Mitochondrial impairment and synaptic dysfunction are associated with neurological defects in iPSCs-derived cortical neurons of MERRF patients.

BACKGROUND: Myoclonic epilepsy with ragged-red fibers (MERRF) syndrome is a rare inherited mitochondrial disease mainly caused by the m.8344A > G mutation in mitochondrial tRNALys gene, and usually manifested as complex neurological disorders and muscle weakness. Currently, the pathogenic mechanism of this disease has not yet been resolved, and there is no effective therapy for MERRF syndrome. In this study, MERRF patients-derived iPSCs were used to model patient-specific neurons for investigation of the pathogenic mechanism of neurological disorders in mitochondrial disease. METHODS: MERRF patient-derived iPSCs were differentiated into excitatory glutamatergic neurons to unravel the effects of the m.8344A > G mutation on mitochondrial bioenergetic function, neural-lineage differentiation and neuronal function. By the well-established differentiation protocol and electrophysiological activity assay platform, we examined the pathophysiological behaviors in cortical neurons of MERRF patients. RESULTS: We have successfully established the iPSCs-derived neural progenitor cells and cortical-like neurons of patients with MERRF syndrome that retained the heteroplasmy of the m.8344A > G mutation from the patients' skin fibroblasts and exhibited the phenotype of the mitochondrial disease. MERRF neural cells harboring the m.8344A > G mutation exhibited impaired mitochondrial bioenergetic function, elevated ROS levels and imbalanced expression of antioxidant enzymes. Our findings indicate that neural immaturity and synaptic protein loss led to the impairment of neuronal activity and plasticity in MERRF neurons harboring the m.8344A > G mutation. By electrophysiological recordings, we monitored the in vivo neuronal behaviors of MERRF neurons and found that neurons harboring a high level of the m.8344A > G mutation exhibited impairment of the spontaneous and evoked potential-stimulated neuronal activities. CONCLUSIONS: We demonstrated for the first time the link of mitochondrial impairment and synaptic dysfunction to neurological defects through impeding synaptic plasticity in excitatory neurons derived from iPSCs of MERRF patients harboring the m.8344A > G mutation. This study has provided new insight into the pathogenic mechanism of the tRNALys gene mutation of mtDNA, which is useful for the development of a patient-specific iPSCs platform for disease modeling and screening of new drugs to treat patients with MERRF syndrome.

Endocrine Challenges in Myoclonic Epilepsy With Ragged Red Fibers Syndrome: A Case Report.

Myoclonic epilepsy with ragged red fibers (MERRF) syndrome is a primary mitochondrial disorder characterized by myoclonus, epilepsy, ataxia, and muscle fiber abnormalities. While traditionally associated with neurological features, MERRF's multisystem nature extends to endocrine dysfunction, including diabetes mellitus, thyroid disorders, and adrenal abnormalities. This case report explores the multifaceted nature of MERRF syndrome by presenting the clinical journey of a 70-year-old woman who sought care at the endocrinology clinic due to coexisting Addison's disease and diabetes mellitus, marked by recurrent hypoglycemia and suboptimal metabolic control. Over time, she developed a history of myoclonic epilepsy, effectively managed with lamotrigine, along with mild sensory axonal polyneuropathy and ataxia. The patient was diagnosed with MERRF syndrome following her son's diagnosis, which had a severe form. This case underscores the intricate interplay between mitochondrial dysfunction and endocrine manifestations in MERRF syndrome, highlighting the importance of a comprehensive and multidisciplinary approach to patient care. MERRF syndrome's array of endocrine manifestations substantially impacts patients' quality of life and morbidity. A comprehensive approach, uniting endocrinologists, neurologists, geneticists, and other specialists, is essential for effective patient care. Further research is warranted to unravel the complex mitochondrial-endocrine interactions in MERRF syndrome, offering potential insights for improved management.

A case of overlap myoclonic epilepsy with ragged-red fibers-Leigh syndrome associated with mitochondrial DNA 8344A>G mutation / 中华神经科杂志

Overlap myoclonic epilepsy with ragged-red fibers (MERRF)-Leigh syndrome is a rare mitochondrial encephalomyopathy. A case of MERRF-Leigh syndrome associated with mitochondrial DNA 8344A>G (m.8344A>G) mutation was reported in this article. The patient has suffered from the disease since 15-year old with myoclonus, exercise intolerance, ataxia, limb weakness, dysphasia, dyspnea, blurred vision and hearing loss. Magnetic resonance imaging revealed lesions on right thalamus, bilateral medulla and lumbar spinal cord and atrophy of cervical spinal cord. Electromyography showed predominantly axonal damage of both sensory nerve and motor nerve. Histochemical analyses revealed ragged red fibers, ragged blue fibers, succinate dehydrogenase-stronghly reactive vessels and decreased cytochrome oxidase activity. Gene tests demonstrated a high level of m.8344A>G mutation and m. 14484T>C mutation. MERRF-Leigh overlap syndrome with m.8344A>G mutation was rare. Bulbar paralysis following myoclonus is the main clinical symptom.

Should we investigate mitochondrial disorders in progressive adult-onset undetermined ataxias?

BACKGROUND: Despite the broad development of next-generation sequencing approaches recently, such as whole-exome sequencing, diagnostic workup of adult-onset progressive cerebellar ataxias without remarkable family history and with negative genetic panel testing for SCAs remains a complex and expensive clinical challenge. CASE PRESENTATION: In this article, we report a Brazilian man with adult-onset slowly progressive pure cerebellar ataxia, which developed neuropathy and hearing loss after fifteen years of ataxia onset, in which a primary mitochondrial DNA defect (MERRF syndrome - myoclonus epilepsy with ragged-red fibers) was confirmed through muscle biopsy evaluation and whole-exome sequencing. CONCLUSIONS: Mitochondrial disorders are a clinically and genetically complex and heterogenous group of metabolic diseases, resulting from pathogenic variants in the mitochondrial DNA or nuclear DNA. In our case, a correlation with histopathological changes identified on muscle biopsy helped to clarify the definitive diagnosis. Moreover, in neurodegenerative and neurogenetic disorders, some symptoms may be evinced later during disease course. We suggest that late-onset and adult pure undetermined ataxias should be considered and investigated for mitochondrial disorders, particularly MERRF syndrome and other primary mitochondrial DNA defects, together with other more commonly known nuclear genes.

Pharmacotherapeutic management of epilepsy in MERRF syndrome.

INTRODUCTION: Epilepsy is a prominent feature of myoclonic epilepsy with ragged-red fibers (MERRF)-syndrome. The most frequent seizure type is myoclonic seizures, of which the treatment is challenging and empiric. AREAS COVERED: Herein, the author summarises and discusses previous and recent findings of antiepileptic drug (AED) treatment in MERRF-syndrome. EXPERT OPINION: MERRF-syndrome is a predominantly maternally inherited, multisystem mitochondrial disorder caused by pathogenic variants predominantly of the mitochondrial DNA (mtDNA). Canonical clinical features of MERRF include myoclonus, epilepsy, ataxia, and myopathy. Additionally, other manifestations in the CNS, peripheral nerves, eyes, ears, heart, gastrointestinal tract, and endocrine organs may occur (MERRF-plus). Today, MERRF is considered rather as myoclonic ataxia than as myoclonic epilepsy. Genotypically, MERRF is due to mutations in 13 mtDNA-located genes and 1 nDNA-located gene. According to the modified Smith-score, the strongest gene-disease relationship exists for MT-TK, MT-TL1, and POLG1. Epilepsy is the second most frequent phenotypic feature of MERRF. Seizure-types associated with MERRF include focal myoclonic, focal clonic, and focal atonic seizures, generalized myoclonic, tonic-clonic, atonic, and myoclonic-atonic seizures, or typical absences. Treatment of myoclonic epilepsy relies on expert judgments recommending levetiracetam, together with clonazepam, or topiramate, zonisamide, or piracetam in monotherapy as the first line AEDs.

MERRF syndrome (Myoclonic Epilepsy with Ragged Red Fibres) presenting with cervicothoracic lipomatosis.

INTRODUCTION: Patients with MERRF syndrome (Myoclonic Epilepsy with Ragged Red Fibres) usually present with encephalomyopathy. However, progressive, recurrent cervicothoracic lipomatosis may be rarely observed. CASE REPORT: The authors report 4 cases of MERRF syndrome associated with lipomatosis. In 3 patients, the diagnosis of MERRF syndrome was established on the basis of the clinical features of the lipomas and clinical interview revealing a personal or family history of lipomas and myopathy. DISCUSSION: In the presence of extensive spinal lipomatosis, the presence of other clinical signs of MERRF syndrome in the patient or the patient's family must be investigated. A diagnosis of MERRF syndrome can guide appropriate genetic counselling.

Myoclonus epilepsy with ragged-red fibers:a case report and literature review / 北京大学学报(医学版)

SUMMARY To demonstrate the clinical manifestation, diagnosis and treatment of myoclonus epilepsy with ragged-red-fibers ( MERRF) , a case of MERRF was presented with review of the literature. A 4-year-7-month-old girl was diagnosed with MERRF. She had tremor, fatigue and developmental delay for more than 2 years. Laboratory tests showed that the serum and urine lactic acid and pyruvic acid increased significantly. Electroencephalogram showed diffuse and focal spike slow wave and slow wave in right central and parietal regions. Electromyogram showed neurological damage. Gene mutational analysis showed mtDNA 8344 A>G mutation. The mutational rate was 78%. Mitochondrial disease MERRF syndrome was diagnosed. Cocktails therapy with vitamins B1, B6, B12, L-carnitine, and coenzyme Q10 was administra-ted to the patient. MERRF is a rare disease. The diagnosis can be made by gene mutational analysis. Cocktail therapy may slow down the deterioration of the disease. Gene therapy is still experimental.

Identification of causative mutations in patients with Leigh syndrome and MERRF by mitochondrial DNA-targeted next-generation sequencing

PURPOSE: Mitochondrial diseases are clinically and genetically heterogeneous disorders, which make their exact diagnosis and classification difficult. The purpose of this study was to identify pathogenic mitochondrial DNA (mtDNA) mutations in 2 Korean families with myoclonic epilepsy with ragged-red fibers (MERRF) and Leigh syndrome, respectively. MATERIALS AND METHODS: Whole mtDNAs were sequenced by the method of mtDNA-targeted next-generation sequencing (NGS). RESULTS: Two causative mtDNA mutations were identified from the NGS data. An m.8344A>G mutation in the tRNA-Lys gene (MT-TK ) was detected in a MERRF patient (family ID: MT132), and an m.9176T>C (p.Leu217Pro) mutation in the mitochondrial ATP6 gene (MT-ATP6) was detected in a Leigh syndrome patient (family ID: MT130). Both mutations, which have been reported several times before in affected individuals, were not found in the control samples. CONCLUSION: This study suggests that mtDNA-targeted NGS will be helpful for the molecular diagnosis of genetically heterogeneous mitochondrial diseases with complex phenotypes.

Distal weakness with respiratory insufficiency caused by the m.8344A > G "MERRF" mutation.

The m.8344A>G mutation in the mt-tRNA(Lys) gene, first described in myoclonic epilepsy and ragged red fibers (MERRF), accounts for approximately 80% of mutations in individuals with MERRF syndrome. Although originally described in families with a classical syndrome of myoclonus, ataxia, epilepsy and ragged red fibers in muscle biopsy, the m.8344A>G mutation is increasingly recognised to exhibit marked phenotypic heterogeneity. This paper describes the clinical, morphological and laboratory features of an unusual phenotype in a patient harboring the m.8344A>G 'MERRF' mutation. We present the case of a middle-aged woman with distal weakness since childhood who also had ptosis and facial weakness and who developed mid-life respiratory insufficiency necessitating non-invasive nocturnal ventilator support. Neurophysiological and acetylcholine receptor antibody analyses excluded myasthenia gravis whilst molecular genetic testing excluded myotonic dystrophy, prompting a diagnostic needle muscle biopsy. Mitochondrial histochemical abnormalities including subsarcolemmal mitochondrial accumulation (ragged-red fibers) and in excess of 90% COX-deficient fibers, was seen leading to sequencing of the mitochondrial genome in muscle. This identified the m.8344A>G mutation commonly associated with the MERRF phenotype. This case extends the evolving phenotypic spectrum of the m.8344A>G mutation and emphasizes that it may cause indolent distal weakness with respiratory insufficiency, with marked histochemical defects in muscle. Our findings support consideration of screening of this gene in cases of indolent myopathy resembling distal limb-girdle muscular dystrophy in which screening of the common genes prove negative.

Treatment of human cells derived from MERRF syndrome by peptide-mediated mitochondrial delivery.

BACKGROUND AIMS: The feasibility of delivering mitochondria using the cell-penetrating peptide Pep-1 for the treatment of MERRF (myoclonic epilepsy with ragged red fibers) syndrome, which is caused by point mutations in the transfer RNA genes of mitochondrial DNA, is examined further using cellular models derived from patients with MERRF syndrome. METHODS: Homogenesis of mitochondria (wild-type mitochondria) isolated from normal donor cells with about 83.5% preserved activity were delivered into MERRF fibroblasts by Pep-1 conjugation (Pep-1-Mito). RESULTS: Delivered doses of 52.5 µg and 105 µg Pep-1-Mito had better delivered efficiency and mitochondrial biogenesis after 15 days of treatment. The recovery of mitochondrial function in deficient cells receiving 3 days of treatment with peptide-mediated mitochondrial delivery was comprehensively demonstrated by restoration of oxidative phosphorylation subunits (complex I, III and IV), mitochondrial membrane potential, adenosine triphosphate synthesis and reduction of reactive oxygen species production. The benefits of enhanced mitochondrial regulation depended on the function of foreign mitochondria and not the existence of mitochondrial DNA and can be maintained for at least 21 days with dramatically elongated mitochondrial morphology. In contrast to delivery of wild-type mitochondria, the specific regulation of Pep-1-Mito during MERRF syndrome progression in cells treated with mutant mitochondria was reflected by the opposite performance, with increase in reactive oxygen species production and matrix metalloproteinase activity. CONCLUSIONS: The present study further illustrates the feasibility of mitochondrial intervention therapy using the novel approach of peptide-mediated mitochondrial delivery and the benefit resulting from mitochondria-organelle manipulation.

Genetics of Mitochondrial Myopathies

Mitochondrion is an intracellular organelle with its own genome. Its function in cellular metabolism is indispensable that mitochondrial dysfunction gives rise to multisystemic failure. The manifestation is most prominent with tissues of high energy demand such as muscle and nerve. Mitochondrial myopathies occur not only by mutations in mitochondrial genome, but also by defects in nuclear genes or secondarily by toxic insult on mitochondrial replication. Currently curative treatment modality does not exist and symptomatic treatment remains mainstay. Administration of L-arginine holds great promise according to the recent reports. Advances in mitochondrial RNA import might enable a new therapeutic strategy.

Clinical spectrum and prognostic factors of acute necrotizing encephalopathy in children.

This study was conducted to investigate the etiology, the clinical characteristics and prognosis of acute necrotizing encephalopathy (ANE) in Korean children. Six children (1 yr to 7 yr) patients with ANE were enrolled. They were diagnosed by clinical and radiological characteristics and their clinical data were retrospectively analyzed. In a search of clinically plausible causes, brain MRI in all patients, mitochondrial DNA studies for mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS) and myoclonus epilepsy and ragged red fibers (MERRF) in four patients, and genomic typing on HLA DRB/HLA DQB genes in three patients were performed. All had precedent illnesses and the main initial symptoms included mental change (83%), seizures (50%), and focal deficits (50%). MRI revealed increased T2 signal density in the bilateral thalami and/or the brainstem in all patients. Mitochodrial DNA studies for MELAS and MERRF were negative in those children and HLA-DRB1*1401, HLA-DRB3*0202, and HLA-DQB1*0502 seemed to be significant. A high dose steroid was given to all patients, which seemed to be partly effective except for 2 patients. In conclusion, ANE is relatively rare, but can result in serious neurological complication in children. Early detection and appropriate treatment may lead to a better neurological outcome.

Mutational analysis of whole mitochondrial DNA in patients with MELAS and MERRF diseases

Mitochondrial diseases are clinically and genetically heterogeneous disorders, which make the exact diagnosis and classification difficult. The purpose of this study was to identify pathogenic mtDNA mutations in 61 Korean unrelated families (or isolated patients) with MELAS or MERRF. In particular, the mtDNA sequences were completely determined for 49 patients. From the mutational analysis of mtDNA obtained from blood, 5 confirmed pathogenic mutations were identified in 17 families, and 4 unreported pathogenically suspected mutations were identified in 4 families. The m.3243A>G in the tRNA(Leu(UUR)) was predominantly observed in 10 MELAS families, and followed by m.8344A>G in the tRNA(Lys) of 4 MERRF families. Most pathogenic mutations showed heteroplasmy, and the rates were considerably different within the familial members. Patients with a higher rate of mutations showed a tendency of having more severe clinical phenotypes, but not in all cases. This study will be helpful for the molecular diagnosis of mitochondrial diseases, as well as establishment of mtDNA database in Koreans.

Clinical Spectrum and Prognostic Factors of Acute Necrotizing Encephalopathy in Children

This study was conducted to investigate the etiology, the clinical characteristics and prognosis of acute necrotizing encephalopathy (ANE) in Korean children. Six children (1 yr to 7 yr) patients with ANE were enrolled. They were diagnosed by clinical and radiological characteristics and their clinical data were retrospectively analyzed. In a search of clinically plausible causes, brain MRI in all patients, mitochondrial DNA studies for mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS) and myoclonus epilepsy and ragged red fibers (MERRF) in four patients, and genomic typing on HLA DRB/HLA DQB genes in three patients were performed. All had precedent illnesses and the main initial symptoms included mental change (83%), seizures (50%), and focal deficits (50%). MRI revealed increased T2 signal density in the bilateral thalami and/or the brainstem in all patients. Mitochodrial DNA studies for MELAS and MERRF were negative in those children and HLA-DRB1*1401, HLA-DRB3*0202, and HLA-DQB1*0502 seemed to be significant. A high dose steroid was given to all patients, which seemed to be partly effective except for 2 patients. In conclusion, ANE is relatively rare, but can result in serious neurological complication in children. Early detection and appropriate treatment may lead to a better neurological outcome.

Histochemical and Molecular Genetic Study of MELAS and MERRF in Korean Patients

Mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episode (MELAS) and myoclonic epilepsy and raggedred fibers (MERRF) are rare disorders caused by point mutation of the tRNA gene of the mitochondrial genome. To understand the pathogenetic mechanism of MELAS and MERRF, we studied four patients. Serially sectioned frozen muscle specimens with a battery of histochemical stains were reviewed under light microscope and ultrastructural changes were observed under electron microscope. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis was performed and the tRNA genes were sequenced to confirm mutations. In two patients with MELAS, strongly succinyl dehydrogenase positive blood vessels (SSVs) and many cytochrome oxidase (COX) positive raggedred fibers (RRFs) were observed, and A3243G mutations were found from the muscle samples. In two patients with MERRF, neither SSV nor COX positive RRFs were seen and A8344G mutations were found from both muscle and blood samples. In the two MERRF families, the identical mutation was observed among family members. The failure to detect the mutation in blood samples of the MELAS suggests a low mutant load in blood cells. The histochemical methods including COX stain are useful for the confirmation and differentiation of mitochondrial diseases. Also, molecular biological study using muscle sample seems essential for the confirmation of the mtDNA mutation.

Molecular Genetic Diagnosis in Korean Patients with Myoclonic Epilepsy with Ragged Red Fiber (MERRF) Syndrome

PURPOSE: Myoclonic epilepsy with ragged red fiber (MERRF) syndrome is a disease of the mitochondrial encephalomyopathies, characterized by progressive myoclonus (action), epilepsy, cerebellar ataxia, intention tremor, muscle weakness, progressive dementia, sensorineural hearing loss and optic atrophy. Its inheritance is maternally inherited mitochondrial mutation, and its pathologic finding is characterized by ragged red fibers (RRF). Biochemically its defects are diverse. This study was undertaken to investigate the pattern of mitochondrial mutation and characterize the clinical features in Korean patients with MERRF. METHODS: We collected 3 clinically suspected MERRF patients from 2 Korean families who have progressive myoclonus, epilepsy, cerebellar ataxia, intention tremor, muscle weakness, progressive dementia etc. We reviewed their clinical findings, electrophysiologic studies, radiologic findings and pathologic findings, retrospectively. Of the 2 patients (case 1 and case 3) who had undergone muscle biopsy, case 1 showed RRF in modified Gomori trichrome staining, increased mitochondrial number and abnormal inclusion body in EM. To examine the pattern of mitochondrial mutation of these patients, molecular study was carried out in 3 patients, 2 mothers, 2 fathers, and 4 siblings. Their genomic DNAs were isolated from peripheral leukocytes, subsequent PCR-direct nucleotide sequencing and Ban II digestion were followed. RESULTS: All mutations in our cases were A to G point mutations in the tRNALys gene at position 8344. CONCLUSION: We confirmed clinically suspected MERRF patients as MERRF and their mothers and siblings as carriers, on the basis of molecular genetic analysis. This study suggests that the molecular genetic analysis can be utilized to diagnose MERRF patients easily and confirm carriers, especially at the presymptomatic stage before the characteristic pathologic changes appear.

Molecular Genetic Diagnosis in Korean Patients with Myoclonic Epilepsy with Ragged Red Fiber (MERRF) Syndrome

PURPOSE: Myoclonic epilepsy with ragged red fiber (MERRF) syndrome is a disease of the mitochondrial encephalomyopathies, characterized by progressive myoclonus (action), epilepsy, cerebellar ataxia, intention tremor, muscle weakness, progressive dementia, sensorineural hearing loss and optic atrophy. Its inheritance is maternally inherited mitochondrial mutation, and its pathologic finding is characterized by ragged red fibers (RRF). Biochemically its defects are diverse. This study was undertaken to investigate the pattern of mitochondrial mutation and characterize the clinical features in Korean patients with MERRF. METHODS: We collected 3 clinically suspected MERRF patients from 2 Korean families who have progressive myoclonus, epilepsy, cerebellar ataxia, intention tremor, muscle weakness, progressive dementia etc. We reviewed their clinical findings, electrophysiologic studies, radiologic findings and pathologic findings, retrospectively. Of the 2 patients (case 1 and case 3) who had undergone muscle biopsy, case 1 showed RRF in modified Gomori trichrome staining, increased mitochondrial number and abnormal inclusion body in EM. To examine the pattern of mitochondrial mutation of these patients, molecular study was carried out in 3 patients, 2 mothers, 2 fathers, and 4 siblings. Their genomic DNAs were isolated from peripheral leukocytes, subsequent PCR-direct nucleotide sequencing and Ban II digestion were followed. RESULTS: All mutations in our cases were A to G point mutations in the tRNALys gene at position 8344. CONCLUSION: We confirmed clinically suspected MERRF patients as MERRF and their mothers and siblings as carriers, on the basis of molecular genetic analysis. This study suggests that the molecular genetic analysis can be utilized to diagnose MERRF patients easily and confirm carriers, especially at the presymptomatic stage before the characteristic pathologic changes appear.