Phenotypic Variability of LGMD 2C/R5 in a Genetically Homogenous Group of Bulgarian Muslim Roma.

Sarcoglycanopathies are among the most frequent and severe forms of autosomal recessive forms of limb-girdle muscular dystrophies (LGMDs) with childhood onset. Four subtypes are known: LGMDR3, LGMDR4, LGMDR5 and LGMDR6, which are caused, respectively, by mutations in the SGCA, SGCB, SGCG and SGCD genes. We present the clinical variability of LGMD 2C/R5 among a genetically homogeneous group of 57 patients, belonging to 35 pedigrees. Molecular genetic analysis showed that all 57 patients were homozygous for the C283Y variant. The muscles of the pelvic girdle and the trunk were affected early and were more severely affected, followed by the shoulder girdle. Macroglossia, hypertrophy of the calves, scapular winging and lumbar hyperlordosis were common in the ambulatory phase. A great intra and interfamilial variability in the clinical presentation of LGMD 2C/R5 was observed, despite having the same underlying molecular defect. Females demonstrated a relatively milder clinical course compared to males. Mean creatine phosphokinase (CK) CK levels were 20 times above normal values. Muscle computer tomography (CT) CT or MRIs showed earlier and more severe involvement of the flexor proximal limb muscles in comparison to extensor muscles.

Clinical and molecular characterization of limb-girdle muscular dystrophy 2G/R7 in a large cohort of Brazilian patients.

Limb-girdle muscular dystrophy type 2G/R7 (LGMD2G/R7) is an ultra-rare condition initially identified within the Brazilian population. We aimed to expand clinical and genetic information about this disease, including its worldwide distribution. A multicenter historical cohort study was performed at 13 centers in Brazil in which data from index cases and their affected relatives from consecutive families with LGMD2G/R7 were reviewed from July 2017 to August 2023. Additionally, a systematic literature review was conducted to identify case reports and series of the disease worldwide. Forty-one LGMD2G/R7 cases were described in the Brazilian cohort, being all subjects homozygous for the c.157C>T/(p.Gln53*) variant in TCAP. Survival curves showed that the median disease duration before individuals required walking aids was 21 years. Notably, women exhibited a slower disease progression, requiring walking aids 13 years later than men. LGMD2G/R7 was frequently reported not only in Brazil but also in China and Bulgaria, with 119 cases identified globally, with possible founder effects in the Brazilian, Eastern European, and Asian populations. These findings are pivotal in raising awareness of LGMD2G/R7, understanding its progression, and identifying potential modifiers. This can significantly contribute to the development of future natural history studies and clinical trials for this disease.

Clinical and Genetic Heterogeneity of Nuclear Envelopathy Related Muscular Dystrophies in an Indian Cohort.

Introduction: Nuclear envelopathies occur due to structural and/or functional defects in various nuclear envelope proteins such as lamin A/C and lamin related proteins. This study is the first report on the phenotype-genotype patterns of nuclear envelopathy-related muscular dystrophies from India. Methods: In this retrospective study, we have described patients with genetically confirmed muscular dystrophy associated with nuclear envelopathy. Data on clinical, laboratory findings and muscle MRI were collected. Results: Sixteen patients were included with median age at onset of 3 years (range: 1 month - 17 years). Three genes were involved: LMNA (11, 68.75%), EMD (4, 25%) and SYNE1 (1, 6.25%). The 11 patients with LMNA variants were Congenital muscular dystrophy (MDCL)=4, Limb Girdle Muscular Dystrophy (LGMD1B)=4 and Emery-Dreifuss Muscular Dystrophy (EDMD2)=3. On muscle biopsy, one patient from each laminopathy phenotype (n = 3) revealed focal perivascular inflammatory infiltrate. Other notable features were ophthalmoparesis in one and facial weakness in one. None had cardiac involvement. Patients with EDMD1 had both upper (UL) and lower limb (LL) proximo-distal weakness. Cardiac rhythm disturbances such as sick sinus syndrome and atrial arrhythmias were noted in two patients with EDMD1. Only one patient with variant c.654_658dup (EMD) lost ambulation in the 3rd decade, 18 years after disease onset. Two had finger contractures with EMD and SYNE1 variants respectively. All patients with LMNA and SYNE1 variants were ambulant at the time of evaluation. Mean duration of illness (years) was 11.6±13 (MDCL), 3.2±1.0 (EDMD2), 10.4±12.8 (LGMD1B), 11.8±8.4 (EDMD1) and 3 (EDMD4). One patient had a novel SYNE1 mutation (c.22472dupA, exon 123) and presented with UL phenotype and prominent finger and wrist contractures. Conclusion: The salient features included ophthalmoparesis and facial weakness in LMNA, prominent finger contractures in EMD and SYNE1 and upper limb phenotype with the novel pathogenic variant in SYNE1.

Exploring Splice-Site Mutations in LAMA2-Related Muscular Dystrophies: A Comprehensive Analysis of Genotypic and Phenotypic Patterns.

LAMA2-related muscular dystrophies (LAMA2-RDs) constitute the most prevalent subtype of congenital muscular dystrophies (CMDs). The clinical spectrum of LAMA2-RDs exhibits considerable diversity, particularly in motor development and disease progression. Phenotypic variability ranges from severe, early-onset presentation, known as merosin-deficient CMD type 1A, to milder, late-onset presentations, including limb-girdle muscular dystrophy-like phenotype. In this study, whole exome sequencing (WES) was applied to a family with a single proband affected by severe muscular dystrophy. The identified causative mutation was a biallelic splice-site mutation in intron 58 of the LAMA2 gene, leading to a premature termination codon in the critical G domain of laminin-α2 and resulting in a severe phenotype. Additionally, we summarized previously reported splice-site mutations to investigate the clinical and transcription consequences of these mutations. Our findings conclude that splice-site mutations predominantly lead to severe MDC1A, whether in a homozygous or heterozygous state, often associated with another loss-of-function mutation. Besides, splice-site mutations with available analysis of their transcriptional consequences were found to be responsible for exon skipping in most cases and the loss of the reading frame. These findings revealed the importance of WES in identifying disease-causing mutations, particularly in highly diversified pathologies like LAMA2-RDs. The results also underscore the importance of transcriptional analysis in determining the impact of splice-site mutations and the phenotype of LAMA2-RDs on patients.

Limb Girdle Muscular Dystrophy Type 2B (LGMD2B): Diagnosis and Therapeutic Possibilities.

Dysferlin is a large transmembrane protein involved in critical cellular processes including membrane repair and vesicle fusion. Mutations in the dysferlin gene (DYSF) can result in rare forms of muscular dystrophy; Miyoshi myopathy; limb girdle muscular dystrophy type 2B (LGMD2B); and distal myopathy. These conditions are collectively known as dysferlinopathies and are caused by more than 600 mutations that have been identified across the DYSF gene to date. In this review, we discuss the key molecular and clinical features of LGMD2B, the causative gene DYSF, and the associated dysferlin protein structure. We also provide an update on current approaches to LGMD2B diagnosis and advances in drug development, including splice switching antisense oligonucleotides. We give a brief update on clinical trials involving adeno-associated viral gene therapy and the current progress on CRISPR/Cas9 mediated therapy for LGMD2B, and then conclude by discussing the prospects of antisense oligomer-based intervention to treat selected mutations causing dysferlinopathies.

Genetic spectrum of sarcoglycanopathies in a cohort of Russian patients.

Sarcoglycanopathies encompass four distinct forms of limb-girdle muscular dystrophies (LGMD), denoted as LGMD R3-R6, arising from mutations within the SGCA, SGCB, SGCG, and SGCD genes. The global prevalence of sarcoglycanopathies is low, making it challenging to study these diseases. The principal objective of this study was to explore the spectrum of mutations in a cohort of Russian patients with sarcoglycanopathies and to ascertain the frequency of these conditions in the Russian Federation. We conducted a retrospective analysis of clinical and molecular genetic data from 49 Russian patients with sarcoglycan genes variants. The results indicated that variants in the SGCA gene were found in 71.4% of cases, with SGCB and SGCG genes each exhibiting variants in 12.2 % of patients. SGCD gene variants were detected in 4.1% of cases. Bi-allelic pathogenic and likely pathogenic variants were identified in 46 of the 49 cases of sarcoglycanopathies: LGMD R3 (n = 34), LGMD R4 (n = 4), LGMD R5 (n = 6), and LGMD R6 (n = 2). A total of 31 distinct variants were identified, comprising 25 previously reported and 6 novel variants. Two major variants, c.229C>T and c.271G>A, were detected within the SGCA, constituting 61.4% of all mutant alleles in Russian patients with LGMD R3. Both LGMD R6 cases were caused by the homozygous nonsense variant c.493C>T p.(Arg165Ter) in the SGCD gene. The incidence of sarcoglycanopathies in the Russian Federation was estimated to be at least 1 in 4,115,039, which is lower than the reported incidence in other populations.

Is Cardiac Transplantation Still a Contraindication in Patients with Muscular Dystrophy-Related End-Stage Dilated Cardiomyopathy? A Systematic Review.

Inherited muscular diseases (MDs) are genetic degenerative disorders typically caused by mutations in a single gene that affect striated muscle and result in progressive weakness and wasting in affected individuals. Cardiac muscle can also be involved with some variability that depends on the genetic basis of the MD (Muscular Dystrophy) phenotype. Heart involvement can manifest with two main clinical pictures: left ventricular systolic dysfunction with evolution towards dilated cardiomyopathy and refractory heart failure, or the presence of conduction system defects and serious life-threatening ventricular arrhythmias. The two pictures can coexist. In these cases, heart transplantation (HTx) is considered the most appropriate option in patients who are not responders to the optimized standard therapeutic protocols. However, cardiac transplant is still considered a relative contraindication in patients with inherited muscle disorders and end-stage cardiomyopathies. High operative risk related to muscle impairment and potential graft involvement secondary to the underlying myopathy have been the two main reasons implicated in the generalized reluctance to consider cardiac transplant as a viable option. We report an overview of cardiac involvement in MDs and its possible association with the underlying molecular defect, as well as a systematic review of HTx outcomes in patients with MD-related end-stage dilated cardiomyopathy, published so far in the literature.

A retrospective study on the clinical and molecular outcomes of calpainopathy in a Turkish patient cohort.

Background and aim: Calpainopathy, also known as limb-girdle muscular dystrophy recessive type 1, is a progressive muscle disorder that impacts the muscles around the hips and shoulders. The disease is caused by defects in the CAPN3 gene and can be inherited in both recessive and dominant forms. In this retrospective study, we aimed to evaluate the clinical and molecular results of our patients with calpainopathy and to examine the CAPN3 variants in Turkish and global populations. Materials and methods: Molecular analyses were performed using the next-generation sequencing (NGS) method. CAPN3 variants were identified through the examination of various databases. Results: In this retrospective study, the cohort consisted of seven patients exhibiting the CAPN3 (NM_000070.3) mutation and a phenotype compatible with calpainopathy at a single center in Türkiye. All patients displayed high CK levels and muscle weakness. We report a novel missense c.2437G>A variant that causes the autosomal dominant form of calpainopathy. Interestingly, the muscle biopsy report for the patient with the novel mutation indicated sarcoglycan deficiency. Molecular findings for the remaining individuals in the cohort included a compound heterozygous variant (frameshift and missense), one homozygous nonsense, one homozygous intronic deletion, and three homozygous missense variants. The most common variant in the Turkish population was c.550del. In both populations, pathogenic variants were most frequently located in exon 21, according to exon length. Variants were stochastically distributed based on consequences in CAPN3 domains. Conclusion: Therefore, the NGS method proves highly effective in diagnosing rare diseases characterized by clinical heterogeneity. Assessing variants based on ethnicity holds significance in the development of precise therapies.

Expert Panel Curation of 31 Genes in Relation to Limb Girdle Muscular Dystrophy.

Introduction: Limb girdle muscular dystrophies (LGMDs) are a group of genetically heterogeneous autosomal conditions with some degree of phenotypic homogeneity. LGMD is defined as having onset >2 years of age with progressive proximal weakness, elevated serum creatine kinase levels and dystrophic features on muscle biopsy. Advances in massively parallel sequencing have led to a surge in genes linked to LGMD. Methods: The ClinGen Muscular Dystrophies and Myopathies gene curation expert panel (MDM GCEP, formerly Limb Girdle Muscular Dystrophy GCEP) convened to evaluate the strength of evidence supporting gene-disease relationships (GDR) using the ClinGen gene-disease clinical validity framework to evaluate 31 genes implicated in LGMD. Results: The GDR was exclusively LGMD for 17 genes, whereas an additional 14 genes were related to a broader phenotype encompassing congenital weakness. Four genes (CAPN3, COL6A1, COL6A2, COL6A3) were split into two separate disease entities, based on each displaying both dominant and recessive inheritance patterns, resulting in curation of 35 GDRs. Of these, 30 (86%) were classified as Definitive, 4 (11%) as Moderate and 1 (3%) as Limited. Two genes, POMGNT1 and DAG1, though definitively related to myopathy, currently have insufficient evidence to support a relationship specifically with LGMD. Conclusions: The expert-reviewed assertions on the clinical validity of genes implicated in LGMDs form an invaluable resource for clinicians and molecular geneticists. We encourage the global neuromuscular community to publish case-level data that help clarify disputed or novel LGMD associations.

TRAPPC11-CDG muscular dystrophy: Review of 54 cases including a novel patient.

The trafficking protein particle (TRAPP) complex is a multisubunit protein complex that functions as a tethering factor involved in intracellular trafficking. TRAPPC11, a crucial subunit of this complex, is associated with pathogenic variants that cause a spectrum of disease, which can range from a limb girdle muscular dystrophy (LGMD) to developmental disability with muscle disease, movement disorder and global developmental delay (GDD)/intellectual disability (ID), or even a congenital muscular dystrophy (CMD). We reviewed the phenotype of all reported individuals with TRAPPC11-opathies, including an additional Mexican patient with novel compound heterozygous missense variants in TRAPPC11 (c.751 T > C and c.1058C > G), restricted to the Latino population. In these 54 patients muscular dystrophy signs are common (early onset muscle weakness, increased serum creatine kinase levels, and dystrophic changes in muscle biopsy). They present two main phenotypes, one with a slowly progressive LGMD with or without GDD/ID (n = 12), and another with systemic involvement characterized by short stature, GDD/ID, microcephaly, hypotonia, poor speech, seizures, cerebral atrophy, cerebellar abnormalities, movement disorder, scoliosis, liver disease, and cataracts (n = 42). In 6 of them CMD was identified. Obstructive hydrocephaly, retrocerebellar cyst, and talipes equinovarus found in the individual reported here has not been described in TRAPPC11 deficiency. As in previous patients, membrane trafficking assays in our patient showed defective abnormal endoplasmic reticulum-Golgi transport as well as decreased expression of LAMP2, and ICAM-1 glycoproteins. This supports previous statements that TRAPPC11-opathies are in fact a congenital disorder of glycosylation (CDG) with muscular dystrophy.

A Bio-Inspired Probabilistic Neural Network Model for Noise-Resistant Collision Perception.

Bio-inspired models based on the lobula giant movement detector (LGMD) in the locust's visual brain have received extensive attention and application for collision perception in various scenarios. These models offer advantages such as low power consumption and high computational efficiency in visual processing. However, current LGMD-based computational models, typically organized as four-layered neural networks, often encounter challenges related to noisy signals, particularly in complex dynamic environments. Biological studies have unveiled the intrinsic stochastic nature of synaptic transmission, which can aid neural computation in mitigating noise. In alignment with these biological findings, this paper introduces a probabilistic LGMD (Prob-LGMD) model that incorporates a probability into the synaptic connections between multiple layers, thereby capturing the uncertainty in signal transmission, interaction, and integration among neurons. Comparative testing of the proposed Prob-LGMD model and two conventional LGMD models was conducted using a range of visual stimuli, including indoor structured scenes and complex outdoor scenes, all subject to artificial noise. Additionally, the model's performance was compared to standard engineering noise-filtering methods. The results clearly demonstrate that the proposed model outperforms all comparative methods, exhibiting a significant improvement in noise tolerance. This study showcases a straightforward yet effective approach to enhance collision perception in noisy environments.

Re-framing bio-plausible collision detection: identifying shared meta-properties through strategic prototyping.

Insects exhibit remarkable abilities in navigating complex natural environments, whether it be evading predators, capturing prey, or seeking out con-specifics, all of which rely on their compact yet reliable neural systems. We explore the field of bio-inspired robotic vision systems, focusing on the locust inspired Lobula Giant Movement Detector (LGMD) models. The existing LGMD models are thoroughly evaluated, identifying their common meta-properties that are essential for their functionality. This article reveals a common framework, characterized by layered structures and computational strategies, which is crucial for enhancing the capability of bio-inspired models for diverse applications. The result of this analysis is the Strategic Prototype, which embodies the identified meta-properties. It represents a modular and more flexible method for developing more responsive and adaptable robotic visual systems. The perspective highlights the potential of the Strategic Prototype: LGMD-Universally Prototype (LGMD-UP), the key to re-framing LGMD models and advancing our understanding and implementation of bio-inspired visual systems in robotics. It might open up more flexible and adaptable avenues for research and practical applications.

A knock down strategy for rapid, generic, and versatile modelling of muscular dystrophies in 3D-tissue-engineered-skeletal muscle.

BACKGROUND: Human iPSC-derived 3D-tissue-engineered-skeletal muscles (3D-TESMs) offer advanced technology for disease modelling. However, due to the inherent genetic heterogeneity among human individuals, it is often difficult to distinguish disease-related readouts from random variability. The generation of genetically matched isogenic controls using gene editing can reduce variability, but the generation of isogenic hiPSC-derived 3D-TESMs can take up to 6 months, thereby reducing throughput. METHODS: Here, by combining 3D-TESM and shRNA technologies, we developed a disease modelling strategy to induce distinct genetic deficiencies in a single hiPSC-derived myogenic progenitor cell line within 1 week. RESULTS: As proof of principle, we recapitulated disease-associated pathology of Duchenne muscular dystrophy and limb-girdle muscular dystrophy type 2A caused by loss of function of DMD and CAPN3, respectively. shRNA-mediated knock down of DMD or CAPN3 induced a loss of contractile function, disruption of tissue architecture, and disease-specific proteomes. Pathology in DMD-deficient 3D-TESMs was partially rescued by a candidate gene therapy treatment using micro-dystrophin, with similar efficacy compared to animal models. CONCLUSIONS: These results show that isogenic shRNA-based humanized 3D-TESM models provide a fast, cheap, and efficient tool to model muscular dystrophies and are useful for the preclinical evaluation of novel therapies.

Management of a 25-Year-Old Female Patient With Limb-Girdle Muscular Dystrophy With Physiotherapy: A Case Report.

Limb-girdle muscular dystrophy (LGMD) is a collection of neuromuscular diseases that develop gradually and are rare, genetically, and clinically diverse. The weakness in muscles affecting the shoulder and pelvic girdles is a defining feature of LGMD. Calpainopathy is another name for limb-girdle muscular dystrophy type 2A (LGMD2A). Limb-girdle muscular dystrophy type 2A results from alterations in the calpain-3 (CAPN3) gene, which results in a CAPN3 protein shortage. Gower's sign is most commonly found in LGMD2A. The prevalence ranges from one person in every 14,500 to one in every 123,000. We present a case of a 25-year-old hypotensive female patient who complained of weakness in all four limbs and easy fatigue with a positive Gower's sign. For subsequent management, the neurologist referred the patient to the physical therapy department. The physical therapy goals included enhanced muscle strength, increased joint mobility, reduced fatigue, normalizing gait, and building dynamic balance and postural stability. Diagnosing LGMD clinical variability is important, emphasizing the importance of precise subtype identification and tailoring therapy. Tackling specific muscular deficits and functional restrictions emerges as a critical component in the holistic care of LGMD by physiotherapists. Continuous monitoring and evaluation using appropriate scales and measurements are essential for tracking performance and tailoring treatment strategies. Regular follow-up consultations with the physiotherapist are needed to identify changes in an individual's health and alter the treatment plan accordingly.

An Angular Acceleration Based Looming Detector for Moving UAVs.

Visual perception equips unmanned aerial vehicles (UAVs) with increasingly comprehensive and instant environmental perception, rendering it a crucial technology in intelligent UAV obstacle avoidance. However, the rapid movements of UAVs cause significant changes in the field of view, affecting the algorithms' ability to extract the visual features of collisions accurately. As a result, algorithms suffer from a high rate of false alarms and a delay in warning time. During the study of visual field angle curves of different orders, it was found that the peak times of the curves of higher-order information on the angular size of looming objects are linearly related to the time to collision (TTC) and occur before collisions. This discovery implies that encoding higher-order information on the angular size could resolve the issue of response lag. Furthermore, the fact that the image of a looming object adjusts to meet several looming visual cues compared to the background interference implies that integrating various field-of-view characteristics will likely enhance the model's resistance to motion interference. Therefore, this paper presents a concise A-LGMD model for detecting looming objects. The model is based on image angular acceleration and addresses problems related to imprecise feature extraction and insufficient time series modeling to enhance the model's ability to rapidly and precisely detect looming objects during the rapid self-motion of UAVs. The model draws inspiration from the lobula giant movement detector (LGMD), which shows high sensitivity to acceleration information. In the proposed model, higher-order information on the angular size is abstracted by the network and fused with multiple visual field angle characteristics to promote the selective response to looming objects. Experiments carried out on synthetic and real-world datasets reveal that the model can efficiently detect the angular acceleration of an image, filter out insignificant background motion, and provide early warnings. These findings indicate that the model could have significant potential in embedded collision detection systems of micro or small UAVs.

A female case report of LGMD2B with compound heterozygous mutations of the DYSF gene and asymptomatic mutation of the X-linked DMD gene.

We report the case of a 31-year-old Chinese woman with a chief complaint of weakness in the lower limbs, which was diagnosed as limb-girdle muscular dystrophy 2B (LGMD2B) with compound heterozygous mutations of the DYSF gene. Meanwhile, this woman is an asymptomatic carrier with the mutation of the X-linked DMD gene. The electromyography, muscle MRI, and muscle biopsy indicated a chronic myogenic injury with dysferlin deletion. As a result of genetic testing, compound heterozygous G-to-T base substitution at position 5,497 in exon 49 of the DYSF gene, leading to a codon change from glutamic acid to termination codon at position 1,833, and a heterozygous C-to-G base change at position 4,638 + 8 in intron 42 of the DYSF gene with a consequence of splice, which has never been reported, were identified as candidate causative mutations. Unfortunately, DMD gene mutation c.3921+12A>G of the DMD gene on the X chromosome was also found in this patient. Finally, the patient was diagnosed as LGMD2B clinically and genetically. In the previous 2 years, the patient's lower limb weakness became slightly worse, resulting in even the total distance walked than before. Fortunately, during the follow-up, her son had not shown slowness or limitation of movement. Genetic testing by next-generation sequencing confirmed the final diagnosis of LGMD2B, and we identified the novel compound heterozygous variants in the DYSF gene, which is of great significance to the accurate diagnosis of genetically coded diseases. Much attention needs to be paid in clinics toward hereditary neuromuscular diseases with multiple pathogenic gene mutations. Genetic counseling and clinical follow-up should be the priorities in future, and promising treatments are also worth exploring.

Dual Adeno-Associated Virus 9 with Codon-Optimized DYSF Gene Promotes In Vivo Muscle Regeneration and May Decrease Inflammatory Response in Limb Girdle Muscular Dystrophy Type R2.

Dysferlinopathy treatment is an active area of investigation. Gene therapy is one potential approach. We studied muscle regeneration and inflammatory response after injection of an AAV-9 with a codon-optimized DYSF gene. A dual-vector system AAV.DYSF.OVERLAP with overlapping DYSF cDNA sequences was generated. Two AAV vectors were separately assembled by a standard triple-transfection protocol from plasmids carrying parts of the DYSF gene. Artificial myoblasts from dysferlin-deficient fibroblasts were obtained by MyoD overexpression. RT-PCR and Western blot were used for RNA and protein detection in vitro. A dysferlinopathy murine model (Bla/J) was used for in vivo studies. Histological assay, morphometry, and IHC were used for the muscle tissue analysis. Dysferlin was detected in vitro and in vivo at subphysiological levels. RT-PCR and Western Blot detected dysferlin mRNA and protein in AAV.DYSF.OVERLAP-transduced cells, and mRNA reached a 7-fold elevated level compared to the reference gene (GAPDH). In vivo, the experimental group showed intermediate median values for the proportion of necrotic muscle fibers, muscle fibers with internalized nuclei, and cross-sectional area of muscle fibers compared to the same parameters in the control groups of WT and Bla/J mice, although the differences were not statistically significant. The inverse relationship between the dosage and the severity of inflammatory changes in the muscles may be attributed to the decrease in the number of necrotic fibers. The share of transduced myofibers reached almost 35% in the group with the highest dose. The use of two-vector systems based on AAV is justified in terms of therapeutic efficacy. The expression of dysferlin at a subphysiological level, within a short observation period, is capable of inducing the restoration of muscle tissue structure, reducing inflammatory activity, and mitigating necrotic processes. Further research is needed to provide a more detailed assessment of the impact of the transgene and viral vector on the inflammatory component, including longer observation periods.

Caveolin-3 loss linked with the P104L LGMD-1C mutation modulates skeletal muscle mTORC1 signalling and cholesterol homeostasis.

BACKGROUND: Caveolins are the principal structural components of plasma membrane caveolae. Dominant pathogenic mutations in the muscle-specific caveolin-3 (Cav3) gene isoform, such as the limb girdle muscular dystrophy type 1C (LGMD-1C) P104L mutation, result in dramatic loss of the Cav3 protein and pathophysiological muscle weakness/wasting. We hypothesize that such muscle degeneration may be linked to disturbances in signalling events that impact protein turnover. Herein, we report studies assessing the effects of Cav3 deficiency on mammalian or mechanistic target of rapamycin complex 1 (mTORC1) signalling in skeletal muscle cells. METHODS: L6 myoblasts were stably transfected with Cav3P104L or expression of native Cav3 was abolished by CRISPR/Cas9 genome editing (Cav3 knockout [Cav3KO]) prior to performing subcellular fractionation and immunoblotting, analysis of real-time mitochondrial respiration or fixed cell immunocytochemistry. Skeletal muscle from wild-type and Cav3-/- mice was processed for immunoblot analysis of downstream mTORC1 substrate phosphorylation. RESULTS: Cav3 was detected in lysosomal-enriched membranes isolated from L6 myoblasts and observed by confocal microscopy to co-localize with lysosomal-specific markers. Cav3P104L expression, which results in significant (~95%) loss of native Cav3, or CRISPR/Cas9-mediated Cav3KO, reduced amino acid-dependent mTORC1 activation. The decline in mTORC1-directed signalling was detected by immunoblot analysis of L6 muscle cells and gastrocnemius Cav3-/- mouse muscle as judged by reduced phosphorylation of mTORC1 substrates that play key roles in the initiation of protein synthesis (4EBP1S65 and S6K1T389 ). S6K1T389 and 4EBP1S65 phosphorylation reduced by over 75% and 80% in Cav3KO muscle cells and by over 90% and 30% in Cav3-/- mouse skeletal muscle, respectively. The reduction in protein synthetic capacity in L6 muscle cells was confirmed by analysis of puromycylated peptides using the SUnSET assay. Cav3 loss was also associated with a 26% increase in lysosomal cholesterol, and pharmacological manipulation of lysosomal cholesterol was effective in replicating the reduction in mTORC1 activity observed in Cav3KO cells. Notably, re-expression of Cav3 in Cav3KO myoblasts normalized lysosomal cholesterol content, which coincided with a recovery in protein translation and an associated increase in mTORC1-directed phosphorylation of downstream targets. CONCLUSIONS: Our findings indicate that Cav3 can localize on lysosomal membranes and is a novel regulator of mTORC1 signalling in muscle. Cav3 deficiency associated with the Cav3P104L mutation impairs mTORC1 activation and protein synthetic capacity in skeletal muscle cells, which may be linked to disturbances in lysosomal cholesterol trafficking and contribute to the pathology of LGMD-1C.

Convergent escape behaviour from distinct visual processing of impending collision in fish and grasshoppers.

In animal species ranging from invertebrate to mammals, visually guided escape behaviours have been studied using looming stimuli, the two-dimensional expanding projection on a screen of an object approaching on a collision course at constant speed. The peak firing rate or membrane potential of neurons responding to looming stimuli often tracks a fixed threshold angular size of the approaching stimulus that contributes to the triggering of escape behaviours. To study whether this result holds more generally, we designed stimuli that simulate acceleration or deceleration over the course of object approach on a collision course. Under these conditions, we found that the angular threshold conveyed by collision detecting neurons in grasshoppers was sensitive to acceleration whereas the triggering of escape behaviours was less so. In contrast, neurons in goldfish identified through the characteristic features of the escape behaviours they trigger, showed little sensitivity to acceleration. This closely mirrored a broader lack of sensitivity to acceleration of the goldfish escape behaviour. Thus, although the sensory coding of simulated colliding stimuli with non-zero acceleration probably differs in grasshoppers and goldfish, the triggering of escape behaviours converges towards similar characteristics. Approaching stimuli with non-zero acceleration may help refine our understanding of neural computations underlying escape behaviours in a broad range of animal species. KEY POINTS: A companion manuscript showed that two mathematical models of collision-detecting neurons in grasshoppers and goldfish make distinct predictions for the timing of their responses to simulated objects approaching on a collision course with non-zero acceleration. Testing these experimental predictions showed that grasshopper neurons are sensitive to acceleration while goldfish neurons are not, in agreement with the distinct models proposed previously in these species using constant velocity approaches. Grasshopper and goldfish escape behaviours occurred after the stimulus reached a fixed angular size insensitive to acceleration, suggesting further downstream processing in grasshopper motor circuits to match what was observed in goldfish. Thus, in spite of different sensory processing in the two species, escape behaviours converge towards similar solutions. The use of object acceleration during approach on a collision course may help better understand the neural computations implemented for collision avoidance in a broad range of species.

Portrait of Dysferlinopathy: Diagnosis and Development of Therapy.

Dysferlinopathy is a disease caused by a dysferlin deficiency due to mutations in the DYSF gene. Dysferlin is a membrane protein in the sarcolemma and is involved in different functions, such as membrane repair and vesicle fusion, T-tubule development and maintenance, Ca2+ signalling, and the regulation of various molecules. Miyoshi Myopathy type 1 (MMD1) and Limb-Girdle Muscular Dystrophy 2B/R2 (LGMD2B/LGMDR2) are two possible clinical presentations, yet the same mutations can cause both presentations in the same family. They are therefore grouped under the name dysferlinopathy. Onset is typically during the teenage years or young adulthood and is characterized by a loss of Achilles tendon reflexes and difficulty in standing on tiptoes or climbing stairs, followed by a slow progressive loss of strength in limb muscles. The MRI pattern of patient muscles and their biopsies show various fibre sizes, necrotic and regenerative fibres, and fat and connective tissue accumulation. Recent tools were developed for diagnosis and research, especially to evaluate the evolution of the patient condition and to prevent misdiagnosis caused by similarities with polymyositis and Charcot-Marie-Tooth disease. The specific characteristic of dysferlinopathy is dysferlin deficiency. Recently, mouse models with patient mutations were developed to study genetic approaches to treat dysferlinopathy. The research fields for dysferlinopathy therapy include symptomatic treatments, as well as antisense-mediated exon skipping, myoblast transplantation, and gene editing.