A single bout of resistance exercise triggers mitophagy, potentially involving the ejection of mitochondria in human skeletal muscle.

AIM: The present study aimed to investigate the effects of a single bout of resistance exercise on mitophagy in human skeletal muscle (SkM). METHODS: Eight healthy men were recruited to complete an acute bout of one-leg resistance exercise. SkM biopsies were obtained one hour after exercise in the resting leg (Rest-leg) and the contracting leg (Ex-leg). Mitophagy was assessed using protein-related abundance, transmission electron microscopy (TEM), and fluorescence microscopy. RESULTS: Our results show that acute resistance exercise increased pro-fission protein phosphorylation (DRP1Ser616) and decreased mitophagy markers such as PARKIN and BNIP3L/NIX protein abundance in the Ex-leg. Additionally, mitochondrial complex IV decreased in the Ex-leg when compared to the Rest-leg. In the Ex-leg, TEM and immunofluorescence images showed mitochondrial cristae abnormalities, a mitochondrial fission phenotype, and increased mitophagosome-like structures in both subsarcolemmal and intermyofibrillar mitochondria. We also observed increased mitophagosome-like structures on the subsarcolemmal cleft and mitochondria in the extracellular space of SkM in the Ex-leg. We stimulated human primary myotubes with CCCP, which mimics mitophagy induction in the Ex-leg, and found that BNIP3L/NIX protein abundance decreased independently of lysosomal degradation. Finally, in another human cohort, we found a negative association between BNIP3L/NIX protein abundance with both mitophagosome-like structures and mitochondrial cristae density in the SkM. CONCLUSION: The findings suggest that a single bout of resistance exercise can initiate mitophagy, potentially involving mitochondrial ejection, in human skeletal muscle. BNIP3L/NIX is proposed as a sensitive marker for assessing mitophagy flux in SkM.

Ablation of Sam50 is associated with fragmentation and alterations in metabolism in murine and human myotubes.

The sorting and assembly machinery (SAM) Complex is responsible for assembling ß-barrel proteins in the mitochondrial membrane. Comprising three subunits, Sam35, Sam37, and Sam50, the SAM complex connects the inner and outer mitochondrial membranes by interacting with the mitochondrial contact site and cristae organizing system complex. Sam50, in particular, stabilizes the mitochondrial intermembrane space bridging (MIB) complex, which is crucial for protein transport, respiratory chain complex assembly, and regulation of cristae integrity. While the role of Sam50 in mitochondrial structure and metabolism in skeletal muscle remains unclear, this study aims to investigate its impact. Serial block-face-scanning electron microscopy and computer-assisted 3D renderings were employed to compare mitochondrial structure and networking in Sam50-deficient myotubes from mice and humans with wild-type (WT) myotubes. Furthermore, autophagosome 3D structure was assessed in human myotubes. Mitochondrial metabolic phenotypes were assessed using Gas Chromatography-Mass Spectrometry-based metabolomics to explore differential changes in WT and Sam50-deficient myotubes. The results revealed increased mitochondrial fragmentation and autophagosome formation in Sam50-deficient myotubes compared to controls. Metabolomic analysis indicated elevated metabolism of propanoate and several amino acids, including ß-Alanine, phenylalanine, and tyrosine, along with increased amino acid and fatty acid metabolism in Sam50-deficient myotubes. Furthermore, impairment of oxidative capacity was observed upon Sam50 ablation in both murine and human myotubes, as measured with the XF24 Seahorse Analyzer. Collectively, these findings support the critical role of Sam50 in establishing and maintaining mitochondrial integrity, cristae structure, and mitochondrial metabolism. By elucidating the impact of Sam50-deficiency, this study enhances our understanding of mitochondrial function in skeletal muscle.

Impact of capillary and sarcolemmal proximity on mitochondrial structure and energetic function in skeletal muscle.

Mitochondria within skeletal muscle cells are located either between the muscle contractile apparatus (interfibrillar mitochondria, IFM) or beneath the cell membrane (subsarcolemmal mitochondria, SSM), with several structural and functional differences reported between IFM and SSM. However, recent 3D imaging studies demonstrate that mitochondria are particularly concentrated in the proximity of capillaries embedded in sarcolemmal grooves rather than in proximity to the sarcolemma itself (paravascular mitochondria, PVM). To evaluate the impact of capillary vs. sarcolemmal proximity, we compared the structure and function of skeletal muscle mitochondria located either lateral to embedded capillaries (PVM), adjacent to the sarcolemma but not in PVM pools (SSM) or interspersed between sarcomeres (IFM). Mitochondrial morphology and interactions were assessed by 3D electron microscopy coupled with machine learning segmentation, whereas mitochondrial energy conversion was assessed by two-photon microscopy of mitochondrial membrane potential, content, calcium, NADH redox and flux in live, intact cells. Structurally, although PVM and SSM were similarly larger than IFM, PVM were larger, rounder and had more physical connections to neighbouring mitochondria compared to both IFM and SSM. Functionally, PVM had similar or greater basal NADH flux compared to SSM and IFM, respectively, despite a more oxidized NADH pool and a greater membrane potential, signifying a greater activation of the electron transport chain in PVM. Together, these data indicate that proximity to capillaries has a greater impact on resting mitochondrial energy conversion and distribution in skeletal muscle than the sarcolemma alone. KEY POINTS: Capillaries have a greater impact on mitochondrial energy conversion in skeletal muscle than the sarcolemma. Paravascular mitochondria are larger, and the outer mitochondrial membrane is more connected with neighbouring mitochondria. Interfibrillar mitochondria are longer and have greater contact sites with other organelles (i.e. sarcoplasmic reticulum and lipid droplets). Paravascular mitochondria have greater activation of oxidative phosphorylation than interfibrillar mitochondria at rest, although this is not regulated by calcium.

Hepatomioneuropatía secundaria a depleción del ADN mitocondrial / Hepatomioneuropathy secondary to mitochondrial DNA depletion

Introducción. La depleción del ADN mitocondrial (ADNmt) consiste en la presencia de un menor número de copias del genoma mitocondrial, siendo sus expresiones fenotípicas muy heterogéneas. Caso clínico. Niña de 22 meses de edad que presenta hipotonía generalizada detectada a partir de los 7 meses de edad asociada a hepatoesplenomegalia con moderada elevación de la transaminasemia, sin datos de fracaso hepático funcional ni hipoglucemia hipocetótica; progresivamente se observó el desarrollo de debilidad muscular, fracaso respiratorio, hipertensión arterial, acidosis láctica y superposición de signos miopáticos y neuropáticos en los estudios neurofisiológicos periféricos. El estudio genético molecular para la atrofia muscular espinal fue normal. El examen con microscopía óptica de la biopsia muscular fue compatible con atrofia neurogénica, con presencia en el examen ultraestructural de gotas lipídicas, acúmulos mitocondriales subsarcolémicos y de gránulos de glucógeno. Los complejos de la cadena respiratoria mitocondrial (CRM) en homogenado muscular fueron normales. El estudio genético molecular a nivel muscular demostró la presencia de una depleción del ADNmt. Conclusiones. Esta observación probablemente represente una nueva expresión fenotípica de depleción del ADNmt que se puede denominar forma hepatomioneuropática. La normalidad de la CRM en músculo no excluye la depleción del ADNmt

Introduction. Mitochondrial DNA depletion (mtDNA) is an highly heterogeneous condition characterized by a decresed number of mtDNA copies. Case report. The patient is a 22-month-old girl with generalized hypotonia, marked weakness, respiratory failure, arterial hypertension, hyperlactacidemia, hepatosplenomegaly and mild hypertransaminasemia without hepatic failure neither hypoketotic hypoglycemia. Electromyographic findings were consistent with neuromyopathy and muscle biopsy suggested a neurogenic atrophy. Electron microscopy revealed lipid droplets, subsarcolemmal accumulation of mitochondrias and glycogen granules. Respiratory chain enzime activities were normal. Genetic study in muscle showed mtDNA depletion, and the diagnosis of spinal muscular atrophy caused by survival motoneuron gene deletion was excluded. Conclusions. This case might be a novel phenotype of mtDNA depletion which could be named hepatomioneuropatyc form. A normal result of respiratory chain enzimes in muscle doesn't excluded mtDNA depletion

Tubular Aggregate Myopathy: A Case Report

We report a first Korean case of presumably dominantly inherited primary tubular aggregate myopathy in a 19-yr-old man, who presented with slowly progressive proximal muscle stiffness and weakness. In hematoxylin and eosin stain, it showed subsarcolemmal, or central pale basophilic granular vacuoles, which stained red with modified Gomori's trichrome and intensive blue with nicotinamide adenonine dinucleotide-tetrazolium reductase, respectively. Ultrastructurally, aggregates of 60 nm-sized hexagonal tubules were found in both type 1 and type 2 fibers. We briefly review the pathologic findings of the previously reported cases of tubular aggregate myopathy and discuss the possible pathogenesis of this disease. We briefly discuss the possible pathogenesis of sarcoplasmic reticulum and review the ultrastructural characteristics.

Effects of thyroidectomy on biochemical and ultrastructural aspects of rat slow and fast muscles

Thyroidectomy provoked the reduction of the oxygen consumption rate in state 3 respiration in both Extensor Digitorum Longus (EDL, fast twitch) and Soleus (slow twitch) muscles of rat, being the last one the most affected. Morphological alterations after 8 weeks of thyroidectomy were found in mitochondria. These organelles exhibited cristae swelling and formation of autophagic vacuoles in subsarcolemmal and intermyofibrillar spaces. Altered mitochondria were also seen in the axon terminal and the postjunctional region of motor end-plate

Two cases of mitochondrial myopathy with predominant respiratory dysfunction

Although it is well known that the respiratory failure is a major cause of death in most patients with chronic neuromuscular disease, predominant respiratory dysfunction without severe involvement of limb muscles is an unusual complication of mitochondrial myopathy in adult age. We experienced two cases of mitochondrial myopathy with severe involvement of respiratory function and only mild involvement of limb muscles. One is a 16 year old female and another is a 22 year old male. The diagnosis is based on morphologic characteristics of "ragged red fibers" under the light microscope and abnormal mitochondrias on the electron microscope in the muscle biopsy.

Encefalomiopatía mitocondrial, acidosis láctica y apariencia de accidentes vasculares cerebrales / Mitochondrial, lactic acidosis and features of cerebrovascular disorders

En una paciente de 12 años de edad, aparentemente sana hasta los 9 años, con epilepsia progresiva, deterioro mental, acidosis láctica y déficit neurológicos que simulaban accidentes cerebrovasculares, la biopsia muscular demostró fibras rojas desflecadas y acúmulos de mitocondrias anormales, confirmando el diagnóstico de encefalomiopatía mitocondrial. Conviene pensar en este grupo de enfermedades en pacientes escolares con cefalea periódica, epilepsia mioclónica que responde mal al tratamiento, hipoacusia sensorioneural y deterioro mental. Las manifestaciones musculares pueden ser frecuentemente tardías

Mitochondrial myopathy and myoclonic epilepsy

Os autores descrevem uma família de raça negra (mäe e três filhos) com miopatia mitocondrial. Duas irmäs tinham acidose láctica concomitante e epilepsia mioclônica. Outros achados observados nos membros mais afetados foram demência, ataxia, fraqueza muscular e neuropatia sensitiva. A mäe era assintomática. Um filho sofreu acidente vascular cerebral isquêmico envolvendo a regiäo temporal direita. Todos os membros da família estudados eram hipertensos. EEG mostrou resposta fotomioclônica na paciente probanda. Biópsia muscular mostrou fibers e mitocôndrias anormais ao estudo de microscopia eletrônica. Análise bioquímica mostrou um defeito no citocromo C oxidase nas mitocôndrias extraídas do músculo esquelético de uma paciente afetada. Aspectos clínicos e genéticos sobre as encefalomiopatias mitocondriais säo discutidos

Miopatia por deficiência de succinato citocromo-c redutase: possível defeito no complexo 11 da cadeia respiratória / Myopathy due to succinate cytochrome-c reductase deficiency: possible deficiency in the complex 11 of the respiratory chain

Relato do caso de mulher de 24 anos de idade que apresentava astenia desde a puberdade, com agravamenteo nos últimos anos, cuja biópsia muscular revelou grande acúmulo de mitocôndrias. As dosagens dos enzimas mitocondriais mostrou importante reduçäo da succinato-citocromo-C-redutase, sugerindo defeito na cadeia respiratória a nível do complexo II. Medicada com altas doses de vitamina C e K, melhorou da força muscular. Säo feitas consideraçöes a respeito das principais síndromes com miopatias mitocondriais, bem como a respeito dos métodos de investigaçäo em defeitos da cadeia respiratória

Non-epileptic myclonus and mitochondrial encephalomyopathy

Foram examinados dois irmäos portadores de uma síndrome miclónica progressiva comn discretos sintomas cerebelares. O exame neurológico mostrava sinais cerebelares moderados e papilas pálidas; mioclonais assíncronas, arrítmicas e assimétricas, um déficit artrestésico e ausência de fraqueza muscular. A atividade de base do EEG era moderadamente lenta e sem atividade irritativa. A TC era normal em ambos os casos. A estimulaçäo fótica intemitente aumentava a freqüência dos abalos mioclônicos que se tornavam bilaterais e sincronos, progredindo para uma crise tônico-clônica generalizada. Potenciais evocados e RMN em um caso foram normais . Drogas anticonvulsivantes foram ineficazes no controle das mioclonias. O diagníostico de encefalomiopatia mitocondrial foi realizado através do achado em espécimes musculares de membranas basais espessadas, com degeneraçäo miofibrilar e um número elevado de mitocondrias distribuídos perifericamente e com uma matriz densa, granular e com alguns vacúolos. Os achados clínicos e eletrográficos sugerem uma origem subcortical para esta síndrome mioclônica

Mitochondrial dysfunction in myasthenia gravis: report of a case

É relatado o caso de um paciente de 11 anos de idade com oftalmoparesia extrínseca, ptose palpebral bilateral e tetraparesia desde os 7 anos de idade. A concentraçäo de anticorpos contra receptor de acetilcolina por radioimunoensaio foi 0,6 nM/1; a pesquisa de anticorpos contra músculo estriado foi negativa. Exame eletromiográfico revelou decremento de 26,1%. Foi tratado com brometo de piridostigmina na dose de 60 mg/d. Submetidos a biópsia de tecido muscular estriado (biceps braquial esquerdo) com avaliaçöes por métodos histoquímicos e microscopia eletrônica, que revelaram: acúmulos de mitocôndrias na regiäo subsarcolemal na coloraçäo pela SDH; aumento da concentraçäo de mitocôndrias e corpúsculos eletrodensos a microscopia eletrônica; esses achados säo sugestivos de miopatia mitocondrial. Em conseqüência da interrupçäo das drogas anticolinesterásicas ocorreu piora das manifestaçöes deficitárias, disfagia e dispnéia. Reintroduzidos os anticolinesterásicos, associados a imunossupressäo com corticosteróides, houve melhora e retomada pelo paciente de suas atividades habituais. Destarte, é discutido o caráter inespecífico das alteraçöes morfogenéticas e disfunçöes de mitocôndrias em outras patologias neuromusuclares, incluindo a miastenia grave, como neste casos em particular

Chronic progressive external ophthalmoplegia: II. A qualitative and quantitative electronnicroscopy study of skeletal muscles

Trata-se de estudo quantitativo das principais alteraçöes encontradas em microscopia eletrônica de biópsia de músculos somáticos 31 entre 34 pacientes com síndrome de oftalmoplegia externa crônica e progressiva (OECP). Os pacientes foram divididos em três grupos clínicos - A) 10 casos esporádicos, com OECP e fraqueza muscular; B) 9 casos com historia familiar positiva, OECP e fraqueza muscular; C) 15 casos com OECP, fraqueza muscular e alguns dos seguintes sintomas: retinopatia pigmentar, ataxia cerebrar, sinais piramidais e neuropatia periférica. Foram encontradas alteraçöes em todos os grupos (8 do grupo A. três do grupo B e 14 do grupo C). As avaliaçöes quantitativas de certos constituintes das fibras musculares, usando a técnica de contagem de pontos revelou: diminuiçäo da fraçäo-volumétrica de miofibrilas, aumento das fraçöes-volumétricas de mitocôndria, glicogênio e lípídeos. As fraçöes-volumétricas de mitocndria correlacionam positivamanete com o conteúdo lipídico, com a proporçäo de fibras do tipo 1 e com a percentagem de fibras com aumento da atividade enzimática oxidativa (definidas em estudo anterior). Os três grupos definidos clinicamente näo mostraram diferenças significativas em termos de proporçöes relativas dos constituintes analisados

Electron Microscopic Observations on the Morphological Changes of Rat Mesentric Mast Cells Induced by Morphine HCl

The effects of morphine HCI on the rat mesenteric mast cells were studied with the electron microscopy. The materials were prepared for electron microscopy by osmium tetroxide fixation and embedding in Epon. The rat mesenteric mast cells showed no distinct morphological changes due to morphine HCl, but the mast cell granlues were changed in various ways. For instance, they formed dusters, showed granular lysis, and an appearance of electron transparency. Frequently, some granules appeared in the extracellular space and the boundary of the granules was not evident. From the results mentioned above, it was suggested that rat mesenteric mast cell granules were affected by morphine HCl in the shape, the granular matrix, and the granular boundaries.