Progressive external ophthalmoplegia due to a recurrent de novo m.15990C>T MT-TP (mt-tRNAPro) gene variant

Progressive external ophthalmoplegia is typically associated with single or multiple mtDNA deletions but occasionally mtDNA single nucleotide variants within mitochondrial transfer RNAs (mt-tRNAs) are identified. We report a 34-year-old female sporadic patient with progressive external ophthalmoplegia accompanied by exercise intolerance but neither fixed weakness nor multisystemic involvement. Histopathologically, abundant COX-deficient fibres were present in muscle with immunofluorescence analysis confirming the loss of mitochondrial complex I and IV proteins. Molecular genetic analysis identified a rare heteroplasmic m.15990C>T mt-tRNA^Pro variant reported previously in a single patient with childhood-onset myopathy. The variant in our patient was restricted to muscle. Single muscle fibre analysis identified higher heteroplasmy load in COX-deficient fibres than COX-normal fibres, confirming segregation of high heteroplasmic load with a biochemical defect. Our case highlights the phenotypic variability typically observed with pathogenic mt-tRNA mutations, whilst the identification of a second case with the m.15990C>T mutation not only confirms pathogenicity but shows that de novo mt-tRNA point mutations can arise in multiple, unrelated patients.     

Are mitochondrial DNA deletions causative in chronic progressive external ophthalmoplegia patients?

Twenty-one patients with long standing unexplained ptosis (3), chronic progressive external ophthalmoplegia (CPEO, 16) or Kearns-Sayre syndrome (KSS, 2) were studied for the presence of mitochondrial DNA (mtDNA) deletions and the major disease-associated mtDNA point mutations with the aim of correlating mitochondrial genetic abnormalities with pathogenesis in these patients. Only 52% were found to have a deletion; of these, 82% harboured the 'common deletion'. Two of 2 KSS patients and 9 of 16 CPEO patients were deletion positive. None of the 3 patients with bilateral ptosis only had a deletion. Of those patients with ragged red fibres (RRF) on histology, 69% had a deletion. No disease associated mtDNA point mutation was observed with the exception of the nucleotide (nt) 11084 A-G mutation associated with mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS) in a patient also harbouring the common deletion. The role of deletions in CPEO patients is discussed.     

A novel mutation in the tRNAIle gene (MTTI) affecting the variable loop in a patient with chronic progressive external ophthalmoplegia (CPEO)

We describe a 62-year-old woman with chronic progressive external ophthalmoplegia (CPEO), multiple lipomas, diabetes mellitus, and a novel mitochondrial DNA (mtDNA) mutation at nucleotide 4302 (4302A>G) of the tRNA^Ile gene (MTTI). This is the first mutation at position 44 in the variable loop (V loop) of any mitochondrial tRNA.The muscle biopsy revealed 10% ragged-red/ragged-blue fibers and 25% cytochrome c oxidase (COX)-deficient fibers.No deletions or duplications were detected by Southern blot analysis. The 4302A>G transition was present only in the patient’s muscle and single-fiber analysis revealed significantly higher levels of the mutation in COX-deficient than in normal fibers. Like tRNA^Leu(UUR), tRNA^Ile appears to be a “hot spot” for mtDNA mutations causing CPEO.     

A novel mitochondrial m.4414T>C MT-TM gene variant causing progressive external ophthalmoplegia and myopathy

We report a novel mitochondrial m.4414T>C variant in the mt-tRNA^Met (MT-TM) gene in an adult patient with chronic progressive external ophthalmoplegia and myopathy whose muscle biopsy revealed focal cytochrome c oxidase (COX)-deficient and ragged red fibres. The m.4414T>C variant occurs at a strongly evolutionary conserved sequence position, disturbing a canonical base pair and disrupting the secondary and tertiary structure of the mt-tRNA^Met. Definitive evidence of pathogenicity is provided by clear segregation of m.4414T>C mutant levels with COX deficiency in single muscle fibres. Interestingly, the variant is present in skeletal muscle at relatively low levels (30%) and undetectable in accessible, non-muscle tissues from the patient and her asymptomatic brother, emphasizing the continuing requirement for a diagnostic muscle biopsy as the preferred tissue for mtDNA genetic investigations of mt-tRNA variants leading to mitochondrial myopathy.     

A case of chronic progressive external ophthalmoplegia associated with familial hypercholesterolemia]

A 52-year-old woman with chronic progressive external ophthalmoplegia (CPEO) with familial hypercholesterolemia (FH) was reported. Her mother died from heart disease, and her elder sister has hypercholesterolemia with swelling of Achilles tendons. She had slowly progressive external ophthalmoplegia, bilateral ptosis, swelling of Achilles tendons since twenties. At 40 years of age, she was pointed out hypercholesterolemia. Physical examination was within normal limits except for bilateral swelling of Achilles tendons. Neurological findings showed bilateral ptosis, disturbance of eye movements, mild proximal muscle weakness and dysesthesia in bilateral hands. Routine laboratory findings were within normal limits except for high serum cholesterol level (512 mg/dl). In the biopsied muscle, there was mild variation in fiber size with several ragged-red fibers and focal cytochrome c oxidase defective fibers. Biochemical analysis of the biopsied muscle revealed normal values in the mitochondrial fraction. Southern blot analysis of the mitochondrial DNA (mtDNA) of the muscle disclosed mixed population of mtDNA, consisting of the normal one and partially deleted (4.9-kilobase). Southern blot analysis of the leukocytes from the patient against the cDNA of LDL receptor was normal at least using the restriction enzyme of BglII, XbaI, EcoRI, PvuII and BamHI. This case has CPEO with deleted mtDNA associated with familial hypercholesterolemia, which is caused to nuclear DNA abnormalities, and is thought to be an important case for us to study the relationship between deleted mtDNA and abnormal nuclear DNA in CPEO.     

Ophthalmoplegia due to mitochondrial DNA disease: the need for genetic diagnosis

We describe a patient with chronic progressive external ophthalmoplegia (CPEO) who underwent muscle biopsy for suspected mitochondrial disease. In spite of normal histocytochemical cytochrome c oxidase (COX) activity and respiratory chain enzyme measurements in muscle, subsequent molecular genetic analysis revealed the presence of a single, large-scale deletion of mitochondrial DNA (mtDNA). The case serves to illustrate the importance of pursuing the proposed mitochondrial genetic abnormality, even in patients with normal biopsy findings.     

A novel heteroplasmic tRNA(Ser(UCN)) mtDNA point mutation associated with progressive external ophthalmoplegia and hearing loss

We sequenced all mitochondrial tRNA genes from a patient with sporadic external ophthalmoplegia (PEO) and 5% COX-negative fibers in muscle biopsy, who had no detectable large mtDNA deletions. Direct sequencing showed a heteroplasmic mutation at nucleotide 7506 in the dihydrouridine stem of the tRNA(Ser(UCN)) gene. RFLP analysis confirmed that 30% of muscle and 20% of urinary epithelium mtDNA harbored the mutation, which was absent in other tissues of the proband as well as in mtDNA of his mother and 100 patients with various encephalomyopathies. Several point mutations on mitochondrial tRNA genes have been reported in PEO patients without large-scale rearrangements of mtDNA but no point mutations have hitherto been found in the gene coding for tRNA(Ser(UCN)).     

Genotype to phenotype correlations in mitochondrial encephalomyopathies associated with the A3243G mutation of mitochondrial DNA

We studied 22 subjects carrying the A3243G point mutation of human mitochondrial DNA (mtDNA). In 14 cases the clinical phenotype was characterized by mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), while 8 patients had chronic progressive external ophthalmoplegia (CPEO). The proportion of A3243G heteroplasmy in muscle was determined by two methods: densitometry on a diagnostic restriction-fragment length polymorphism and solid-phase mini-sequencing. We found a highly significant inverse correlation between the percentage of A3243G mutation and the specific activity of complex 1, the respiratory complex with the highest number of mtDNA-encoded subunits, suggesting a direct effect of the mutation on mtDNA translation. No correlation was observed between the percentage of mutated mtDNA and the presence or absence of specific clinical features, such as stroke, ophthalmoplegia and diabetes mellitus. However, in the MELAS group the percentage of mutated mtDNA molecules was strongly correlated with the age of onset, while no such correlation was found in the CPEO group, suggesting a different time-dependent evolution of the mutation in the two groups. Finally, in contrast with other mtDNA mutations associated with ragged-red fibres (RRF), in both MELAS³²⁴³ and CPE0³²⁴³ we observed a high proportion of RRF that were positive to the histochemical reaction to cytochromec oxidase, a morphological feature that seems to be specific for the neuromuscular phenotypes associated with mutations affecting the tRNA^Leu(UUR) gene.     

Late-onset mitochondrial myopathy with dystrophic changes due to a G7497A mutation in the mitochondrial tRNASer (UCN) gene

Mutations of mitochondrial tRNA genes are usually associated with multi-systemic disorders with onset of symptoms in childhood or early adulthood. Dystrophic myopathic changes are not typical features of these disorders. We report two siblings with a severe progressive myopathy of late onset without external ophthalmoplegia and without involvement of the central and peripheral nervous system. Muscle biopsy specimens showed severe myopathic changes similar to those found in muscular dystrophies. Molecular analysis revealed a G7497A mutation in the mitochondrial tRNA^Ser(UCN) gene. In both patients, the proportion of mutated mitochondrial DNA in muscle was more than 97%. Mitochondrial disorder associated with the G7497A mutation has to be included into the differential diagnosis of severe progressive late-onset myopathy with histopathological dystrophic myopathic changes. Mitochondrial myopathy and high level of mutated mtDNA might be a characteristic of the G7497A tRNA^Ser(UCN) mutation.     

Mitochondrial encephalomyopathy with autosomal dominant inheritance: A clinical and genetic entity of mitochondrial diseases

We report a Japanese family with chronic progressive external ophthal-moplegia (CPEO) with autosomal dominant inheritance, and review 54 reported CPEO patients in seven families (including the present family) with autosomal dominant inheritance and mtDNA deletions in the skeletal muscle. Mean age at onset in the CPEO was 26 years, which is older than that in published solitary cases. In addition to blepharoptosis and external ophthalmoplegia, proximal muscle atrophy and weakness were found in 62%, hearing loss in 25%, and ataxia in 17% of the patients. Retinal degeneration was not found, and cardiac involvement was very rare. mtDNA deletions in the muscle were multiple and large scale, and all such deletions were located in the non–D-loop region. Autosomal dominant CPEO has unique clinical features which differ from those of solitary CPEO, and is associated with multiple large-scale mtDNA deletions. Thus, autosomal dominant CPEO can be considered a clinical and genetic entity of mitochondrial diseases. © 1995 John Wiley & Sons, Inc.     

Frequency of dystrophic muscle abnormalities in chronic progressive external ophthalmoplegia: analysis of 86 patients

BACKGROUND: There are few reports describing the coexistence of dystrophic features with those typical of mitochondrial myopathies in muscle biopsy. A recent study suggested that dystrophic features are frequent in patients with chronic progressive external ophthalmoplegia (CPEO) with a high mutation load, but the actual frequency of these abnormalities in CPEO remains undetermined. OBJECTIVE: To review the occurrence of dystrophic abnormalities in a large series of patients with CPEO to assess the frequency of such abnormalities and to verify whether they are correlated with specific mitochondrial DNA (mtDNA) mutations. METHODS: Retrospective survey of case series (86 patients with CPEO). RESULTS: Only three cases with dystrophic abnormalities were found: two with a large scale mtDNA deletion and one with the A3251G mutation. All three patients showed predominantly proximal muscular weakness resembling limb girdle muscular dystrophy. CONCLUSIONS: Dystrophic abnormalities are rare in CPEO and are not correlated with a specific molecular defect.     

Multiple mitochondrial DNA deletions in hereditary inclusion body myopathy

We have recently described an autosomal dominant hereditary inclusion body myopathy (h-IBM). Clinically it is is characterized by congenital joint contractures and slowly progressive, proximal muscle weakness and ophthalmoplegia. There is deterioration of muscle function between 30 and 50 years of age. While young patients show minor pathological changes in muscle, the middle-aged and old patients show rimmed vacuoles and inclusions of filaments measuring 15–18 nm in diameter. Except for the absence of significant inflammation the histopathology is similar to that found in sporadic inclusion body myositis (s-IBM). In s-IBM mitochondrial alterations including cytochrome c oxidase (COX) -deficient muscle fibers are common. These are due to multiple mitochondrial DNA (mtDNA) deletions. In this study we investigated the occurrence of mitochondrial alterations in autosomal dominant h-IBM. Young affected individuals showed no mitochondrial changes but three patients aged 38, 51 and 59 years, respectively, showed ragged red fibers and COX-deficient muscle fibers. Polymerase chain reaction analysis showed multiple mtDNA deletions. By in situ hybridization clonal expansions of mtDNA with deletions were demonstrated in COX-deficient muscle fibers. Most of the analyzed deletion breakpoints showed nucleotide repeats flanking the deletions. The results show that COX-deficient muscle fibers and somatic mtDNA deletions are present in this family with h-IBM. The same factors may be involved in the development of mtDNA deletions in s-IBM and this family with h-IBM. Received: 13 July 1999 / Revised: 6 October 1999 · Accepted: 12 October 1999     

Sensorineural hearing loss in patients with chronic progressive external ophthalmoplegia or Kearns–Sayre syndrome

In the present study we assessed the prevalence and nature of hearing loss in patients with chronic progressive external ophthalmoplegia (CPEO) or Kearns–Sayre syndrome (KSS) due to single large–scale mitochondrial DNA (mtDNA) deletion or mtDNA tRNA ^{Leu (UUR)} A3243G point mutation (A3243G PM). 14 patients with mtDNA deletion and three patients with A3243G PM underwent audiological evaluation comprising pure–tone and speech audiometry as well as transient evoked otoacoustic emissions (OAE). Audiological evaluation revealed hearing impairment in 10/17 patients. Hearing loss was mild to moderate predominantly affecting high frequencies in five patients with subjective hearing problems (three patients with mtDNA deletions, two patients with A3243G PM). Subclinical hearing deficits restricted to high frequencies were seen in further five asymptomatic patients (four patients with mtDNA deletions, one patients with A3243G PM). Audiological findings suggested a cochlear origin of hearing loss in all subjects. Our results demonstrate that CPEO or KSS patients due to mtDNA deletion or A3243G PM are at high risk of developing sensorineural hearing deficits.     

Distinct segregation of the pathogenic m.5667G>A mitochondrial tRNAAsn mutation in extraocular and skeletal muscle in chronic progressive external ophthalmoplegia

Chronic progressive external ophthalmoplegia (CPEO) is a frequent clinical manifestation of disorders caused by pathogenic mitochondrial DNA mutations. However, for diagnostic purposes skeletal muscle tissue is used, since extraocular muscle tissue is usually not available for work-up. In the present study we aimed to identify causative factors that are responsible for extraocular muscle to be primarily affected in CPEO. We performed comparative histochemical and molecular genetic analyses of extraocular muscle and skeletal muscle single fibers in a case of isolated CPEO caused by the heteroplasmic m.5667G>A mutation in the mitochondrial tRNA^Asn gene (MT-TN). Histochemical analyses revealed higher proportion of cytochrome c oxidase deficient fibers in extraocular muscle (41%) compared to skeletal muscle (10%). However, genetic analyses of single fibers revealed no significant difference either in the mutation loads between extraocular muscle and skeletal muscle cytochrome c oxidase deficient single fibers (extraocular muscle 86% ± 4.6%; skeletal muscle 87.8 %± 5.7%, p = 0.246) nor in the mutation threshold (extraocular muscle 74% ± 3%; skeletal muscle 74% ± 4%). We hypothesize that higher proportion of cytochrome c oxidase deficient fibers in extraocular muscle compared to skeletal muscle might be due to facilitated segregation of the m.5667G>A mutation into extraocular muscle, which may explain the preferential ocular manifestation and clinically isolated CPEO.     

Neuromuscular disease presentation with three genetic defects involving two genomes

An extensive range of molecular defects have been identified in the human mitochondrial genome (mtDNA), many associated with well-characterised, progressive neurological syndromes. We describe a patient who presented to a mitochondrial clinic with progressive bilateral ptosis, external opthalmoplegia and increasing difficulty with walking. He had previously been diagnosed with a dominant, demyelinating polyneuropathy due to PMP22 gene duplication and had also developed gout, presenting in acute renal failure, due to an X-linked recessive HPRT gene mutation. Muscle biopsy revealed many COX-deficient fibres which we show contain high levels of a third genetic defect – a novel, mitochondrial tRNA^Leu(CUN) (MTTL2) gene mutation.     

慢性进行性眼外肌麻痹的临床、病理及诊断（附6例报道）

目的:探讨慢性进行性眼外肌麻痹(CPEO)的临床、病理及诊断。方法:对6例CPEO患者的临床表现、病理特点进行分析并与另5组CPEO比较。结果:6例患者中男3例,女3例,平均起病年龄13岁。2例为同胞兄弟。6例均有进行性加重的双睑下垂和眼球活动障碍,其中1例起病不对称。除眼外肌麻痹外,3例闭目肌力减退、1例轻度吞咽困难、1例轻度肢体无力。3例伴内分泌功能异常。肌活检破碎红纤维(RRF)阳性肌纤维数和细胞色素氧化酶(cytochromeoxidase,COX)阴性肌纤维数均明显>2%。结论:CPEO的主要临床表现为进行性发展的眼外肌麻痹,可伴肢体肌无力、视网膜色素变性、听力障碍、心脏传导异常、内分泌异常等。诊断主要依赖临床和肌肉活检中发现>2%的RRF 肌纤维、COX-肌纤维。     

Residual muscle cytochrome c oxidase activity accounts for submaximal exercise lactate threshold in chronic progressive external ophthalmoplegia

The data from histological, biochemical, and mitochondrial DNA (mtDNA) studies of muscle biopsies from 10 patients affected with chronic progressive external ophthalmoplegia (CPEO) were related to dynamic and metabolic parameters of incremental submaximal exercise. Maximum power output was reduced in all patients as compared to controls. Analysis of the venous lactate curve during exercise revealed a lactate threshold at exercise levels ranging from 40 to 50% of the predicted maximal power output. An earlier significant increase in lactate could be detected by calculating the mean δ lactate. Lactate values were inversely correlated with the cytochrome c oxidase (COX) activity of isolated muscle mitochondria. No relationship was found between lactate values and the number of ragged red fibers, or cytochrome c oxidase-negative fibers or the proportion of deleted mtDNA measured in muscle biopsy specimens. The discussion underscores the value of lactate kinetics in assessing skeletal muscle function, as well as the use of muscle COX levels to predict the effectiveness of wild-type complementation of deleted skeletal muscle mtDNA in in vivo contractile performance of CPEO subjects. © 1996 John Wiley & Sons, Inc.     

Mitochondrial myopathies: divergences of genetic deletions,biochemical defects and the clinical syndromes

Summary Genomic Southern analysis of muscle mitochondrial (mt) DNA from 16 patients with mitochondrial myopathies was performed; 14 of 16 patients had chronic progressive external ophthalmoplegia (CPEO), while 2 patients had mitochondrial myopathies without CPEO. Eleven patients with CPEO, including 5 who exhibited the complete triad of symptoms characteristic of the Kearns-Sayre syndrome (i.e. CPEO, retinal degeneration and heart block) had hetero-plasmic mtDNA with deletions ranging from 2.0 to 8.0 kb in length. There was no clear-cut correlation between the size and location of the deletions, on the one hand, and the histo-chemical and biochemical data or the severity of the disease, on the other.     

Progressive fat replacement of muscle contributes to the disease mechanism of patients with single,large-scale deletions of mitochondrial DNA

Muscle dysfunction in mitochondrial myopathy is predominantly caused by insufficient generation of energy. We hypothesise that structural changes in muscles could also contribute to their pathophysiology. The aims of this study were to determine fat fractions and strength in selected muscles in patients with chronic progressive external ophthalmoplegia (CPEO), and compare progression of muscle fat fraction with age in individuals with CPEO vs. healthy controls and patients with the m.3243A>G mutation of mitochondrial DNA (mtDNA). Seventeen patients with CPEO and single large-scale deletions of mtDNA, 52 healthy controls, and 12 patients carrying the m.3243A>G mtDNA mutation were included. Muscle fat fractions were measured from cross-sections of paraspinal and leg muscles. Peak muscle strength was assessed from a static dynamometer. There was a direct correlation between age and fat fraction in all muscle groups in CPEO patients and healthy controls (p < 0.05). Analysis of covariance showed a higher progression rate of fat replacement in CPEO patients vs. healthy controls in studied muscle groups (p < 0.05). Patients with the m.3243A>G mutation had slower progression rates of fat replacement. Muscle strength decreased with increasing muscular fat fraction in CPEO patients, no correlation was seen in other groups. This indicates that structural muscle changes contribute to the phenotype of older patients affected by CPEO and large-scale deletions. It should therefore be considered, along with known energy deficiencies, as the cause of exercise intolerance.     

Late-onset mitochondrial disorder with electromyographic evidence of myotonia

We describe a patient with chronic progressive external ophthalmoplegia (CPEO) due to a deletion of mitochondrial DNA (mtDNA) who had electromyographic evidence of myotonic discharges. Myotonia has not previously been described in association with mitochondrial disease and this report extends the known phenotypic expression of these disorders.