Browsing by Subject "dystrophin"

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    Acute contraction-induced strength loss, muscle inexcitability and recovery after exercise in males with Duchenne muscular dystrophy
    (2020-11) Trost, Joyceann
    This dissertation had three objectives with the overall goal to explore contraction-induced strength loss and recovery in males with Duchenne muscular dystrophy (DMD) specifically to: 1) evaluate a novel protocol, combining voluntary and evoked contractions to measure strength and excitability of wrist extensor muscles, for safety, feasibility, reliability and discriminant validity for males with DMD and aged-matched controls, 2) explore strength loss and muscle fiber inexcitability contribution to strength loss after submaximal isometric contractions in males with DMD, and. 3) determine the recovery of strength and muscle excitability immediately following contraction-induced force loss in males with DMD. Through the literature review, I discovered a need for a reliable and valid measurement protocol of muscle contractile function that could be used for DMD across all disease levels. Several measures for walking and upper extremity function are currently used to measure outcomes during clinical trials. However, none of the current measurements for DMD incorporate a way to quantitatively measure both voluntary and evoked strength along with muscle excitability over disease progression or in response to an intervention. The key to the assessment and protocol design needed to be feasibility and safety for boys and men with DMD at different stages of the disease process. I was able to design and evaluate a novel protocol, combining voluntary and evoked contractions to measure strength and excitability of wrist extensor muscles for safety, feasibility, reliability and discriminant validity between males with DMD and controls (Chapter 3). Wrist extensor muscle strength and excitability were assessed in males with DMD (N=10; mean 15.4 [SD 5.9] years) (Brooke 1-6) and age-matched healthy male controls (N=15; mean 15.5 [SD 5.0] years). Torque and EMG measurements were analyzed under maximum voluntary and stimulated conditions at two visits. I discovered that our protocol of multiple maximal voluntary contractions (MVC) and evoked twitch contractions was feasible and safe, with 96% of the participants able to complete the assessment protocol maintaining >93% strength both for DMD and controls (P≥0.074). Reliability was excellent for voluntary and evoked measurements. Torque, EMG and timing of twitch onset measurements discriminated between DMD and controls (P<0.001). This first part of the study demonstrated a useful protocol for measuring skeletal muscle function in clinical trials in males with DMD across various ages and disease levels. The second part of the study addressed in the context of a neuromuscular disease the concern of injury to dystrophic deficient skeletal muscle during and after repeated contractions (exercise) that result in acute strength loss. I knew that a feature of dystrophin-deficient skeletal muscle in the mdx mouse model was a hypersensitivity to strength loss from eccentric exercise due to fibers becoming unexcitable. I wanted to explore how this feature translated to humans lacking dystrophin. We hypothesized that there would be no difference in strength loss during exercise and that males with Duchenne muscular dystrophy (DMD) would have more significant impairment in muscle excitability corresponding with a loss of strength than age-matched controls (Chapter 4). Males with DMD and a group of age-matched controls performed a sub-maximal voluntary isometric wrist extensor exercise protocol until 55% of maximal voluntary contraction (MVC) could no longer be sustained. Voluntary and evoked force and EMG were accessed before, during, and after exercise. There was a significant interaction (time*group) (p<0.001) for MVC torque during exercise, suggesting that the two groups were different in how they reached muscle fatigue. No difference was measured between groups in MVC torque decrement at the time of exercise cessation (by design). Evoked twitch torque decrement was 34% for DMD and 36% for control with no group*time interaction observed (p=0.834). Muscle excitability contributed to the evoked torque variance in DMD (76%) and control (59%). The groups were not different in RMS EMG decrement (P=0.986) or M-wave decrement (P=0.911) during exercise, which does not support our hypothesis that the DMD groups had a more considerable decline in muscle excitability with fatigue during exercise. Lastly, the recovery of both strength and muscle excitability was explored and compared between groups. The DMD group recovered MVC baseline strength by 10 minutes post-exercise (P=0.530) and evoked torque by 5 minutes (P=0.266). In contrast, controls were still different from MVC and evoked torque baseline at 15 minutes after exercise (P<0.002). Strength loss from submaximal intermittent isometric exercise does not result in more significant impairment of muscle excitability is transient, and recovers faster in males with DMD than controls, suggesting a different mechanism of peripheral fatigue between the groups. The work contained in my dissertation begins to address the fear of exercise that has been a common concern for individuals with DMD and will contribute to knowledge of evidence-based exercise prescription in the future.
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    Structural And Intrinsic Disorder In The Regulation Of Protein-Protein Interactions
    (2019-06) Fealey, Michael E.
    This thesis applied spectroscopy and molecular dynamics simulation to study the structural biology of actin-binding domains (ABDs) from the spectrin superfamily of proteins as well as an intrinsically disordered region (IDR) of an integral membrane protein called synaptotagmin 1. In the former case, the structural hypothesis being tested was that actin-binding domains exist in distinct conformational states that are either permissive to or inhibitory towards binding of actin filaments. This question was probed using pulsed-EPR, which measured distances between the calponin homology (CH) domains that make up the ABD as proxy for conformation in the presence or absence of actin or with and without disease-causing mutation. The initial hypothesis of a closed compact state being unable to bind actin and an open extended state being binding-competent was largely supported by the data. However, the hypothesis was ultimately refined to conclude that an “open” state is likely to still be a fairly collapsed structure that is dynamically disordered. With this model, future efforts will be able use the model to look for small molecules that perturb the conformational equilibrium of ABDs harboring disease-causing mutations in potentially therapeutically efficacious ways. Moreover, the model can be tested in other ABDs of the protein superfamily to assess similarities and differences in mechanism. In the case of the intrinsically disordered region of synaptotagmin 1, it was hypothesized that a post-translational modification, specifically phosphorylation of a threonine residue, caused a structural change in the IDR that then results in a change in neurotransmitter release. This hypothesis was also tested with spectroscopic methods, mainly FRET and circular dichroism, but also with molecular dynamics. It was found that mimicking the low dielectric environment of the membrane with co-solvents in solution and artificially in silico caused the synaptotagmin 1 IDR to fold into helical structure. The post-translational modification, however, was found to interfere with the formation of helical structure, providing a still incomplete but novel molecular explanation for the effect it has on potentiation of neurotransmitter release observed in vivo. At the very least, the structural model provides a working hypothesis that can be further explored in further work.