Respiratory and upper limb function as outcome measures in ambulant and non-ambulant subjects with Duchenne muscular dystrophy: A prospective multicentre study.

The field of translational research in Duchenne muscular dystrophy (DMD) has been transformed in the last decade by a number of therapeutic targets, mostly studied in ambulant patients. A paucity of studies focus on measures that capture the non-ambulant stage of the disease, and the transition between the ambulant and non-ambulant phase. In this prospective natural history study, we report the results of a comprehensive assessment of respiratory, upper limb function and upper limb muscle strength in a group of 89 DMD boys followed in 3 European countries, 81 receiving corticosteroids, spanning a wide age range (5-18 years) and functional abilities, from ambulant (n = 60) to non-ambulant (n = 29). Respiratory decline could be detected in the early ambulatory phase using Peak Expiratory Flow percentage predicted (PEF%), despite glucocorticoid use (mean annual decline: 4.08, 95% CI [-7.44,-0.72], p = 0.02 in ambulant; 4.81, 95% CI [-6.79,-2.82], p < 0.001 in non-ambulant). FVC% captured disease progression in non-ambulant DMD subjects, with an annual loss of 5.47% (95% CI [-6.48,-4.45], p < 0.001). Upper limb function measured with the Performance of Upper Limb (PUL 1.2) showed an annual loss of 4.13 points (95% CI [-4.79,3.47], p < 0.001) in the non-ambulant cohort. Measures of upper limb strength (MyoGrip and MyoPinch) showed a continuous decline independent of the ambulatory status, when reported as percentage predicted (grip force -5.51%, 95% CI [-6.54,-4.48], p < 0.001 in ambulant and a slower decline -2.86%; 95% CI -3.29,-2.43, p < 0.001, in non-ambulant; pinch force: -2.66%, 95% CI [-3.82,-1.51], p < 0.001 in ambulant and -2.23%, 95% CI [-2.92,-1.53], p < 0.001 in non-ambulant). Furthermore, we also explored the novel concept of a composite endpoint by combining respiratory, upper limb function and force domains: we were able to identify clear clinical progression in patients in whom an isolated measurement of only one of these domains failed to appreciate the yearly change. Our study contributes to the field of natural history of DMD, linking the ambulant and non-ambulant phases of the disease, and suggests that composite scores should be explored further.

Development of a patient-reported outcome measure for upper limb function in Duchenne muscular dystrophy: DMD Upper Limb PROM.

AIM: To develop a patient-reported outcome measure (PROM) assessing upper limb function related to activities of daily living (ADL) that cannot be observed in a clinical setting, specifically for patients with Duchenne muscular dystrophy (DMD) across a wide age range, applicable in the different stages of the disease. METHOD: The developmental process was based on US Food and Drug Administration guidelines. This included item generation from a systematic review of existing tools and expert opinion on task difficulty and relevance, involving individuals with DMD. Cultural aspects affecting ADL were taken into consideration to make this tool applicable to the broad DMD community. Items were selected in relation to a conceptual framework reflecting disease progression covering the full range of upper limb function across different ADL domains. RESULTS: After pilot testing and iterative Rasch analyses, redundant or clinically irrelevant items were removed. The final questionnaire consists of 32 items covering four domains of ADL (food, self-care, household and environment, leisure and communication). Test-retest reliability was excellent. INTERPRETATION: A DMD-specific upper limb PROM was developed on the basis of clinical relevance and psychometric robustness. Its main purpose is to document the patient self-reported natural history of DMD and assess the efficacy of interventions.

Effect of temperature on the working stroke of muscle myosin.

Muscle contraction is due to myosin motors that transiently attach with their globular head to an actin filament and generate force. After a sudden reduction of the load below the maximum isometric force (T0), the attached myosin heads execute an axial movement (the working stroke) that drives the sliding of the actin filament toward the center of the sarcomere by an amount that is larger at lower load and is 11 nm near zero load. Here, we show that an increase in temperature from 2 to 17 degrees C, which increases the average isometric force per attached myosin head by 60%, does not affect the amount of filament sliding promoted by a reduction in force from T0 to 0.7T0, whereas it reduces the sliding under low load by 2.5 nm. These results exclude the possibility that the myosin working stroke is due to the release of the mechanical energy stored in the initial endothermic force-generating process and show that, at higher temperatures, the working stroke energy is greater because of higher force, although the stroke length is smaller at low load. We conclude the following: (i) the working stroke is made by a series of state transitions in the attached myosin head; (ii) the temperature increases the probability for the first transition, competent for isometric force generation; and (iii) the temperature-dependent rise in work at high load can be accounted for by the larger free energy drop that explains the rise in isometric force.

Laminopathies affecting skeletal and cardiac muscles: clinical and pathophysiological aspects.

Laminopathies are caused by mutations in the LMNA gene encoding the ubiquitous proteins lamins A/C that are components of the lamina, a fibrous meshwork located at the inner face of the nuclear envelope. Laminopathies may affect one or several tissues such as striated muscles, peripheral nerves and adipose tissue in isolate or combined fashion. This review focuses on laminopathies affecting the striated muscle tissue only, namely Emery-Dreifuss muscular dystrophy (EDMD), limb girdle muscular dystrophy type 1B (LGMD1B) and dilated cardiomyopathy with conduction defects (DCM-CD). The phenotype of animal models in which the same mutation as that identified in EDMD or DCM-CD patients has been reproduced is presented as well as the pathophysiological mechanisms known to date.

Temperature dependence of the force-generating process in single fibres from frog skeletal muscle.

Generation of force and shortening in striated muscle is due to the cyclic interactions of the globular portion (the head) of the myosin molecule, extending from the thick filament, with the actin filament. The work produced in each interaction is due to a conformational change (the working stroke) driven by the hydrolysis of ATP on the catalytic site of the myosin head. However, the precise mechanism and the size of the force and length step generated in one interaction are still under question. Here we reinvestigate the endothermic nature of the force-generating process by precisely determining, in tetanized intact frog muscle fibres under sarcomere length control, the effect of temperature on both isometric force and force response to length changes. We show that raising the temperature: (1) increases the force and the strain of the myosin heads attached in the isometric contraction by the same amount (approximately 70 %, from 2 to 17 degrees C); (2) increases the rate of quick force recovery following small length steps (range between -3 and 2 nm (half-sarcomere)-1) with a Q10 (between 2 and 12 degrees C) of 1.9 (releases) and 2.3 (stretches); (3) does not affect the maximum extent of filament sliding accounted for by the working stroke in the attached heads (10 nm (half-sarcomere)-1). These results indicate that in isometric conditions the structural change leading to force generation in the attached myosin heads can be modulated by temperature at the expense of the structural change responsible for the working stroke that drives filament sliding. The energy stored in the elasticity of the attached myosin heads at the plateau of the isometric tetanus increases with temperature, but even at high temperature this energy is only a fraction of the mechanical energy released by attached heads during filament sliding.

Intracellular pH regulation in isolated fast-twitch skeletal muscle from dystrophin-deficient mouse.

In muscles from anaesthetized dystrophin-deficient mdx mice, exercise results in a stronger acidification and a slower intracellular pH recovery compared to control mice. We examined whether this observation could be attributed to defective H+-carriers in dystrophin-lacking muscles. Immunohistochemistry and Western blots revealed no defect in mdx muscles for the presence of the lactate-/H+co-transporter MCT4 and of the Na+/H+ antiporter NHE1, the main H+-carriers active in fast-twitch skeletal muscle after exercise. Functional tests of the H+-transporters, on isolated muscles submitted to identical flow of superfusion, were performed in conditions meant to lower intracellular pH: repetitive electrical stimulation or NH4Cl pre-pulse. These revealed no defect in intracellular pH recovery in mdx muscles. Therefore, we conclude that impaired intracellular pH regulation in anaesthetized mdx mice is not attributable to a reduced presence or activity of H+-extruders. We propose that CO2 washout might be slowed down in vivo in mdx muscles because of the defective vascular response in contracting muscles from these mice.

Effect of adrenaline on the post-tetanic potentiation in mouse skeletal muscle.

We report the influence of adrenergic stimulation on the amplitude and time course of post-tetanic potentiation of twitch contraction. This was complemented by measurements of the peak of [Ca2+]i transients in twitches and of the level of myosin light chain 2 (LC2) phosphorylation, before, 20 and 300 s after the conditioning tetanus. Soon after the tetanus, twitch potentiation and increases of LC2 phosphorylation and of [Ca2+]i peak were similar in control conditions and in the presence of adrenaline. In control conditions, twitch potentiation, LC2 phosphorylation and [Ca2+]i peak returned to, or close to, pre-tetanic values in 300 s. On the contrary, in the presence of adrenaline, twitch potentiation and LC2 phosphorylation were partially or fully maintained respectively, while the increase of [Ca2+]i peak was not. This situation allowed us to analyse the relative contributions of elevated LC2 phosphorylation and [Ca2+]i peak in the twitch post-tetanic potentiation phenomenon. Moreover, it was shown that the increase of LC2 phosphorylation (up to 0.5 mol P/mol LC2) affected neither the kinetic parameters of the twitch nor the maximal velocity of shortening. It is proposed that the maintenance of LC2 phosphorylation in the presence of adrenaline results from the inhibition of myosin light chain phosphatase. This could be achieved through the production of the active, phosphorylated form of the inhibitor-1, an endogenous inhibitor, which binds to the catalytic sub-units common to class 1 protein phosphatases.