Pathogenic biallelic variants in GYG1, encoding for glycogenin-1, are associated with polyglucosan bodies myopathy characterized by muscle accumulation of deposits of amylopectin-like polysaccharides (MIM 616199). So far, only few cases (<50) with molecular defects in GYG1 have been reported. The proband is a 79-year-old Italian woman presenting with subacute onset of diffuse soreness, weakness in the upper limbs and diffuse muscle atrophy without cardiac or respiratory involvement. Electromyography showed myopathic features. Muscle biopsy revealed several type I muscle fibers containing intensely PAS-positive, diastase-resistant vacuoles of variable dimension. Ultrastructural analysis showed vacuoles with granular-fibrillar storage material localized in subsarcolemmal and intermyofibrillar areas, small amounts of free glycogen and jagged Z-line appearance of some sarcomeres. Clinical exome sequencing revealed two heterozygous pathogenic variants in GYG1. Our findings provide clinical and molecular characterization of a novel case of GYG1-related polyglucosan bodies myopathy and highlight the histological clues leading to the diagnosis of this rare clinical phenotype.

Myosin heavy chain (MyHC)-related congenital myopathies display variable age of onset and clinical manifestations depending on the mutated isoform. Cardiomyopathy, ophthalmoplegia and primarily proximal weakness may be part of the clinical picture. A 57-year-old male patient with a history of arterial hypertension and hyperferritinemia (thalassemic trait) began to experience lower limb proximal weakness at the age of 23 years and he got progressively worse over the years when he also reported mild dyspnea, easy fatigability, fasciculations and a sort of discomfort in the posterior muscles of both thighs even at rest. He had no diplopia or dysphagia. Peculiar clinical features were bilateral exophthalmos, slight eyelid ptosis, limited ocular motility in all gaze directions, marked lower limb proximal weakness with posteromedial thigh hypotrophy and hypertrophic calves with increased consistency. Severe signs of both myopathic and neurogenic degeneration were seen at quadriceps skeletal muscle biopsy. Serum CK values were slightly elevated (less than 300 U/L). His parents are second-degree cousins and have normal serum CK levels, a proband’s first cousin had a diagnosis of muscular dystrophy and died at the age of 60 years, wheelchair bound. A 46-year-old sister is healthy. EMG showed signs of reinnervation in all muscles, myopathic signs being evident in the gastrocnemius muscles. Clear signs of fibro-adipose degeneration were observed at muscle MRI, more evidently so in the semitendinosus, rectus femoris, sartorius, gracilis and gastrocnemius muscles. Genetic analysis revealed the new homozygous variant c.3901G > T in exon 29 of MYH2 gene (NM_017534), predicted to introduce the premature stop codon p.(Glu1301*), thus leading to a diagnosis of MYH2-related myopathy. This variant shows a clear genotype-phenotype correlation, as it leads to a near-complete loss of MyHC IIa expression and results in a recessive myopathy characterized by adult-onset progressive muscle weakness with ophthalmoplegia and myopathic changes consistent with biallelic truncating mutations.

Core myopathies are congenital diseases with clinical, pathological and genetic heterogeneity. Main histological features are fiber “cores” showing a focally reduced oxidative enzyme activity. Dusty Core Disease (DuCD) differs from Central Core Myopathy for the presence of irregular areas, without clear borders and round/ovoidal shape, and myofibrillar disorganization characterized by reddish purple granular material depositions. This disorder is defined clinically by severe phenotypes with early onset of disease and molecularly by low level of RyR1 in muscle. Until now DuCD was associated only to biallelic recessive RYR1 mutations. We analyzed the clinical aspects, pathological features and mutational spectrum of four DuCD patients, belonging to our cohort of Congenital Myopathy probands. Molecular analysis detected 5 different RYR1 pathogenic variants, two of them so far unreported. Patients presented a heterogeneous phenotype ranging from severe recessive infantile forms to moderate dominant adult-onset presentations. Histological, immunological and ultrastructural techniques were employed to validate these dominant cases, which expand our knowledge on the inheritance of this subgroup of diseases.

Pompe disease is a rare, progressive lysosomal storage disorder caused by acid α-glucosidase deficiency, leading to glycogen accumulation, proximal muscle weakness, and respiratory decline. Enzyme replacement therapy (ERT) significantly improves survival and stabilizes motor function, but IgE-mediated hypersensitivity reactions (HSRs) can critically compromise treatment, posing a major clinical challenge. Desensitization protocols allow temporary tolerance to ERT, yet breakthrough reactions may occur, necessitating adjunctive strategies such as omalizumab. We report a 40-year-old woman with late-onset Pompe disease who developed severe IgE-mediated HSRs to alglucosidase alfa after years of uneventful therapy. Basophil activation testing (BAT) and serum-specific IgE confirmed an IgE-mediated mechanism. A 15-step, 5-bag desensitization protocol allowed temporary tolerance, but breakthrough reactions required therapy interruption. Upon switching to avalglucosidase alfa, BAT demonstrated IgE cross-reactivity, and a new desensitization protocol was implemented. Initial infusions were complicated by recurrent HSRs. The addition of subcutaneous omalizumab (300 mg monthly), administered two days before ERT, enabled safe reintroduction. Moreover, therapy was resumed gradually, starting at 50

Objectives Ataxia with vitamin E deficiency (AVED) is a chronic progressive syndrome with low vitamin E levels, caused by biallelic pathogenic variants in the alpha-tocopherol transfer protein (TTPA) gene on chromosome 8q13.1. According to family segregation studies, we describe a novel putative variant in compound heterozygosity with a known pathogenic variant. Methods Clinical and instrumental evaluations were performed at our Neuromuscular Unit. Results Two siblings had been experiencing clumsiness, which later evolved into progressive gait unsteadiness, with falls and dysarthria. Brain MRI, EMG, and nerve conduction studies were inconclusive, but focused clinical exome sequencing revealed biallelic variants in the TTPA gene in both, the pathogenetic maternally inherited variant c.513_514insTT (p.Thr172Leufs*5) on exon 3 and the previously undescribed paternally inherited variant c.158T > C (p.Leu53Pro) on exon 1. Their plasma vitamin E levels were low (<1.7 mg/L). Their relatives carried 1 single variant each, with normal vitamin E levels. After 6 months of oral vitamin supplementation with a laboratory response, disease progression has stopped, although their neurologic examination has not clearly ameliorated. Discussion Exome sequencing identified a new potentially pathogenic variant in AVED, an ultra-rare but treatable cause of inherited ataxia. We recommend prompt vitamin E supplementation in patients with AVED.

ObjectiveThe recent examination of a woman who presented a migraine attack with visual aura and unilateral mydriasis, followed, a few days later, by an episode of Takotsubo Syndrome, prompted us to review the existing literature correlating migraine and TS as well as the most recent studies regarding the association between Takotsubo Syndrome and neurological dysfunction. Our aim is to define the role of dysautonomia and the contribution of the Autonomic Nervous System in the pathogenesis of both diseases, also, to define the functional role of the hypothalamus in the both the determination of headache attacks and the onset of clinical manifestations in other organs and tissues.BackgroundMigraine with aura and Takotsubo Syndrome (TS) share emotional, psychological and physical triggering factors which are implicated in the pathogenetic process. The autonomic nervous system plays a crucial, though antipodal, role in both pathologies. Indeed, migraine clinical manifestations imply the activation of the parasympathetic system whereas a massive catecholamine release following sympathetic nervous system activation is pivotal in triggering the Takotsubo Syndrome.MethodsReview of anatomy, physiology and neural regulation in both physiological and stress responses. Examination of the functional connections of the hypothalamus.Results and discussionThe coexistence of psycho-emotional stressors, endocrine/hormonal factors and a possible genetic predisposition might explain the pathophysiological overlap between these two distinct clinical manifestations. The hypothalamus is functionally connected with all the pain-related brain regions and pathways and can be responsible for the clinical manifestations in both the nervous system and the cardiovascular apparatus. Migraine is the most frequent chronic, early onset condition associated with TS.

Inflammatory myopathies represent a heterogeneous group of autoimmune disorders affecting skeletal muscle, with distinct pathological features. While muscle biopsy remains a key diagnostic tool, conventional immunohistochemical approaches are limited in both number of available markers and spatial resolution. To explore the feasibility and potential of the MACSima™ Imaging Platform for high-dimensional, spatially resolved analysis of skeletal muscle tissue in inflammatory myopathies, we applied a panel of antibodies targeting immune cells, extracellular matrix components, blood vessels, and lymphatic markers to skeletal muscle cryosections from patients with different inflammatory myopathies and age-matched controls. Using the MACSima™ platform, we performed iterative immunofluorescence staining and imaging cycles to generate multiplexed protein maps. The platform enabled robust signal acquisition and preserved tissue morphology throughout all cycles. Preliminary spatial analysis revealed distinct immune cell infiltration patterns, ECM remodeling signatures, and vascular changes across disease subtypes. This pilot study demonstrates the feasibility of applying high-dimensional multiplex imaging to skeletal muscle using the MACSima™ system. Our results represent a first partial spatial profiling of muscle pathology in autoimmune myopathies using an innovative methodological approach.

The Idiopathic Inflammatory Myopathies (IIMs) are a group of autoimmune disorders characterised by persistent muscle inflammation and diverse clinical manifestations. Common symptoms include muscle weakness, myalgia, and elevated serum creatine kinase levels. Recent findings highlight the relevance of muscle fibre necrosis in IIMs. We therefore grouped our IIM patients according to the percentage of necrotic fibres in muscle biopsy. Our clustered patients were analysed for the inflammatory milieu, the capillary network, and the endoplasmic reticulum stress. In patients with a higher percentage of necrotic fibres we detected a more marked presence of CD206 positive cells, an activation of the endothelial-mesenchymal transition process, an altered capillary network, more marked ER stress and connective tissue deposition. Furthermore, our study revealed a key role of C/EBP-homologous protein (CHOP), a multifunctional transcription factor that contributes to cellular functions including apoptosis, autophagy, inflammation, mediation of ER stress and induction of fibrosis. Our study suggests that CD206 positive cells and CHOP have an important role in pathogenetic mechanisms and could therefore be considered possible therapeutic targets to modulate the inflammatory response of these patients, namely to contain or slow down the progression of fibrosis.

Glycogen storage disease II or Pompe disease (PD), is a rare autosomal recessive disorder due to biallelic pathogenic variants in GAA, resulting in the enzymatic deficiency of alpha-1,4-glucosidase. Two clinical forms are recognized, namely, early onset (EOPD) and late-onset (LOPD). We present the case of an asymptomatic 33-year-old man who underwent a genetic screening for autosomal recessive disorders (parental prenatal counselling) and was found to carry the homozygous pathogenic GAA substitution NM_000152.5(GAA):c.-32-13T>G (IVS1). Neurological examination, serum CK levels, electromyography, muscle MRI, respiratory and cardiac screening were reported normal. We investigated the effects of the variant at transcript and protein levels in available tissues from the proband and his parents. The IVS1-32-13T>G variant (dbSNP: rs386834236, Clin Var ID: 4,027) occurs in 90% of Caucasian LOPD patients and is associated with a broad range of symptom onset. About 50 subjects have been reported harboring this variant in homozygosis and most of them are asymptomatic, although a subset develops symptoms with time. Residual levels of alpha-1,4-glucosidase activity and protein content do not seem to reflect clinical severity in homozygous IVS1 LOPD patients.

Skeletal muscle dystrophies comprise a group of inherited disorders characterized by progressive muscle weakness, with Duchenne and Becker muscular dystrophies (DMD/BMD) being among the most severe. These dystrophies are caused by mutations in the dystrophin gene, resulting in muscle cell instability, chronic inflammation, fibrosis, and impaired muscle regeneration. Although skeletal muscle has intrinsic regenerative potential via satellite cells, the ongoing muscle damage in DMD/BMD depletes these cells and promotes fibrosis. Inflammation also plays a pivotal role, with immune cell infiltration correlating with disease severity. This study investigates fibrosis, inflammation, and capillarization in BMD patients across different age groups to clarify how disease progression varies over time. Morphological analyses of muscle biopsies revealed an increase in connective tissue, particularly in adult patients. Pediatric patients showed reduced capillarization, whereas adult patients displayed vascular adaptations, including elevated capillary-to-fibre ratios and capillary contacts, indicative of compensatory mechanisms in response to chronic muscle degeneration. Inflammatory profiles also varied with age: younger adult patients exhibited a predominance of CD68-positive macrophages, while older adults demonstrated increased CD4/CD8 T-cell activity. Our findings highlight pronounced age-dependent differences in muscle pathology, encompassing structural adaptations, fibrosis, and inflammation, which may be crucial for developing age-tailored therapeutic approaches.

Magnetization transfer imaging (MTI) evaluates the exchange of magnetization between protons in free water molecules and protons bound to macromolecules, including lipids. Widely used in the study of central nervous system diseases, its application in the neuromuscular field has been previously explored only in a Spanish cohort of patients with late-onset Pompe disease (LOPD). To investigate the potential role of MTR as an early biomarker of muscle involvement, we here evaluate magnetization transfer ratio (MTR) and fat fraction (FF) in patients with LOPD in various stages of disease compared to healthy controls (HCs). Quantitative muscle MRI (qMRI) was performed on 31 LOPD patients (21 with mild and 10 with moderate/severe clinical involvement) and 31 matched HCs using 3T MRI. FF and MTR were measured in 11 thigh muscles. Correlations between FF and MTR were assessed. Additionally, FF and MTR were compared between groups of HCs vs. early vs. moderate/severe LOPD. We also explored whether MTR can detect muscle involvement in not yet fat-infiltrated muscles (FF ≤ 10%) in early LOPD. MTR of thigh muscles with FF ≤ 10% was significantly lower in LOPD compared to HCs. Changes in MTR could be detected even in mildly symptomatic patients, particularly in the medial and posterior compartments (Mann-Whitney U: p < 0.05). MTR and FF were inversely correlated in all subjects groups. We found significant differences in MTR and FF changes at the group level between mild and moderate/severe vs HCs. We conclude that MTI has potentially high sensitivity to detect mild muscle fiber damage, even before fat replacement has occurred, making it a useful biomarker to monitor early signs of disease, disease progression, and the efficacy of treatment approaches (Sanofi company provided support for this study, but had no role in study design, data collection and analysis, decision to publish, or preparation of the abstract). (The first and second authors contributed equally to this work)

Inclusion body myositis (IBM) is a slowly progressive disorder belonging to the idiopathic inflammatory myopathies, and it represents the most common adult-onset acquired myopathy. The main clinical features include proximal or distal muscular asymmetric weakness, with major involvement of long finger flexors and knee extensors. The main histological findings are the presence of fiber infiltrations, rimmed vacuoles, and amyloid inclusions. The etiopathogenesis is a challenge because both environmental and genetic factors are implicated in muscle degeneration and a distinction has been made previously between sporadic and hereditary forms. Here, we describe an Italian patient affected with a hereditary form of IBM with onset in his mid-forties. Next-generation sequencing analysis disclosed a heterozygous mutation c.76C>T (p.Pro26Ser) in the PDZ motif of the LDB3/ZASP gene, a mutation already described in a family with a late-onset myopathy and highly heterogenous degree of skeletal muscle weakness. In the proband’s muscle biopsy, the expression of ZASP, myotilin, and desmin were increased. In our family, in addition to the earlier age of onset, the clinical picture is even more peculiar given the evidence, in one of the affected family members, of complete ophthalmoplegia in the vertical gaze. These findings help extend our knowledge of the clinical and genetic background associated with inclusion body myopathic disorders.

Objective:Myotonic dystrophy type 2 (DM2; PROMM) is characterized by myotonia and muscle dysfunction, episodic muscle pain, proximal and axial weakness of the neck flexors. We describe the case of a young woman affected with a clinically silent form of DM2 disclosed by her return to physical exercise, a 7 km walk, after Covid-19 lockdown. Methods:The patient underwent neurological examination, serum CK dosage and electromyography after assessing the Emergency Room complaining of cramps and severe myalgia. Molecular screening for CNBP expansions was carried out on the patient and her family. Results:Clinical signs were generalized muscle weakness, more evident in the lower limb-girdle, myotonia at hands and foot fingers and dramatic elevation in CK levels. DM2 genetic assay revealed a pathological expansion in intron 1 of CNBP gene, confirming the clinical suspicion. Conclusions:The case we describe is the first, to our knowledge, addressing the impact of Covid pandemia on DM2 patients. In particular, we discuss the role of physical training in modulating the onset and the severity of clinical manifestations of DM2, since sustained regular exercise can mask the disease whereas prolonged suspension can cause massive muscle damage. Recent works investigate possible molecular mechanisms altered by forced physical inactivity, preventing skeletal muscle from adapting to the sudden, non-progressive training reactivation. Additional observations on DM2 patients, other myopathic subjects and elders will help clarify this important issue and provide useful behavioural advice.

ObjectiveMitochondrial DNA (mtDNA) depletion/deletions syndrome (MDDS) comprises a group of diseases caused by primary autosomal defects of mtDNA maintenance. Our objective was to study the etiology of MDDS in 4 patients who lack pathogenic variants in known genetic causes.MethodsWhole exome sequencing of the probands was performed to identify pathogenic variants. We validated the mitochondrial defect by analyzing mtDNA, mitochondrial dNTP pools, respiratory chain activities, and GUK1 activity. To confirm pathogenicity of GUK1 deficiency, we expressed 2 GUK1 isoforms in patient cells.ResultsWe identified biallelic GUK1 pathogenic variants in all 4 probands who presented with ptosis, ophthalmoparesis, and myopathic proximal limb weakness, as well as variable hepatopathy and altered T‐lymphocyte profiles. Muscle biopsies from all probands showed mtDNA depletion, deletions, or both, as well as reduced activities of mitochondrial respiratory chain enzymes. GUK1 encodes guanylate kinase, originally identified as a cytosolic enzyme. Long and short isoforms of GUK1 exist. We observed that the long isoform is intramitochondrial and the short is cytosolic. In probands’ fibroblasts, we noted decreased GUK1 activity causing unbalanced mitochondrial dNTP pools and mtDNA depletion in both replicating and quiescent fibroblasts indicating that GUK1 deficiency impairs de novo and salvage nucleotide pathways. Proband fibroblasts treated with deoxyguanosine and/or forodesine, a purine phosphatase inhibitor, ameliorated mtDNA depletion, indicating potential pharmacological therapies.InterpretationPrimary GUK1 deficiency is a new and potentially treatable cause of MDDS. The cytosolic isoform of GUK1 may contribute to the T‐lymphocyte abnormality, which has not been observed in other MDDS disorders. ANN NEUROL 2024

A 53-year-old man approached our Neuromuscular Unit following an incidental finding of hyperckemia. Similar to his mother who had died at the age of 77 years, he was diabetic and had a few lipomas. The patient’s two sisters, aged 60 and 50 years, did not have any neurological symptoms. Proband’s skeletal muscle biopsy showed several COX-negative fibers, many of which were “ragged red”. Genetic analysis revealed the presence of the A8344G mtDNA mutation, which is most commonly associated with a maternally inherited multisystem mitochondrial disorder known as MERRF (myoclonus epilepsy with ragged-red fibers). The two sisters also carry the mutation. Family members on the maternal side were reported healthy. Although atypical phenotypes have been reported in association with the A8344G mutation, central nervous system (CSN) manifestations other than myoclonic epilepsy are always reported in the family tree. If present, our four-generation family manifestations are late-onset and do not affect CNS. This could be explained by the fact that the mutational load remains low and therefore prevents tissues/organs from reaching the pathologic threshold. The fact that this occurs throughout generations and that CNS, which has the highest energetic demand, is clinically spared, suggests that regulatory genes and/or pathways affect mitochondrial segregation and replication, and protect organs from progressive dysfunction.

Collagen VI is a heterotrimeric protein expressed in several tissues and involved in the maintenance of cell integrity. It localizes at the cell surface, creating a microfilamentous network that links the cytoskeleton to the extracellular matrix. The heterotrimer consists of three chains encoded by COL6A1, COL6A2 and COL6A3 genes. Recessive and dominant molecular defects cause two main disorders, the severe Ullrich congenital muscular dystrophy and the relatively mild and slowly progressive Bethlem myopathy. We analyzed the clinical aspects, pathological features and mutational spectrum of 15 COL6-mutated patients belonging to our cohort of muscular dystrophy probands. Patients presented a heterogeneous phenotype ranging from severe forms to mild adult-onset presentations. Molecular analysis by NGS detected 14 different pathogenic variants, three of them so far unreported. Two changes, localized in the triple-helical domain of COL6A1, were associated with a more severe phenotype. Histological, immunological and ultrastructural techniques were employed for the validation of the genetic variants; they documented the high variability in COL6 distribution and the extracellular matrix disorganization, highlighting the clinical heterogeneity of our cohort. The combined use of these different technologies is pivotal in the diagnosis of COL6 patients.

Leigh syndrome (LS) or subacute necrotizing encephalomyelopathy is a progressive, lethal, mitochondrial disease mostly presenting in childhood. We report a 36-year-old African male presenting to the Emergency Department with a 6-month history of progressive dysarthria. Examination also showed oculomotor limitations for downgaze and convergence, mild right hemiparesis with Babinski sign, and absent lower limb tendon reflexes. He revealed he was presenting mild walking difficulties since the age of 25. Family history was unremarkable. A brain magnetic resonance showed diffuse white matter lesions without enhancing around the third ventricle, in the medulla oblongata, and bilaterally to the caudate and the putamen. Polymerase-chain-reaction amplification of the mitochondrial genes, followed by direct sequencing, found a 10191T>C variant related to LS. To now, only 8 late-onset patients share the same pathogenic variant.