Mitochondrial Network Configuration Influences Sarcomere and Myosin Filament Structure in Striated Muscles

Sustained muscle contraction occurs through interactions between actin and myosin filaments within sarcomeres and requires a constant supply of adenosine triphosphate (ATP) from nearby mitochondria. However, it remains unclear how different physical configurations between sarcomeres and mitochondria alter the energetic support for contractile function. Here, we show that sarcomere cross-sectional area (CSA) varies along its length in a cell type-dependent manner where the reduction in Z-disk CSA relative to the sarcomere center is closely coordinated with mitochondrial network configuration. Further, we find myosin filaments near the sarcomere periphery are curved relative to interior filaments with greater curvature for filaments near mitochondria compared to the sarcoplasmic reticulum. Finally, we demonstrate myosin filament lattice spacing is smaller at filament ends than filament centers in a cell type-dependent manner. These data suggest that both sarcomere structure and myofilament interactions are influenced by the location and orientation of mitochondria within a muscle cell.

Accidental monensin poisoning in buffaloes in Bahia, Brazil

ABSTRACT: Monensin is an ionophore antibiotic (IA) widely used for growth promotion and weight gain in the production of ruminants. However, it has caused intoxication in several species, including buffaloes, mainly because of the ignorance or disrespect of the recommendations for use in each animal species. The objective of this study was to describe, for the first time, clinical-epidemiological and anatomopathological data of an outbreak of accidental poisoning by monensin in buffalos and rediscuss the recommendation of the use of IA in the production of this species. The outbreak affected 21 adult buffaloes after consumption of remains from a feed formulated on the farm and whose constituents were mixed by hand. Clinical and first death signs were observed 24 hours after ingestion of this food. In general, the clinical picture was characterized by muscle weakness, tremors, difficulty in locomotion, and decubitus. Fifteen buffaloes presented clinical signs of poisoning (71.5% morbidity), followed by death (100% lethality), after acute to subacute evolution (<24h to 96h). Laboratory tests indicated elevated serum activity of creatine phosphokinase and aspartate aminotransferase enzymes. Three buffaloes underwent necropsy, and samples from several organs were collected for histopathological examination. The main injuries found were hyaline degeneration and multifocal segmental necrosis in the skeletal and cardiac striated muscles (myopathy and degenerative-necrotic multifocal multifocal-necrotic cardiopathy). The diagnosis was confirmed by the toxicological evaluation of suspected ration remains, which detected 461.67mg/kg of monensin. The death of 71.5% buffaloes in this lot occurred due to a succession of errors, which included faults in the formulation of the ration and, above all, due to the use of monensin in a highly sensitive species. Despite the possible beneficial effects of IA use as a dietary supplement for buffaloes, we are of the opinion that IAs should never be used in bubalinoculture since any increment in production does not compensate for the imminent risk of death due to a small safety margin for this species and the absence of antidotes.

Multiple regions of junctin drive interaction with calsequestrin-1 and localization at triads in skeletal muscle

Junctin is a transmembrane protein of striated muscles, localized at the junctional sarcoplasmic reticulum (j-SR). It is characterized by a luminal C-terminal tail, through which it functionally interacts with calsequestrin and the ryanodine receptor. Interaction with calsequestrin was ascribed to the presence of stretches of charged amino acids. However, the regions able to bind calsequestrin have not been defined in detail. We report here that, in non-muscle cells, junctin and calsequestrin assemble in long linear regions within the endoplasmic reticulum, mirroring the formation of calsequestrin polymers. In differentiating myotubes, the two proteins co-localize at triads, where they assemble with other j-SR proteins. By performing GST pull-down assays with distinct regions of the junctin tail, we identified two KEKE motifs able to bind calsequestrin. In addition, stretches of charged amino acids downstream these motifs were found to be also able to bind calsequestrin and the ryanodine receptor. Deletion of even one of these regions impaired the ability of junctin to localize at the j-SR, suggesting that interaction with other proteins at this site represents a key element in junctin targeting.

Preclinical Advances of Therapies for Laminopathies

Laminopathies are a group of rare disorders due to mutation in LMNA gene. Depending on the mutation, they may affect striated muscles, adipose tissues, nerves or are multisystemic with various accelerated ageing syndromes. Although the diverse pathomechanisms responsible for laminopathies are not fully understood, several therapeutic approaches have been evaluated in patient cells or animal models, ranging from gene therapies to cell and drug therapies. This review is focused on these therapies with a strong focus on striated muscle laminopathies and premature ageing syndromes.

Disrupted Calcium Homeostasis in Duchenne Muscular Dystrophy: A Common Mechanism behind Diverse Consequences

Duchenne muscular dystrophy (DMD) leads to disability and death in young men. This disease is caused by mutations in the DMD gene encoding diverse isoforms of dystrophin. Loss of full-length dystrophins is both necessary and sufficient for causing degeneration and wasting of striated muscles, neuropsychological impairment, and bone deformities. Among this spectrum of defects, abnormalities of calcium homeostasis are the common dystrophic feature. Given the fundamental role of Ca2+ in all cells, this biochemical alteration might be underlying all the DMD abnormalities. However, its mechanism is not completely understood. While abnormally elevated resting cytosolic Ca2+ concentration is found in all dystrophic cells, the aberrant mechanisms leading to that outcome have cell-specific components. We probe the diverse aspects of calcium response in various affected tissues. In skeletal muscles, cardiomyocytes, and neurons, dystrophin appears to serve as a scaffold for proteins engaged in calcium homeostasis, while its interactions with actin cytoskeleton influence endoplasmic reticulum organisation and motility. However, in myoblasts, lymphocytes, endotheliocytes, and mesenchymal and myogenic cells, calcium abnormalities cannot be clearly attributed to the loss of interaction between dystrophin and the calcium toolbox proteins. Nevertheless, DMD gene mutations in these cells lead to significant defects and the calcium anomalies are a symptom of the early developmental phase of this pathology. As the impaired calcium homeostasis appears to underpin multiple DMD abnormalities, understanding this alteration may lead to the development of new therapies. In fact, it appears possible to mitigate the impact of the abnormal calcium homeostasis and the dystrophic phenotype in the total absence of dystrophin. This opens new treatment avenues for this incurable disease.

THE RESULTS OF THE SEARCH FOR LABORATORY SIGNS OF AUTOIMMUNE REACTIONS TO CEREBRAL AND EXTRACEREBRAL AUTOANTIGENS IN CHILDREN WITH AUTISM SPECTRUM DISORDERS ASSOCIATED WITH GENETIC DEFICIENCY OF THE FOLATE CYCLE

Relevance. The results of five meta-analyzes of randomized controlled clinical trials indicate an association between genetic deficiency of the folate cycle (GDFC) and autism spectrum disorders (ASD) in children. Autoimmune mechanisms play a special role in the pathogenesis of encephalopathy in children with ASD associated with GDFC. Objective: to study the structure of autoimmune reactions in children with ASD associated with GDFC, according to the accumulated evidence base and to identify associations of laboratory signs of autoimmunity and microorganisms to improve understanding of encephalopathy pathogenesis and diagnostic, monitoring and treatment algorithms. Materials and methods. The medical data of 225 children aged 2 to 9 years with GDFC, who had clinical manifestations of ASD (183 boys and 42 girls) were retrospectively analyzed. The diagnosis of ASD was made by child psychiatrists according to the criteria DSM-IV-TR (Diagnostic and Statistical Manual of mental disorders) and ICD-10 (The International Statistical Classification of Diseases and Related Health Problems) (study group; SG). The control group (CG) included 51 clinically healthy children (37 boys and 14 girls) of similar age and gender distribution who did not suffer from GDFC and ASD. Pathogenic polymorphic variants of folate cycle genes were determined by PCR with restriction (Sinevo, Ukraine). Autoantibodies to autoantigens of CNS subcortical ganglion neurons in blood serum were determined using a Cunningham panel (Moleculera Labs, Inc, USA). Serum autoantibodies to neurons of the mesolimbic system of the brain were identified by ELISA (MDI Limbach Berlin GmbH, Germany). Autoimmunization to myelin was assessed by serum autoantibody titer to basic myelin protein (ELISA) and signs of neutrophil and CD8+ T-lymphocyte sensitization to hemispheric white matter autoantigens (cell-based assay; department of neuroimmunology at the Neurosurgery Institute; Ukraine). Serum autoantibodies to nuclei of connective tissue cells and striated muscle proteins were determined by western blot analysis (Sinevo, Ukraine). To determine the significance of the differences between the indicators in the observation groups, we used the Student's parametric T-test with the confidence probability p and the nonparametric criterion – the number of signs Z according to Urbach Yu.V. The odds ratio (OR) and the 95% confidence interval (95% CI) were used to study the associations between the studied indicators. The study was performed as a fragment of research work commissioned by the Ministry of Health of Ukraine (№ state registration 0121U107940). Research. Positive results of the Cunningham panel occurred in 32%, laboratory signs of autoimmunization to neurons of the mesolimbic system – 36%, myelin of white matter of the hemispheres – 43%, nuclei autoantigens of connective tissue cells – 53%, proteins of striated muscles – 48% of cases among children SG (in general – 68% of cases; p < 0.05; Z < Z0.05). Serological signs of autoimmunization to autoantigens of the subcortical ganglia of the cerebral hemispheres were associated with Streptococcus pyogenes and Borrelia, to neurons of the mesolimbic system – EBV, HHV-6, HHV-7, Toxoplasma and TTV, to CNS myelin – EBV, HHV-6, HHV-7, Borrelia and TTV, to the nuclei of connective tissue cells and striated muscles – EBV, HHV-6, HHV-7, Borrelia and TTV. Conclusions. In children with ASD associated with GDFC laboratory sings of microbial-induced autoimmunity to a number of cerebral and extracerebral autoantigens has been evaluated, which affects the mental and physical health of patients and is a potential target for effective therapeutic interventions.

Electromyographic features of the perineum and pelvic floor in patients with an artificial bladder

Aim – to assess the electromyographic features of the pelvic floor muscles and the sphincter apparatus in patients who underwent radical cystprostatectomy with the formation of an artificial bladder.The main study group consisted of 57 patients with muscle invasive bladder cancer who underwent a standard radical cystprostatectomy with ileocystoplasty. The study of the pelvic floor muscular bioelectric activity with computed electromyography (EMG) of the sphincter apparatus of the pelvic organs was carried out on a 2-channel computer electromyograph "NeuroTrac ™ MyoPlus4". The obtained results of the study of the bioelectric activity of the pelvic floor muscles showed a decrease in the amplitude of contractions in the Work Average mode by 42.1% (p≤0.001) for the perineal electrode, and by 35.7% (p≤0.05) for the rectal electrode, compared with the control group, which indicates a low contractility of the muscular diaphragm of the pelvis and may cause incontinence in patients with an artificial bladder. The average deviation over the entire duration of the session in Work mode in the group of patients with neobladder in channels A and B was 2.3 (p≤0.05) and 1.9 (p≤0.05) times higher, respectively, compared with control group. These data indicate an imbalance in the muscle tone of the pelvic floor in patients after extensive reconstructive intervention on the pelvic organs and can potentiate urodynamic disorders in the study group of patients. The average amplitude (Rest Average) of the activity of biopotentials in the resting state of the pelvic floor muscles along channels A and B is higher by 42.4% and 47.6% (p≤0.05), in comparison with the control group, which indicates insufficient relaxation and rest of striated muscles and sphincter. Despite the change in bioelectric potentials from the rectal electrode in the study group of patients, there were no signs of functional insufficiency of the anal sphincter, in contrast to the severity of urinary incontinence, which correlated and corresponded to the results of bioelectrical changes obtained through channels A and B, up to oscillations and loss of complete control. The EMG analysis of the pelvic floor muscles revealed characteristic changes in the biopotentials of the pelvic sphincters and indicated their relationship with the clinical features of the rehabilitation of this group of patients and the prospect of including the biofeedback method. It is a derivative form of the electromyographic signal in the treatment of incontinence in patients with neobladder.

Troponin Variants as Markers of Skeletal Muscle Health and Diseases

Ca2+-regulated contractility is a key determinant of the quality of muscles. The sarcomeric myofilament proteins are essential players in the contraction of striated muscles. The troponin complex in the actin thin filaments plays a central role in the Ca2+-regulation of muscle contraction and relaxation. Among the three subunits of troponin, the Ca2+-binding subunit troponin C (TnC) is a member of the calmodulin super family whereas troponin I (TnI, the inhibitory subunit) and troponin T (TnT, the tropomyosin-binding and thin filament anchoring subunit) are striated muscle-specific regulatory proteins. Muscle type-specific isoforms of troponin subunits are expressed in fast and slow twitch fibers and are regulated during development and aging, and in adaptation to exercise or disuse. TnT also evolved with various alternative splice forms as an added capacity of muscle functional diversity. Mutations of troponin subunits cause myopathies. Owing to their physiological and pathological importance, troponin variants can be used as specific markers to define muscle quality. In this focused review, we will explore the use of troponin variants as markers for the fiber contents, developmental and differentiation states, contractile functions, and physiological or pathophysiological adaptations of skeletal muscle. As protein structure defines function, profile of troponin variants illustrates how changes at the myofilament level confer functional qualities at the fiber level. Moreover, understanding of the role of troponin modifications and mutants in determining muscle contractility in age-related decline of muscle function and in myopathies informs an approach to improve human health.

Complexity of skeletal muscle degeneration: multi-systems pathophysiology and organ crosstalk in dystrophinopathy

Duchenne muscular dystrophy is a highly progressive muscle wasting disorder due to primary abnormalities in one of the largest genes in the human genome, the DMD gene, which encodes various tissue-specific isoforms of the protein dystrophin. Although dystrophinopathies are classified as primary neuromuscular disorders, the body-wide abnormalities that are associated with this disorder and the occurrence of organ crosstalk suggest that a multi-systems pathophysiological view should be taken for a better overall understanding of the complex aetiology of X-linked muscular dystrophy. This article reviews the molecular and cellular effects of deficiency in dystrophin isoforms in relation to voluntary striated muscles, the cardio-respiratory system, the kidney, the liver, the gastrointestinal tract, the nervous system and the immune system. Based on the establishment of comprehensive biomarker signatures of X-linked muscular dystrophy using large-scale screening of both patient specimens and genetic animal models, this article also discusses the potential usefulness of novel disease markers for more inclusive approaches to differential diagnosis, prognosis and therapy monitoring that also take into account multi-systems aspects of dystrophinopathy. Current therapeutic approaches to combat muscular dystrophy are summarised.