[Current and Future Status of Muscle Pathology: The Position of Muscle Pathology Diagnosis in the Future].

Many muscle disease names are mostly based on muscle pathology findings. Naturally, muscle pathology is important in the diagnosis of muscle diseases. Moreover, in recent years, extensive genetic analysis and autoantibody testing for myositis have been applied clinically, although muscle biopsies are less performed. However, muscle pathology should be proactively considered when a single gene presents multiple phenotypes, when variants of unknown pathological significance are detected, or in cases of autoimmune myositis that may be misdiagnosed as muscular dystrophy.

Genetic analysis of 37 cases with primary periodic paralysis in Chinese patients.

BACKGROUND: Primary periodic paralysis (PPP) is an inherited disorders of ion channel dysfunction characterized by recurrent episodes of flaccid muscle weakness, which can classified as hypokalemic (HypoPP), normokalemic (NormoPP), or hyperkalemic (HyperPP) according to the potassium level during the paralytic attacks. However, PPP is charactered by remarkable clinical and genetic heterogeneity, and the diagnosis of suspected patients is based on the characteristic clinical presentation then confirmed by genetic testing. At present, there are only limited cohort studies on PPP in the Chinese population. RESULTS: We included 37 patients with a clinical diagnosis of PPP. Eleven (29.7%) patients were tested using a specific gene panel and 26 (70.3%) by the whole-exome sequencing (WES). Twenty-two cases had a genetic variant identified, representing a diagnostic rate of 59.5% (22/37). All the identified mutations were either in the SCN4A or the CACNA1S gene. The overall detection rate was comparable between the panel (54.5%: 6/11) and WES (61.5%: 16/26). The remaining patients unresolved through panel sequencing were further analyzed by WES, without the detection of any mutation. The novel atypical splicing variant c.2020-5G > A affects the normal splicing of the SCN4A mRNA, which was confirmed by minigene splicing assay. Among 21 patients with HypoPP, 15 patients were classified as HypoPP-2 with SCN4A variants, and 6 HypoPP-1 patients had CACNA1S variants. CONCLUSIONS: Our results suggest that SCN4A alleles are the main cause in our cohort, with the remainder caused by CACNA1S alleles, which are the predominant cause in Europe and the United States. Additionally, this study identified 3 novel SCN4A and 2 novel CACNA1S variants, broadening the mutation spectrum of genes associated with PPP.

Challenges and Considerations of Preclinical Development for iPSC-Based Myogenic Cell Therapy.

Cell therapies derived from induced pluripotent stem cells (iPSCs) offer a promising avenue in the field of regenerative medicine due to iPSCs' expandability, immune compatibility, and pluripotent potential. An increasing number of preclinical and clinical trials have been carried out, exploring the application of iPSC-based therapies for challenging diseases, such as muscular dystrophies. The unique syncytial nature of skeletal muscle allows stem/progenitor cells to integrate, forming new myonuclei and restoring the expression of genes affected by myopathies. This characteristic makes genome-editing techniques especially attractive in these therapies. With genetic modification and iPSC lineage specification methodologies, immune-compatible healthy iPSC-derived muscle cells can be manufactured to reverse the progression of muscle diseases or facilitate tissue regeneration. Despite this exciting advancement, much of the development of iPSC-based therapies for muscle diseases and tissue regeneration is limited to academic settings, with no successful clinical translation reported. The unknown differentiation process in vivo, potential tumorigenicity, and epigenetic abnormality of transplanted cells are preventing their clinical application. In this review, we give an overview on preclinical development of iPSC-derived myogenic cell transplantation therapies including processes related to iPSC-derived myogenic cells such as differentiation, scaling-up, delivery, and cGMP compliance. And we discuss the potential challenges of each step of clinical translation. Additionally, preclinical model systems for testing myogenic cells intended for clinical applications are described.

Effect of epicatechin consumption on the inflammatory pathway and mitochondria morphology in PBMC from a R350P desminopathy patient: A case report.

Desminopathy R350P is a human myopathy that is characterized by the progressive loss of muscle fiber organization. This results in the loss of muscle size, mobility, and strength. In desminopathy, inflammation affects muscle homeostasis and repair, and contributes to progressive muscle deterioration. Mitochondria morphology was also suggested to affect desminopathy progression. Epicatechin (Epi)-a natural compound found in cacao-has been proposed to regulate inflammatory signaling and mitochondria morphology in human and animal models. Hence, we hypothesize chronic Epi consumption to improve inflammatory pathway and mitochondria morphology in the peripheral blood mononuclear cells (PBMCs) of a desminopathy R350P patient. We found that 12 weeks of Epi consumption partially restored TRL4 signaling, indicative of inflammatory signaling and mitochondria morphology in the desminopathy patient. Moreover, Epi consumption improved blood health parameters, including reduced HOMA-IR and IL-6 levels in the desminopathy patient. This indicates that Epi consumption could be a useful tool to slow disease progression in desminopathy patients.

The Dysferlinopathies Conundrum: Clinical Spectra, Disease Mechanism and Genetic Approaches for Treatments.

Dysferlinopathies refer to a spectrum of muscular dystrophies that cause progressive muscle weakness and degeneration. They are caused by mutations in the DYSF gene, which encodes the dysferlin protein that is crucial for repairing muscle membranes. This review delves into the clinical spectra of dysferlinopathies, their molecular mechanisms, and the spectrum of emerging therapeutic strategies. We examine the phenotypic heterogeneity of dysferlinopathies, highlighting the incomplete understanding of genotype-phenotype correlations and discussing the implications of various DYSF mutations. In addition, we explore the potential of symptomatic, pharmacological, molecular, and genetic therapies in mitigating the disease's progression. We also consider the roles of diet and metabolism in managing dysferlinopathies, as well as the impact of clinical trials on treatment paradigms. Furthermore, we examine the utility of animal models in elucidating disease mechanisms. By culminating the complexities inherent in dysferlinopathies, this write up emphasizes the need for multidisciplinary approaches, precision medicine, and extensive collaboration in research and clinical trial design to advance our understanding and treatment of these challenging disorders.

Clinical Reasoning: A 19-Month-Old Girl With Infantile-Onset Myopathy and White Matter Changes.

We describe the case of a 19-month-old girl presenting with gross motor delays, hypotonia, diminished deep tendon reflexes, hyperCKaemia, extensive white matter changes on MRI brain, and electromyography studies consistent with myopathy. The differential diagnosis for infantile-onset hypotonia and muscle weakness is broad. It includes numerous subtypes of genetic disorders, including congenital muscular dystrophies, congenital myopathies, congenital myasthenic syndromes, spinal muscular atrophy, single-gene genetic syndromes, and inborn errors of metabolism. We outline our clinical approach leading to the diagnosis of a distinctive genetic neuromuscular condition essential for neurologists and geneticists working with patients of all ages to recognize.

Collagen VI Deficiency Impairs Tendon Fibroblasts Mechanoresponse in Ullrich Congenital Muscular Dystrophy.

The pericellular matrix (PCM) is a specialized extracellular matrix that surrounds cells. Interactions with the PCM enable the cells to sense and respond to mechanical signals, triggering a proper adaptive response. Collagen VI is a component of muscle and tendon PCM. Mutations in collagen VI genes cause a distinctive group of inherited skeletal muscle diseases, and Ullrich congenital muscular dystrophy (UCMD) is the most severe form. In addition to muscle weakness, UCMD patients show structural and functional changes of the tendon PCM. In this study, we investigated whether PCM alterations due to collagen VI mutations affect the response of tendon fibroblasts to mechanical stimulation. By taking advantage of human tendon cultures obtained from unaffected donors and from UCMD patients, we analyzed the morphological and functional properties of cellular mechanosensors. We found that the length of the primary cilia of UCMD cells was longer than that of controls. Unlike controls, in UCMD cells, both cilia prevalence and length were not recovered after mechanical stimulation. Accordingly, under the same experimental conditions, the activation of the Hedgehog signaling pathway, which is related to cilia activity, was impaired in UCMD cells. Finally, UCMD tendon cells exposed to mechanical stimuli showed altered focal adhesions, as well as impaired activation of Akt, ERK1/2, p38MAPK, and mechanoresponsive genes downstream of YAP. By exploring the response to mechanical stimulation, for the first time, our findings uncover novel unreported mechanistic aspects of the physiopathology of UCMD-derived tendon fibroblasts and point at a role for collagen VI in the modulation of mechanotransduction in tendons.

SNUPN deficiency causes a recessive muscular dystrophy due to RNA mis-splicing and ECM dysregulation.

SNURPORTIN-1, encoded by SNUPN, plays a central role in the nuclear import of spliceosomal small nuclear ribonucleoproteins. However, its physiological function remains unexplored. In this study, we investigate 18 children from 15 unrelated families who present with atypical muscular dystrophy and neurological defects. Nine hypomorphic SNUPN biallelic variants, predominantly clustered in the last coding exon, are ascertained to segregate with the disease. We demonstrate that mutant SPN1 failed to oligomerize leading to cytoplasmic aggregation in patients' primary fibroblasts and CRISPR/Cas9-mediated mutant cell lines. Additionally, mutant nuclei exhibit defective spliceosomal maturation and breakdown of Cajal bodies. Transcriptome analyses reveal splicing and mRNA expression dysregulation, particularly in sarcolemmal components, causing disruption of cytoskeletal organization in mutant cells and patient muscle tissues. Our findings establish SNUPN deficiency as the genetic etiology of a previously unrecognized subtype of muscular dystrophy and provide robust evidence of the role of SPN1 for muscle homeostasis.

The structure and function of lamin A/C: Special focus on cardiomyopathy and therapeutic interventions.

Lamins are inner nuclear membrane proteins that belong to the intermediate filament family. Lamin A/C lie adjacent to the heterochromatin structure in polymer form, providing skeletal to the nucleus. Based on the localization, lamin A/C provides nuclear stability and cytoskeleton to the nucleus and modulates chromatin organization and gene expression. Besides being the structural protein making the inner nuclear membrane in polymer form, lamin A/C functions as a signalling molecule involved in gene expression as an enhancer inside the nucleus. Lamin A/C regulates various cellular pathways like autophagy and energy balance in the cytoplasm. Its expression is highly variable in differentiated tissues, higher in hard tissues like bone and muscle cells, and lower in soft tissues like the liver and brain. In muscle cells, including the heart, lamin A/C must be expressed in a balanced state. Lamin A/C mutation is linked with various diseases, such as muscular dystrophy, lipodystrophy, and cardiomyopathies. It has been observed that a good number of mutations in the LMNA gene impact cardiac activity and its function. Although several works have been published, there are still several unexplored areas left regarding the lamin A/C function and structure in the cardiovascular system and its pathological state. In this review, we focus on the structural organization, expression pattern, and function of lamin A/C, its interacting partners, and the pathophysiology associated with mutations in the lamin A/C gene, with special emphasis on cardiovascular diseases. With the recent finding on lamin A/C, we have summarized the possible therapeutic interventions to treat cardiovascular symptoms and reverse the molecular changes.

ABHD7-mediated depalmitoylation of lamin A promotes myoblast differentiation.

LMNA gene mutation can cause muscular dystrophy, and post-translational modification plays a critical role in regulating its function. Here, we identify that lamin A is palmitoylated at cysteine 522, 588, and 591 residues, which are reversely catalyzed by palmitoyltransferase zinc finger DHHC-type palmitoyltransferase 5 (ZDHHC5) and depalmitoylase α/ß hydrolase domain 7 (ABHD7). Furthermore, the metabolite lactate promotes palmitoylation of lamin A by inhibiting the interaction between it and ABHD7. Interestingly, low-level palmitoylation of lamin A promotes, whereas high-level palmitoylation of lamin A inhibits, murine myoblast differentiation. Together, these observations suggest that ABHD7-mediated depalmitoylation of lamin A controls myoblast differentiation.

European Joint Programme on Rare Diseases workshop: LAMA2-muscular dystrophy: paving the road to therapy March 17-19, 2023, Barcelona, Spain.

The European Joint Programme on Rare Diseases (EJPRD) funded the workshop "LAMA2-Muscular Dystrophy: Paving the road to therapy", bringing together 40 health-care professionals, researchers, patient-advocacy groups, Early-Career Scientists and other stakeholders from 14 countries. Progress in natural history, pathophysiology, trial readiness, and treatment strategies was discussed together with efforts to increase patient-awareness and strengthen collaborations. Key outcomes were (a) ongoing natural history studies in 7 countries already covered more than 350 patients. The next steps are to include additional countries, harmonise data collection and define a minimal dataset; (b) therapy development was largely complementary. Approaches included LAMA2-replacement and correction, LAMA1-reactivation, mRNA modulation, linker-protein expression, targeting downstream processes and identifying modifiers, using viral vectors, muscle stem cells, iPSC and mouse models and patient lines; (c) LAMA2-Europe will inform patients (-representatives) worldwide on standards of care and scientific progress, and enable sharing experiences. Follow-up monthly online meetings and research repositories have been established to create sustainable collaborations.

Analysis of the GFP-labelled ß-dystroglycan interactome in HEK-293 transfected cells reveals novel intracellular networks.

Dystroglycan (DG) is a cell adhesion complex that is widely expressed in tissues. It is composed by two subunits, α-DG, a highly glycosylated protein that interacts with several extracellular matrix proteins, and transmembrane ß-DG whose, cytodomain binds to the actin cytoskeleton. Glycosylation of α-DG is crucial for functioning as a receptor for its multiple extracellular binding partners. Perturbation of α-DG glycosylation is the central event in the pathogenesis of severe pathologies such as muscular dystrophy and cancer. ß-DG acts as a scaffold for several cytoskeletal and nuclear proteins and very little is known about the fine regulation of some of these intracellular interactions and how they are perturbed in diseases. To start filling this gap by identifying uncharacterized intracellular networks preferentially associated with ß-DG, HEK-293 cells were transiently transfected with a plasmid carrying the ß-DG subunit with GFP fused at its C-terminus. With this strategy, we aimed at forcing ß-DG to occupy multiple intracellular locations instead of sitting tightly at its canonical plasma membrane milieu, where it is commonly found in association with α-DG. Immunoprecipitation by anti-GFP antibodies followed by shotgun proteomic analysis led to the identification of an interactome formed by 313 exclusive protein matches for ß-DG binding. A series of already known ß-DG interactors have been found, including ezrin and emerin, whilst significant new matches, which include potential novel ß-DG interactors and their related networks, were identified in diverse subcellular compartments, such as cytoskeleton, endoplasmic reticulum/Golgi, mitochondria, nuclear membrane and the nucleus itself. Of particular interest amongst the novel identified matches, Lamina-Associated Polypeptide-1B (LAP1B), an inner nuclear membrane protein, whose mutations are known to cause nuclear envelopathies characterized by muscular dystrophy, was found to interact with ß-DG in HEK-293 cells. This evidence was confirmed by immunoprecipitation, Western blotting and immunofluorescence experiments. We also found by immunofluorescence experiments that LAP1B looses its nuclear envelope localization in C2C12 DG-knock-out cells, suggesting that LAP1B requires ß-DG for a proper nuclear localization. These results expand the role of ß-DG as a nuclear scaffolding protein and provide novel evidence of a possible link between dystroglycanopathies and nuclear envelopathies displaying with muscular dystrophy.

A comparative study on riboflavin responsive multiple acyl-CoA dehydrogenation deficiency due to variants in FLAD1 and ETFDH gene.

Lipid storage myopathy (LSM) is a heterogeneous group of lipid metabolism disorders predominantly affecting skeletal muscle by triglyceride accumulation in muscle fibers. Riboflavin therapy has been shown to ameliorate symptoms in some LSM patients who are essentially concerned with multiple acyl-CoA dehydrogenation deficiency (MADD). It is proved that riboflavin responsive LSM caused by MADD is mainly due to ETFDH gene variant (ETFDH-RRMADD). We described here a case with riboflavin responsive LSM and MADD resulting from FLAD1 gene variants (c.1588 C > T p.Arg530Cys and c.1589 G > C p.Arg530Pro, FLAD1-RRMADD). And we compared our patient together with 9 FLAD1-RRMADD cases from literature to 106 ETFDH-RRMADD cases in our neuromuscular center on clinical history, laboratory investigations and pathological features. Furthermore, the transcriptomics study on FLAD1-RRMADD and ETFDH-RRMADD were carried out. On muscle pathology, both FLAD1-RRMADD and ETFDH-RRMADD were proved with lipid storage myopathy in which atypical ragged red fibers were more frequent in ETFDH-RRMADD, while fibers with faint COX staining were more common in FLAD1-RRMADD. Molecular study revealed that the expression of GDF15 gene in muscle and GDF15 protein in both serum and muscle was significantly increased in FLAD1-RRMADD and ETFDH-RRMADD groups. Our data revealed that FLAD1-RRMADD (p.Arg530) has similar clinical, biochemical, and fatty acid metabolism changes to ETFDH-RRMADD except for muscle pathological features.

Derivation of two human induced pluripotent stem cell lines carrying a missense mutation in FHL1 (c.377G > A, p.C126Y) linked to familial muscular dystrophy.

FHL1 gene locates in the Xq26 region and encodes for four and half LIM domain protein 1. It plays a crucial role in muscle cells and mutations in FHL1 are related to muscular dystrophy (MD). Peripheral blood mononuclear cells (PBMCs) were obtained from 2 family patients with MD that carry a pathogenic missense mutation in FHL1 (c.377G > A, p.C126Y). Induced pluripotent stem cells (iPSCs) were generated by PBMCs reprogramming using the lentiviral-hSTEMCCA-loxP vector, obtaining FHL1-T and FHL1-V iPSCs lines from patients. FHL1 genotype was maintained, and stemness and pluripotency were confirmed in both iPSCs lines.

Effect of sequential burr passes on minimally invasive akin and first metatarsal dorsiflexion osteotomies.

BACKGROUND: Minimally invasive surgical (MIS) osteotomies are increasing as a surgical option for treating midfoot and forefoot conditions. This study aimed to evaluate the impact of each burr pass on the degree of correction, gap size, and alignment in MIS Akin and first metatarsal dorsiflexion osteotomies (DFO). METHODS: MIS Akin and first metatarsal DFO were performed on ten cadaveric specimens. Fluoroscopic measurements included the metatarsal dorsiflexion angle (MDA), dorsal cortical length (MDCL), first phalangeal medial cortical length (PCML) and proximal to distal phalangeal articular angle (PDPAA). RESULTS: The average decrease in PCML with each burr pass was as follows: 1.53, 1.33, 1.27, 1.23 and 1.13 mm at the 1st to 5th pass, respectively. The MDCL sequentially decreased by 1.80, 1.59, 1.35, 0.75, and 0.60 mm. The MDA consistently decreased, and the PDPAA incrementally became more valgus oriented. CONCLUSION: On average, a first metatarsal dorsal wedge resection of 4.7 mm and first phalangeal medial wedge resection of 2.9 mm was achieved after 3 and 2 burr passes, respectively. This data may aid surgeons determine the optimal number of burr passes required to achieve the desired patient-specific surgical correction.

The super-healing MRL strain promotes muscle growth in muscular dystrophy through a regenerative extracellular matrix.

The Murphy Roths Large (MRL) mouse strain has "super-healing" properties that enhance recovery from injury. In mice, the DBA/2J strain intensifies many aspects of muscular dystrophy, so we evaluated the ability of the MRL strain to suppress muscular dystrophy in the Sgcg-null mouse model of limb girdle muscular dystrophy. A comparative analysis of Sgcg-null mice in the DBA/2J versus MRL strains showed greater myofiber regeneration, with reduced structural degradation of muscle in the MRL strain. Transcriptomic profiling of dystrophic muscle indicated strain-dependent expression of extracellular matrix (ECM) and TGF-ß signaling genes. To investigate the MRL ECM, cellular components were removed from dystrophic muscle sections to generate decellularized myoscaffolds. Decellularized myoscaffolds from dystrophic mice in the protective MRL strain had significantly less deposition of collagen and matrix-bound TGF-ß1 and TGF-ß3 throughout the matrix. Dystrophic myoscaffolds from the MRL background, but not the DBA/2J background, were enriched in myokines like IGF-1 and IL-6. C2C12 myoblasts seeded onto decellularized matrices from Sgcg-/- MRL and Sgcg-/- DBA/2J muscles showed the MRL background induced greater myoblast differentiation compared with dystrophic DBA/2J myoscaffolds. Thus, the MRL background imparts its effect through a highly regenerative ECM, which is active even in muscular dystrophy.

DAG1 haploinsufficiency is associated with sporadic and familial isolated or pauci-symptomatic hyperCKemia.

DAG1 encodes for dystroglycan, a key component of the dystrophin-glycoprotein complex (DGC) with a pivotal role in skeletal muscle function and maintenance. Biallelic loss-of-function DAG1 variants cause severe muscular dystrophy and muscle-eye-brain disease. A possible contribution of DAG1 deficiency to milder muscular phenotypes has been suggested. We investigated the genetic background of twelve subjects with persistent mild-to-severe hyperCKemia to dissect the role of DAG1 in this condition. Genetic testing was performed through exome sequencing (ES) or custom NGS panels including various genes involved in a spectrum of muscular disorders. Histopathological and Western blot analyses were performed on muscle biopsy samples obtained from three patients. We identified seven novel heterozygous truncating variants in DAG1 segregating with isolated or pauci-symptomatic hyperCKemia in all families. The variants were rare and predicted to lead to nonsense-mediated mRNA decay or the formation of a truncated transcript. In four cases, DAG1 variants were inherited from similarly affected parents. Histopathological analysis revealed a decreased expression of dystroglycan subunits and Western blot confirmed a significantly reduced expression of beta-dystroglycan in muscle samples. This study supports the pathogenic role of DAG1 haploinsufficiency in isolated or pauci-symptomatic hyperCKemia, with implications for clinical management and genetic counseling.

Plectin Deficiency in Fibroblasts Deranges Intermediate Filament and Organelle Morphology, Migration, and Adhesion.

Plectin, a highly versatile and multifunctional cytolinker, has been implicated in several multisystemic disorders. Most sequence variations in the human plectin gene (PLEC) cause epidermolysis bullosa simplex with muscular dystrophy (EBS-MD), an autosomal recessive skin-blistering disorder associated with progressive muscle weakness. In this study, we performed a comprehensive cell biological analysis of dermal fibroblasts from three different patients with EBS-MD, where PLEC expression analyses revealed preserved mRNA levels in all cases, whereas full-length plectin protein content was significantly reduced or completely absent. Downstream effects of pathogenic PLEC sequence alterations included massive bundling of vimentin intermediate filament networks, including the occurrence of ring-like nuclei-encasing filament bundles, elongated mitochondrial networks, and abnormal nuclear morphologies. We found that essential fibroblast functions such as wound healing, migration, or orientation upon cyclic stretch were significantly impaired in the cells of patients with EBS-MD. Finally, EBS-MD fibroblasts displayed reduced adhesion capacities, which could be attributed to smaller focal adhesion contacts. Our study not only emphasizes plectin's functional role in human skin fibroblasts, it also provides further insights into the understanding of EBS-MD-associated disease mechanisms.

LMNA-related muscular dystrophy involving myoblast proliferation and apoptosis through the FOXO1/GADD45A pathway.

LMNA-related muscular dystrophy is a major disease phenotype causing mortality and morbidity in laminopathies, but its pathogenesis is still unclear. To explore the molecular pathogenesis, a knock-in mouse harbouring the Lmna-W520R mutation was modelled. Morphological and motor functional analyses showed that homozygous mutant mice revealed severe muscular atrophy, profound motor dysfunction, and shortened lifespan, while heterozygotes showed a variant arrangement of muscle bundles and mildly reduced motor capacity. Mechanistically, the FOXO1/GADD45A pathway involving muscle atrophy processes was found to be altered in vitro and in vivo assays. The expression levels of FOXO1 and its downstream regulatory molecule GADD45A significantly increased in atrophic muscle tissue. The elevated expression of FOXO1 was associated with decreased H3K27me3 in its gene promotor region. Overexpression of GADD45A induced apoptosis and cell cycle arrest of myoblasts in vitro, and it could be partially restored by the FOXO1 inhibitor AS1842856, which also slowed the muscle atrophy process with improved motor function and prolonged survival time of homozygous mutant mice in vivo. Notably, the inhibitor also partly rescued the apoptosis and cell cycle arrest of hiPSC-derived myoblasts harbouring the LMNA-W520R mutation. Together, these data suggest that the activation of the FOXO1/GADD45A pathway contributes to the pathogenesis of LMNA-related muscle atrophy, and it might serve as a potential therapeutic target for laminopathies.

Riboflavin-responsive lipid-storage myopathy in elderly patients.

There are scarce reports of riboflavin-responsive lipid storage myopathy in elderly patients with onset in their sixties. We describe three elderly patients with riboflavin-responsive lipid-storage myopathy. All three patients (aged 67-71 years on first examination) had subacute onset of neck extensors and proximal limb weakness progressing to inability to rise from a sitting position or to walk. Muscle biopsies showed vacuoles with lipid content, mainly in type 1 fibers. Genetic analysis failed to identify any pathogenic variant in one patient, identified a heterozygous variant of uncertain significance c.812 A > G; p.Tyr271Cys in the ETFDH gene in the second patient, and revealed a heterozygote likely pathogenic variant c.1286-2 A > C in the ETFDH gene predicted to cause abnormal splicing in the third patient. All patients responded to treatment with riboflavin and carnitine, and regained normal strength. This report emphasizes the importance of muscle biopsy in revealing treatable lipid storage myopathy in elderly patients with progressive myopathy of unidentifiable cause.