[DNA diagnosis of Leber's hereditary optic neuropathy performed at Keio University Hospital].

PURPOSE: To learn the clinical value of DNA diagnosis for Leber's hereditary optic neuropathy (LHON), we reviewed the results of DNA diagnosis performed at Keio University Hospital. METHODS AND PATIENTS: Included were 224 patients, 87 patients at Keio University Hospital and 137 patients from other clinics, with bilateral optic neuropathy who were suspected of having LHON. With informed consent, the 3460, 9804, 11,778, 13,730, and 14,484 mutations of mitochondria DNA (mt-DNA) were examined form 1990 to 1998. Percentage of male patients, age at onset of the disease, and percentage of familial history were compared between patients with and without the mutations. The clinical diagnosis at the time of DNA analysis were examined in patients without the mutation. RESULTS: Seventy two(32%) of the 224 patients had one of the five mtDNA mutations, 63(88%) patients had the 11,778 mutation, 6(8%) had the 14,484 mutation, and 3(4%) had the 3460 mutation. In 72 patients with one of the LHON mutations, 89% of the patients were male, the average age of the disease onset was 24.3 years, and 42% of the patients had a familial history of the disease. Eighty (53%) of 152 patients who did not have one of the 5 mutations were diagnosed as having bilateral optic atrophy with unknown causes. CONCLUSION: Although DNA diagnosis of LHON is a useful clinical test, we must know the clinical characteristics of the disease, before taking advantage of this analysis.

Novel mutation in RP2 gene in two brothers with X-linked retinitis pigmentosa and mtDNA mutation of leber hereditary optic neuropathy who showed marked differences in clinical severity.

PURPOSE: To report the identification of a novel mutation of the RP2 gene in two Japanese brothers with X-linked retinitis pigmentosa of a differing clinical severity. The mother was a carrier of both retinitis pigmentosa and optic atrophy. METHODS: The older brother had a severe form of retinitis pigmentosa associated with macular degeneration and total optic atrophy, whereas the younger brother presented typical X-linked retinitis pigmentosa. RESULTS: Each patient exhibited a novel 2-bp insertion at codon 278 in exon 3 of the RP2 gene as well as a 11778 mutation in mitochondrial DNA. This suggests that the older brother may have developed Leber hereditary optic neuropathy as well as retinitis pigmentosa. CONCLUSION: Molecular testing confirmed the clinical diagnosis in each case. However, such testing did not explain the differences in the severity of the ophthalmoscopic findings between the two brothers.

Progression of visual field loss in patients with retinitis pigmentosa of sporadic and autosomal recessive types.

PURPOSE: We examined the natural course of patients with retinitis pigmentosa of the eight sporadic and five autosomal recessive forms over 5 years. METHODS: We measured the areas of the visual fields by Goldmann perimetry using a digitizer and a computer software. RESULTS: The visual field of V-4 isopters in 4 sporadic cases was approximately 200 cm2 during 30 years after the initial examination, but decreased down to 40 cm2 in the next 10 years. The visual field was reduced to half the normal field in 3 autosomal recessive cases early below the age of 25 years. In 4 sporadic and 2 autosomal recessive cases, the inferior temporal visual field was the widest at the onset of the disease, but exhibited the most severe loss. The superior nasal area was the narrowest initially, and showed the mildest progression. CONCLUSIONS: The visual field in retinitis pigmentosa is constricted age-dependently with severe loss of the inferior temporal visual area and mild damage to the superior nasal area.

Quantitative determination of heteroplasmy in Leber's hereditary optic neuropathy by single-strand conformation polymorphism.

PURPOSE: The maternal inheritance of Leber's hereditary optic neuropathy (LHON) is caused by defects in the genes of mitochondrial DNA (mtDNA). The most prevalent mtDNA mutation, present in 40% to 90% of families with this disease, is a G to A substitution at nucleotide position 11778. The rapid and accurate quantification of heteroplasmy of this mutation will help determine the relative risk for disease expression. METHODS: The authors conducted screening tests for heteroplasmy in 44 visually affected patients with the 11778 mutation and 34 unaffected members of 36 Japanese families with LHON using the single-strand conformation polymorphism analysis. This method can detect even a single base difference between the sequences of wild type and mutant DNA strands. The percentage of mutant mtDNA was calculated using an image analyzer. RESULTS: Single-strand conformation polymorphism analysis allowed the detection of heteroplasmy ranging from 5% to 95%. Five (14%) of the 36 families showed heteroplasmy, and 14 (18%) of the 78 persons tested had heteroplasmy ranging from 10% to 94%. Seven patients with heteroplasmy with visual loss had mutant mtDNA ranging from 62% to 94%. CONCLUSIONS: Single-strand conformation polymorphism analysis is rapid, efficient, and accurate for detecting point mutations and quantifying heteroplasmy in mtDNA. Individuals with heteroplasmy with less than 60% of mutant mtDNA in circulating leukocytes are probably at lesser risk for developing optic atrophy.

[Molecular genetic analysis of Leber's hereditary optic neuropathy with the 3460 mutation in Japanese pedigrees].

We have identified a point mutation at nucleotide position 3460 in the ND1 gene of complex I in a Japanese pedigree with Leber's hereditary optic neuropathy by sequencing the ND genes in mitochondrial DNA. None of the 60 healthy Japanese had the 3460 mutation. The proband and his mother also had the 7444 mutation in the COI gene of complex IV and became nearly blind at age 19 with visual acuities of 0.02 OD and 0.04 OS We screened 30 patients with bilateral optic atrophy for the 3460 mutation, and identified one male patient who had the 3460 mutation in heteroplasmic fashion without carrying the 7444 mutation. He lost his sight at age 14 but it recovered to 1.2 OD and 0.7 OS about two years and half after the onset. The difference in final visual acuity between these two patients may reflect the degree of reduction in mitochondrial energy production.

Lack of differences among mitochondrial DNA in family members with Leber's hereditary optic neuropathy and differing visual outcomes.

Investigation of a maternal family of three generations of Leber's hereditary optic neuropathy (LHON) showed four affected and three unaffected individuals. Two of the four patients had recovered near-normal vision, one spontaneously, and one following treatment with idebenone, a quinol compound. One patient whose visual impairment persisted was a heavy consumer of alcohol and tobacco. Molecular genetic analysis of 12 known primary or secondary mutations in mitochondrial DNA (mtDNA) associated with LHON revealed only the 11778 mutation in a homoplasmic fashion with no secondary mutations. The variations in clinical outcome thus could not be explained by synergistically interacting secondary mutations in mtDNA. Environmental factors may play an etiologic role in the development of optic atrophy.

Risk of false-positive molecular genetic diagnosis of Leber's hereditary optic neuropathy.

PURPOSE/METHODS: The most common pathogenic mitochondrial mutation at nucleotide 11778 in Leber's hereditary optic neuropathy is usually detected by the loss of an SfaNI restriction site. To evaluate a false-positive diagnostic error in this molecular genetic assay, we investigated SfaNI polymorphism in 120 patients with bilateral optic atrophy. RESULTS/CONCLUSIONS: The ratio of false-positive to true-positive results was 1:36. Mitochondrial DNA polymorphism at nucleotide 11779 reflects a false-positive genetic error.

Clinical features of Japanese Leber's hereditary optic neuropathy with 11778 mutation of mitochondrial DNA.

The G to A transition of nucleotide position (nt) 11778 of mitochondrial DNA (mtDNA) has been frequently observed in Japanese Leber's hereditary optic neuropathy (LHON) cases. Therefore, we performed a multi-institutional study in Japan of LHON cases with this 11778 mutation of the mtDNA. Genetic and clinical data on 108 cases (90 affected and 18 carriers) in 79 unrelated families were obtained from 64 Japanese institutions. Detection of the nt11778 mutation was performed using restriction enzymes (74 cases) or dot blot with allele specific oligonucleotide (34 cases). Heteroplasmy was observed in 13 of the 90 affected cases and in 8 of the 18 carrier cases. Forty-five families had family history of LHON (44 maternal inheritance, 1 undetermined), and in 28 families (35.9%) there were isolated cases. The male-to-female ratio in the affected was 82:7 (92.1% male). The age at onset of visual loss ranged from 7 to 59 years (average: 23.4 years). All cases had bilateral involvement except one case with a blind eye resulting from ocular infection during childhood. Onset interval between the two eyes ranged from simultaneous to 17 months (average: 2.5 months), in 91.3% of cases being under 6 months. Visual acuity was 0.1 or worse in 152 (85.9%) of 177 eyes, only 6 eyes showing over 0.5. Progression of visual loss ranged from 0 to 48 months (average: 6.2 months). Central visual field abnormality was observed in 162 eyes (96.4%) of 168 eyes. Nonsuspect fundus in the ophthalmoscopic examination constituted 22.8% of eyes. Systemic corticosteroid was given to 45 (52.9%) of 85 cases and visual acuity was improved in only 2 cases (4.4%). Arrhythmia, neurological and muscular abnormality were observed as rare general complications. The present survey indicates that the male-to-female ratio is higher than the previous Japanese LHON statistics and that the visual outcome is better than in American LHON cases with the 11778 mutation.

High frequency of mutations at position 11778 in mitochondrial ND4 gene in Japanese families with Leber's hereditary optic neuropathy.

We have investigated the presence of a point mutation at position 11778 in the ND4 gene of mitochondrial DNA in 17 Japanese families with Leber's hereditary optic neuropathy (LHON), and have identified the mutation in 14 (82.4%) of the 17 families. The prevalence of this mutation appears to be much higher in Japanese patients with LHON than in patients of other ethnic origins, such as Finnish, Dutch, German, and English families.

Mitochondrial DNA analysis of Leber's hereditary optic neuropathy.

A mitochondrial DNA (mtDNA) mutation associated with Leber's hereditary optic neuropathy (LHON) was recently observed. The presence or absence of the mutation was analyzed in 10 Japanese patients whose clinical course and fundus findings were consistent with LHON. Four of them had at least one maternally related individual who also had bilateral optic atrophy, and were diagnosed as "definite LHON". The other 6 cases lacked any record of optic nerve disease in maternally related individuals, and were diagnosed as "possible LHON". We found the mutation at the SfaNI site of mtDNA in 3 out of the former 4 cases, and in 5 out of the latter 6 cases. This result demonstrates the clinical and diagnostic importance of mtDNA analysis, especially with possible cases of LHON, and suggests that an alternative mutation associated with LHON is also present in Japanese patients.