Deleting titin's C-terminal PEVK exons increases passive stiffness, alters splicing, and induces cross-sectional and longitudinal hypertrophy in skeletal muscle

The Proline, Glutamate, Valine and Lysine-rich (PEVK) region of titin constitutes an entropic spring that provides passive tension to striated muscle. To study the functional and structural repercussions of a small reduction in the size of the PEVK region, we investigated skeletal muscles of a mouse with the constitutively expressed C-terminal PEVK exons 219–225 deleted, the Ttn(Δ219–225) model (MGI: Ttn(TM 2.1Mgot)). Based on this deletion, passive tension in skeletal muscle was predicted to be increased by ∼17% (sarcomere length 3.0 μm). In contrast, measured passive tension (sarcomere length 3.0 μm) in both soleus and EDL muscles was increased 53 ± 11% and 62 ± 4%, respectively. This unexpected increase was due to changes in titin, not to alterations in the extracellular matrix, and is likely caused by co-expression of two titin isoforms in Ttn(Δ219–225) muscles: a larger isoform that represents the Ttn(Δ219–225) N2A titin and a smaller isoform, referred to as N2A2. N2A2 represents a splicing adaption with reduced expression of spring element exons, as determined by titin exon microarray analysis. Maximal tetanic tension was increased in Ttn(Δ219–225) soleus muscle (WT 240 ± 9; Ttn(Δ219–225) 276 ± 17 mN/mm2), but was reduced in EDL muscle (WT 315 ± 9; Ttn(Δ219–225) 280 ± 14 mN/mm2). The changes in active tension coincided with a switch toward slow fiber types and, unexpectedly, faster kinetics of tension generation and relaxation. Functional overload (FO; ablation) and hindlimb suspension (HS; unloading) experiments were also conducted. Ttn(Δ219–225) mice showed increases in both longitudinal hypertrophy (increased number of sarcomeres in series) and cross-sectional hypertrophy (increased number of sarcomeres in parallel) in response to FO and attenuated cross-sectional atrophy in response to HS. In summary, slow- and fast-twitch muscles in a mouse model devoid of titin's PEVK exons 219–225 have high passive tension, due in part to alterations elsewhere in splicing of titin’s spring region, increased kinetics of tension generation and relaxation, and altered trophic responses to both functional overload and unloading. This implicates titin’s C-terminal PEVK region in regulating passive and active muscle mechanics and muscle plasticity

Titin contribution to active muscle

The aim of this dissertation is to compare the traditional ‘muscle as motor’ viewpoint to the alternative viewpoint ‘muscle as composite material’. We make three parts to achieve the aim. We first investigate whether the contribution of activation and length perturbation on muscle force depends on length history. The effect of length history is to alter muscle stiffness. The alteration of muscle stiffness determines the role of activation and length perturbation in determining muscle force. That is, muscle responds to activation and length perturbation depending on length history. Muscle models based on the ‘muscle as motor’ viewpoint fail to predict muscle force accurately under dynamic conditions in part because they cannot account for the history dependence of muscle force. An alternative muscle viewpoint should be able to explain the history dependent muscle force to enlarge our knowledge about how muscle works under dynamic conditions. In the second part, we focus on the ratio of muscle force to stiffness. The ratio of force to stiffness has been believed to be relatively constant during isometric contraction because both isometric force and stiffness are linearly related to the number of formed cross-bridge. However, the ratio depends on shortening velocity as well as is not constant during the isometric force redevelopment period following active shortening. This finding invokes ‘stress-induced inhibition’ where weakly-bound cross bridge generated by active shortening contributes only to stiffness with no contribution to force. The weakly-bound cross bridge is able to explain non-zero y-intercepts of the relationship between force and stiffness, but not the lower slope for the slower velocity. In the ‘stress-induced inhibition’, the slower shortening velocity should distort actin more due to the bigger stress. The more distorted actin induces the bigger number of weakly-bound cross bridge, which should result in the bigger slope of the relationship between force and stiffness. This is not compatible with our results. Another possible explanation of the finding is a change in titin stiffness. According to the winding filament hypothesis, titin equilibrium length is modulated by cross-bridge force. Active shortening at different velocity could generate the different initial conditions for titin strain and equilibrium position, and then titin differently responds to an increase in cross bridge force depending on the initial conditions. The tunable titin stiffness contributes to the variable muscle stiffness. Finally, this dissertation develops a titin-clutch model based on a composite material viewpoint. The titin-clutch model predicts frequency-dependent muscle force better than the Hill model. The model has three subunits: a contractile, titin, and series elastic element. The pulley connects the titin to the contractile element in series and parallel. The history-dependent pulley position implements the history-dependent contributions of length perturbation and activation to force. When the contractile force increases, pulley rotates counterclockwise direction, then titin element wraps around a pulley. The interaction between them regulates titin stiffness. The dissertation concludes that incorporating a tunable titin spring in muscle models improves the predictions of muscle force under dynamic conditions

Clinical, Biomechanical, and Physiological Translational Interpretations of Human Resting Myofascial Tone or Tension

Background: Myofascial tissues generate integrated webs and networks of passive and active tensional forces that provide stabilizing support and that control movement in the body. Passive [central nervous system (CNS)–independent] resting myofascial tension is present in the body and provides a low-level stabilizing component to help maintain balanced postures. This property was recently called “human resting myofascial tone” (HRMT). The HRMT model evolved from electromyography (EMG) research in the 1950s that showed lumbar muscles usually to be EMG-silent in relaxed gravity-neutral upright postures. Methods: Biomechanical, clinical, and physiological studies were reviewed to interpret the passive stiffness properties of HRMT that help to stabilize various relaxed functions such as quiet balanced standing. Biomechanical analyses and experimental studies of the lumbar multifidus were reviewed to interpret its passive stiffness properties. The lumbar multifidus was illustrated as the major core stabilizing muscle of the spine, serving an important passive biomechanical role in the body. Results: Research into muscle physiology suggests that passive resting tension (CNS-independent) is generated in sarcomeres by the molecular elasticity of low-level cycling cross-bridges between the actomyosin filaments. In turn, tension is complexly transmitted to intimately enveloping fascial matrix fibrils and other molecular elements in connective tissue, which, collectively, constitute the myofascial unit. Postural myofascial tonus varies with age and sex. Also, individuals in the population are proposed to vary in a polymorphism of postural HRMT. A few people are expected to have outlier degrees of innate postural hypotonicity or hypertonicity. Such biomechanical variations likely predispose to greater risk of related musculoskeletal disorders, a situation that deserves greater attention in clinical practice and research. Axial myofascial hypertonicity was hypothesized to predispose to ankylosing spondylitis. This often-progressive deforming condition of vertebrae and sacroiliac joints is characterized by stiffness features and particular localization of bony lesions at entheseal sites. Such unique features imply concentrations and transmissions of excessive force, leading to tissue micro-injury and maladaptive repair reactions. Conclusions: The HRMT model is now expanded and translated for clinical relevance to therapists. Its passive role in helping to maintain balanced postures is supported by biomechanical principles of myofascial elasticity, tension, stress, stiffness, and tensegrity. Further research is needed to determine the molecular basis of HRMT in sarcomeres, the transmission of tension by the enveloping fascial elements, and the means by which the myofascia helps to maintain efficient passive postural balance in the body. Significant deficiencies or excesses of postural HRMT may predispose to symptomatic or pathologic musculoskeletal disorders whose mechanisms are currently unexplained

Cardiac sarcomere mechanics in health and disease

The sarcomere is the fundamental structural and functional unit of striated muscle and is directly responsible for most of its mechanical properties. The sarcomere generates active or contractile forces and determines the passive or elastic properties of striated muscle. In the heart, mutations in sarcomeric proteins are responsible for the majority of genetically inherited cardiomyopathies. Here, we review the major determinants of cardiac sarcomere mechanics including the key structural components that contribute to active and passive tension. We dissect the molecular and structural basis of active force generation, including sarcomere composition, structure, activation, and relaxation. We then explore the giant sarcomere-resident protein titin, the major contributor to cardiac passive tension. We discuss sarcomere dynamics exemplified by the regulation of titin-based stiffness and the titin life cycle. Finally, we provide an overview of therapeutic strategies that target the sarcomere to improve cardiac contraction and filling

Sarcomere Dynamics During Contraction-Induced Injury to Permeabilized Single Muscle Fibers.

A contraction-induced injury results when an activated skeletal muscle or a single fiber is stretched in a ‘lengthening contraction’. This type of injury to muscle fibers is a leading cause of the loss of mobility that is associated with old age. Manifestations of the injury revealed by electron microscopy show focally disrupted single sarcomeres, or small groups of sarcomeres, that are surrounded by intact sarcomeres. The purpose of this dissertation was to study the dynamic behavior of sarcomeres during lengthening contractions of single muscle fibers. The working hypothesis was that contraction-induced injury occurs during a lengthening contraction when sarcomeres undergo elongation at varying rates and those lengthened at the highest rates reach longer lengths and are damaged. To test this hypothesis, a laser scanning system was developed that recorded rapidly (500 s−1) the lengths of sarcomeres contained in each of 20 contiguous regions along ~1 mm segments of a fiber. The fiber segments were obtained from soleus muscles of adult male rats and the membranes of the fibers were permeabilized chemically. The experiments performed with the laser scanning apparatus, indicated that: (i) during a lengthening contraction, the regions of fibers that contain the longer sarcomeres at the onset of the lengthening elongate more compared with the regions that contain shorter sarcomeres; (ii) lengthening contractions increased the variability in the lengths of sarcomeres in relaxed fibers; and (iii) during a lengthening contraction, within any given fiber region, the rate of elongation of sarcomeres was stable, but different regions stretched at different rates as the contraction proceeded. The present findings support the hypothesis that, within single fibers, non-uniformities develop in the lengths of sarcomeres that are increased by a lengthening contraction. The finding that all fiber regions elongate at stable rates during lengthening contractions contradicts a previous hypothesis that during lengthening contractions, the longest sarcomeres undergo rapid and sudden elongation to extreme lengths. The conclusion is that during lengthening contractions, the longer sarcomeres in series with shorter sarcomeres undergo elongation at higher velocities and upon return to original length, structural elements within some of these longer sarcomeres are functionally compromised.Ph.D.Biomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/58375/1/appaji_1.pd