Thesis (Ph. D.)--Michigan State University, 2007. / Includes bibliographical references (leaves 143-154). Also available online (PDF file) by a subscription to the set or by purchasing the individual file.

Thesis (Ph. D.)--University of Washington, 2002. / Vita. Includes bibliographical references (leaves 115-132).

Thesis (Ph.D.)--Aberdeen University, 2009. / Title from web page (viewed on Mar. 30, 2010). Includes bibliographical references.

Duchenne Muscular Dystrophy (DMD) is a fatal X-linked disorder affecting 1 out of every 3500 male births. The underlying cause of DMD is mutations within the dystrophin gene resulting in loss of protein expression, which leads to myofiber instability and damage. The constant damage of skeletal muscle causes sustained immune infiltration, marked by increased levels of cytokines, such as TGF-beta. Interestingly, TGF-beta can decrease the myogenic potential of satellite cells, thus preventing muscle regeneration. Previously, our lab has shown that knockdown of the Ste20 Like Kinase, SLK, in normal mammary epithelial cells was sufficient to delay TGF-beta induced epithelial to mesenchymal transition. Therefore, we speculated that decreasing SLK levels would be sufficient to decrease the anti-myogenic effects of TGF-beta both in cultured myoblasts and in a mouse model of muscular dystrophy. In the first section of this study, we explored the effect of muscle specific deletion of SLK on muscle development and regeneration. Skeletal muscle specific deletion of SLK did not impair muscle development, but caused a myopathy in older mice. Additionally, muscle regeneration was delayed, but not inhibited by SLK deletion. These findings indicated that SLK has beneficial roles in skeletal muscle, but was not absolutely required for optimal muscle development and regeneration. In the second section, we investigated the potential for SLK knockdown to mitigate the anti-myogenic effects of TGF-beta in vitro. Decreasing levels of SLK restored myoblast differentiation in the presence of TGF-beta in a p38 dependent manner. In the final section, we determined that SLK levels are elevated in dystrophic muscle and that subsequent deletion of SLK in the mdx mouse enhances terminal differentiation of myoblasts without further exacerbating the pathology of the disease. Collectively, this work demonstrates that SLK inhibition can provide a protective effect against the anti-myogenic effects of TGF-beta via upregulation of p38 activity.

The importance of activity during the development of central components of the nervous system such as the visual system has long been recognized (Wiesel & Hubel 1963) and it is beginning to be understood that sensory experience and motor behavior are equally important for neuromuscular development (Brumley et al. 2015; Sharp & Bekoff 2015). The chick embryo model has proven to be especially useful in studying the relationships among motor behavior, sensory experience, and neuromuscular development (Oppenheim et al. 1978; Sharp & Bekoff 2001) due to its accessibility and early onset of movement behavior. Traditionally, neuromuscular blockers have been used to broadly study the role of neural activity and muscle activity during development (Oppenheim et al. 1978; Ding et al. 1983). In order to noninvasively alter neural activity in specific populations of cells, the Sharp lab has developed an optogenetic approach that allows the expression of ChIEF, a variant of channelrhodopsin-2, in the spinal cord of living chick embryos (Sharp & Fromherz 2011). In order to better understand the unique role that muscle activity plays in neuromuscular development, it would be advantageous to directly and noninvasively control muscle activity through light-activation of ChIEF expressed in muscle fibers. Therefore, the primary objective of this thesis research was to achieve ChIEF expression in the plasma membrane of myotubes in living chick embryos. Initial attempts to express ChIEF in chick muscle resulted in low success rates. The CAG promoter in pPB-ChIEF-Tom, the plasmid vector that encodes ChIEF, was likely hindering expression of ChIEF in muscle tissue. Therefore, standard molecular cloning techniques were used to replace the CAG promoter with the myosin light chain promoter which was known to drive transgene expression in chick muscle (Wang et al. 2011). The new DNA construct that resulted from modifying pPB-ChIEF-Tom was identified as pPB-MLC-ChIEF-Tom (mChIEF). ChIEF was successfully expressed in hindlimb muscles of chick embryos via somite electroporation of mChIEF and observed between E7 and E18. Expression patterns corresponded with the current understanding of muscle progenitor contributions of somites to hindlimb muscles (Rees et al. 2003). ChIEF was located in the outer membrane of muscle fibers on E9, E14, and E18 when tissue was histologically examined in conjunction with myosin heavy chain immunofluorescence. Importantly, light-activation of ChIEF in the hindlimb muscle of living chick embryos resulted in muscle contraction and light-evoked hindlimb movements. In addition to demonstrating the functionality of ChIEF expression, an effort was made to characterize the effects of altered parameters of light stimuli on light-evoked movement and determine whether light-evoked muscle contraction could be used to imitate normal, neuronal muscle control. Light intensity was directly related to amplitude and rate of light-evoked movement. Light duration was directly related to amplitude and latency of peak movement. Unfused and fused tetanus were observed when bursts of short duration light pulses with varying interpulse intervals were used to activate ChIEF. This thesis research strongly suggests that light-activation of ChIEF expressed in living, chick embryo hindlimb muscle results in muscle contractions in manner similar to normal, neurally-driven muscle contraction.

Orientador: Edmundo Medici Filho / Banca: Elisa Emi Tanaka Carloto / Banca: Antônio Francisco David / Banca: Lucia Teramoto / Banca: Julio Cezar de Melo Castilho / Resumo: As alterações anatômicas das vértebras cervicais têm sido empregadas na análise de maturação óssea em Ortodontia e Ortopedia Funcional dos Maxilares. Como essas mudanças são sutis, o objetivo neste estudo foi desenvolver e avaliar um método de análise de maturação óssea empregando-se medidas dessas estruturas. Foram utilizados 246 pares de radiografias cefalométricas laterais e de mão/punho, 135 de indivíduos do sexo feminino e 111 do masculino, divididos em 5 grupos segundo a Curva de Crescimento de Mercadante. Nas radiografias cefalométricas laterais, 15 razões foram obtidas das vértebras C2, C3 e C4, utilizando o programa Radiocef 4.0®. A média de cada razão foi comparada entre os grupos por meio das análises ANOVA e Tukey, que apontaram 10 razões como as que melhor diferem cada fase e que foram utilizadas na criação do novo método. Neste método, denominado Maturação por Razões em Vértebras Cervicais (MRVC), a maturação óssea é classificada em 7 estágios a partir dos resultados das 10 razões. Para sua validação, utilizou-se outra amostra de 58 pares de radiografias cefalométricas laterais e de mão/punho, 28 do sexo feminino e 30 do masculino, submetidas à avaliação de 4 examinadores por três métodos: Curva de Crescimento, Hassel e Farman e método MRVC. Os resultados apontaram melhor desempenho intra e inter-examinadores para a Curva de Crescimento e desempenho semelhante entre o método MRVC e o de Hassel e Farman. Por outro lado, o método MRVC proporcionou resultados próximos ao da Curva de Crescimento na determinação da maturação óssea do indivíduo em relação ao pico do surto de crescimento puberal. Concluiu-se que foi possível criar um método de análise de maturação óssea por medidas das vértebras cervicais em radiografias cefalométricas laterais com eficácia comparável à da Curva de Crescimento. / Abstract: Anatomic alterations of the cervical vertebrae have been used on the assessment of bone maturation in Orthodontic/Functional orthopedics. Since these changes are subtle, our aim was to create and analyze a method for the assessment of skeletal maturation using measurements of the cervical vertebrae. Two hundred and forty six pairs of lateral cephalometric and hand/wrist radiographs (135 male and 111 female) were used on this study. The sample was separated in 5 groups according to the stage of Mercadante's Pubertal Growth Curve. Fifteen ratios were calculated with the measurements of C2, C3 and C4 vertebrae. The mean for each ratio was compared among the groups with the ANOVA and Tukey tests, which indicated 10 ratios that better distinguish each stage. These ratios were used on the development of the method Maturation by Ratios on Cervical Vertebrae (MRCV). This method ranks the maturation in 7 stages based on the results of the ratios. For the validation of the method, four examiners assessed 58 pairs of lateral cephalometric and hand/wrist radiographs using 3 methods: Pubertal Growth Curve, Hassel and Farman and MRCV. Better intra and inter-examiner performance were observed for Pubertal Growth Curve method, while the performance with Hassel and Farman and MRCV were almost the same. However, the results for MRCV were close to the Pubertal Growth Curve on the determination of skeletal maturation related to the peak of the pubertal growth. Based on the results, we concluded that it was possible to create a method for skeletal maturation assessment with measurements of the cervical vertebrae in lateral cephalometric radiographs with efficacy comparable to the Pubertal Growth Curve's. / Doutor

The role of glycogen in endurance performance has been accepted in theory. It has been shown that higher resting muscle glycogen levels prolong endurance performance. On the other hand, low glycogen levels have been associated with fatigue. Ultimately, a person's muscle glycogen level dictates the duration in which an activity can be maintained at a maximal effort, after which time, performance will decrease. As of yet, there has been no evidence as to what happens to the fatigued muscle. Force production in skeletal muscle is dictated by the release and uptake of Ca²⁺ from the sarcoplasmic reticulum (SR). Force production is proportional to [Ca²⁺], as [Ca²⁺] increases so does force. At the point of fatigue, there is a decrease in force production. Since fatigue has been associated with glycogen depletion, it is likely that SR function has been altered causing this decrease in force. The purpose of this study was to determine the effect of glycogen depletion on the SR. Twenty male Sprague-Dawley (Harlan Sprague-Dawley, Indianapolis, IN) rats weighing, 345 ± 70 gm were housed two per cage in the Virginia Tech Lab Animal Resources facility. They were fed ad libitum (Purina Rodent Laboratory Chow and water) until time of experiment. Ten of the rats were used as control animals and the other ten were assigned to the experimental group. Rats were allowed a minimum of 5 days to acclimate to their housing. On the morning of the day of testing, rats were selected in pairs according to the housing cage in an effort to decrease variations in food consumption. To reduce muscle glycogen levels, experimental rats were given an initial injection of either epinephrine (1mg/g: ip) while control rats were injected with saline (equal volume) at 0 hr. Thirty minutes later they received another injection of epinephrine or saline (0.5 mg/g: ip). At the end of the hour the rats were anesthetized with pentobarbital sodium (60 mg/kg:ip) for tissue harvesting. Upon reaching a surgical plane of anesthesia one gastrocnemious muscle was extracted for the muscle glycogen assay and the other removed for SR vesicle preparation. Rats were then euthanized with an overdose of pentobarbital sodium. The tissue was assayed for glycogen and glucose levels as well as for Ca²⁺ uptake and release and ATPase activity. It was found that epinephrine animals had 23% less glycogen than did the control animals and almost twice the amount of glucose (control — 2.9 nmol/g and epinephrine — 5.9 nmol/g). Ca²⁺ uptake rates in epinephrine animals were significantly decreased by 19.7% (p < .05). Control animals had a release rate of 77.15 ± 1.26 nmol/mg/min and epinephrine animals had a release rate of 75.01 ± 1.86 nmol/mg/min. Ca²⁺ release rates were decreased but not significantly. Ca²⁺ stimulated ATPase activity was significantly decreased by 17.7% in epinephrine animals (p < .05). This is one of the first studies that demonstrate that glycogen reduction in a rested muscle causes altered SR function similar to those caused by exercise. This study shows that low glycogen levels are associated with decreased SR function, which is the primary reason for causing the loss of force in muscle. Ultimately, this study suggests that glycogen loading will enhance endurance performance. / Master of Science

Skeletal muscle has been used for biomechanical assist in experimental and clinical studies. Central to the success of these procedures is the generation of sufficient muscle force for the lifetime of the subject. Burst (tetanic) stimulation results in summation of individual twitches and generates higher power output. However, the superiority of paraneural versus intramuscular as well as proximal versus middle and distal intramuscular stimulations remains unclear. Electrophysiological mapping and mechanical performance of seven canine latissimus dorsi muscles were analyzed. The mechanism of higher tension generation produced by: (1) increased temporal summation; (2) greater motor units activated; or (3) result of both were determined. The parameters primarily dependent on the number of activated motor units are significantly greater following paraneural and proximal intramuscular stimulations. The parameters mainly related to temporal summation are not different between various electrode configurations. For intramuscular stimulation, it is the location of interelectrode field rather than the location of the cathode perse that determines the mechanical performance of the skeletal muscle. Furthermore, tension development of skeletal muscle is primary nerve activation rather than direct muscle stimulation. The higher tension generation that resulted from different electrode configurations is produced by activating a higher number of muscle fibers through the neuromuscular junctions.

Both civilian and defense industry rely heavily on robotics which continues to gain a more prominent role. To exemplify, defense strategies in Middle East have relied upon robotic drones and teleoperative assistant robots for mission oriented tasks. These operations have been crucial in saving the lives of soldiers and giving us the edge in mitigating disasters. Future assistive robotics will have direct human interaction and will reside in normal human environments. As the advancement in technology continues to occur, there will be a focus towards eliminating the direct human control and replacing it with higher level autonomy. Further, advancements in electronics and electromechanical components will reduce the cost and makes the assistive robotics accessible to the masses. This thesis focuses on robotic teleoperation technology and the future high level control of assistant robotics. A dexterous 16 degree of freedom hand with bend sensors for precise joint positions was designed, modeled, fabricated and characterized. The design features a unique motor actuation mechanism that was 3D printed to reduce the cost and increase the modularity. The upper body was designed to be biomimetic with dimensions similar to that of a typical six foot tall male. The upper body of the humanoid consists of a 4 degree of freedom shoulder and upper arm with direct feedback at each joint. A theoretical nonlinear switching controller was designed to control these 4 degrees of freedom. The entire system was teleoperative controlled with an Xbox Kinect that tracks the skeletal points of a user and emulates these 3D points to the joints of humanoid upper body. This allows for a direct user control over a robotic assistive upper body with nothing more than a human emulating the desired movements. / Master of Science

Viviparity has evolved multiple times within squamates. Eggshells are reduced in viviparous forms, which reduces calcium available to embryos during development. This study tested the hypothesis that reduced calcium impacts neonates of viviparous forms. Developmental series from oviparous and viviparous populations of Zootoca vivipara (reproductively bimodal) were cleared & stained to reveal bone development. Photographs (high magnification, calibrated to size) were used to obtain measurements of lengths of the body, humerus, femur, skull and lower jaw, and of ossified portions of limb bones. Percent ossification was scored for targeted skull bones. Results were analyzed using general linear models and revealed no differences in ossification in either limbs or skull. Overall size of oviparous neonates was significantly larger. Findings do not support our hypothesis and indicate that reduction in eggshell calcium in embryos of viviparous populations does not negatively impact limb or skull ossification during development but may influence overall size.