited percentage of mutations in the individual's mitochondrial DNA and the individual's age. Age-related expression of mitochondrial disease has also been demonstrated for mitochondrial DNA deletions. Although deletions that retain both origins of replication result in late-onset disease because of the progressive enrichment of the deleted mitochondrial DNA, a 10.4-kb deletion that lacks the light-strand replication origin and maintains a stable mutant percentage in both tissues and cultured cells has been discovered. This deletion is associated with adult-onset diabetes and deafness, but not with ophthalmoplegia, ptosis, or mitochondrial myopathy. Biochemically, it causes a generalized defect in mitochondrial protein synthesis and oxidative phosphorylation. The age-related decline in oxidative phosphorylation could reflect the accumulation of somatic mitochondrial DNA mutations. Inhibition of oxidative phosphorylation stimulates this accumulation. The general paradigm for mitochondrial DNA diseases may be that inherited mutations inhibit the electron transport chain. This damages the mitochondrial DNA, further reducing oxidative phosphorylation. Ultimately, oxidative phosphorylation drops below the expression threshold of cells and tissues, and clinical symptoms appear.

Leber's hereditary optic neuropathy: a model for mitochondrial neurodegenerative diseases.

A number of human diseases have been attributed to defects in oxidative phosphorylation (OXPHOS) resulting from mutations in the mitochondrial DNA (mtDNA). One such disease is Leber's hereditary optic neuropathy (LHON), a neurodegenerative disease of young adults that results in blindness due to atrophy of the optic nerve. The etiology of LHON is genetically heterogeneous and in some cases multifactorial. Eleven mtDNA mutations have been associated with LHON, all of which are missense mutations in the subunit genes for the subunits of the electron transport chain complexes I, III, and IV. Molecular, biochemical, and population genetic studies have categorized these mutations as high risk (class I), low risk (class II), or intermediate risk (class I/II). Class I mutations appear to be primary genetic causes of LHON, while class II mutations are frequently found associated with class I genotypes and may serve as exacerbating genetic factors. Different LHON pedigrees can harbor different combinations of class I, II, or I/II mtDNA mutations, as shown by the complete sequence analysis of the mtDNAs of four LHON probands. The various mtDNA genotypes included an isolated class I mutation, combined class I+II mutations, and combined class I/II+II mutations. The occurrence of such genotypes supports the hypothesis that LHON may result from the additive effects of various genetic and environmental insults to OXPHOS, each of which increases the probability of blindness.

Diseases resulting from mitochondrial DNA point mutations.

A number of mitochondrial DNA (mtDNA) mutations have been identified which cause familial, late onset neuromuscular degenerative diseases. These include missense mutations in most of the mtDNA polypeptide genes as well as base substitutions in several tRNA genes. Missense mutations in the mitochondrial electron-transport genes cause Leber hereditary optic neuropathy. Ten mutations have been associated with this disease, but four at nps 11,178, 3460, 4160 and 15,257 appear sufficient in themselves to cause the disease. One missense mutation in the ATPase 6 gene at np 8993 causes a second phenotype, neurogenic muscle weakness, ataxia and retinitis pigmentosum. Transfer RNA mutations have been identified for myoclonic epilepsy and ragged-red fibre disease in the tRNA(Lys) gene at np 8344 and for the mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes syndrome and for maternal mitochondrial myopathy and cardiomyopathy syndrome in the tRNA(Leu)(UUR) gene at nps 3234 and 3260, respectively. Deficiencies in mitochondrial oxidative phosphorylation enzymes have been observed in several common neurodegenerative diseases such as Alzheimer and Parkinson diseases. Perhaps mtDNA mutations play a role in these as well.

Hypoxemia is associated with mitochondrial DNA damage and gene induction. Implications for cardiac disease.

OBJECTIVE: --Oxidative phosphorylation (OXPHOS) deficiency due to hypoxemia or other causes was hypothesized to increase oxygen radical generation, damage mitochondrial DNA (mtDNA), and reduce adenosine triphosphate synthesis, resulting in compensatory OXPHOS gene induction. Therefore, we investigated the levels of mtDNA damage and OXPHOS transcripts in normal and ischemic hearts, and then in other forms of heart disease. DESIGN: --DNA was extracted from the heart and the levels of the common 4977 base pair mtDNA deletion were quantitated as an index for mtDNA damage. Total RNA was extracted from hearts and analyzed for OXPHOS transcript levels. RESULTS: --In control hearts, the 4977 base pair mtDNA deletion appeared at age 40 years and reached a maximum deletion of 0.0035%. Much higher levels were found in ischemic hearts (0.02% to 0.85%), as well as in three of 10 cases with other types of heart disease (0.017% to 0.16%). The OXPHOS transcripts were increased in all diseased hearts. CONCLUSION: --Ischemic hearts have increased mtDNA damage and OXPHOS gene expression, suggesting that mtDNA damage is associated with OXPHOS deficiency. Oxidative phosphorylation defects may also play a role in some other forms of cardiac disease.