The overall goals of the four studies presented herein are 1) to identify how the stiffnesses that govern the mechanical behavior of the muscle-tendon unit (MTU) during locomotion shift during aging and 2) to investigate how these stiffness changes arise at the cellular level and how these cellular changes can be affected by crosstalk between muscle and bone. These studies were performed using computational and experimental models of vertical hopping, in conjunction with in vitro studies.

In Chapter 1, the current state of the literature describing the stiffness changes attributed to aging in bone, muscle, and tendon is addressed. In Chapter 2, we used a previously described Hill-type model to sweep a 2D parameter space of muscle and tendon stiffnesses centered on a “typical” stiffness for a healthy young human. Results from the study suggest that tendon stiffness governs more of the function of the entire muscle-tendon unit (MTU) than does muscle stiffness. We hypothesize that aging may induce the observed MTU changes seen in humans in one of two ways:​ (1) stiffer muscle and tendon, or (2) stiffer muscle but more compliant tendon.

Studies investigating the stiffness changes of muscle-tendon units across age have demonstrated conflicting findings. Therefore, in Chapter 3, we replaced the modeled MTU with a biological one from either a young or old FN344 x BN1 aging rat hybrid and simulated the inertial environments from Chapter 2. We characterized the stiffness utilizing isometric testing, determined the resonant frequency of the MTU, and drove muscle contraction by direct nerve stimulation across a range of frequencies centered on the resonant frequency. We found that, while neither overall MTU stiffness nor muscle and tendon stiffness was not significantly different between young and old rats, older MTUs tended to have increased muscle stiffness and decreased tendon stiffness. This trade off led to small changes in force, length, velocity, and power output of the muscle. This concludes that the tradeoff of stiffnesses in the MTU provide a robustness of the system to function changes.

While higher-level structural changes are often reported in muscle functional studies, how these changes arise at the cellular level are rarely investigated. In Chapter 4, we optimized a cellular co-culture system to enable experiments that examine the age-related changes in the growth and function of muscle myoblasts and bone osteoblasts. We selected bone due to its proximity to muscle and role in muscle growth and repair. We used a Brown Norway rat model and isolated cells from hindlimb muscles and bones. We tested several growth conditions to determine the one best for promoting both muscle and bone cell growth. Our results demonstrated that, while cell plate coating did not alter cell growth, the culture media altered how the cells grew and differentiated, with muscle and bone cells both preferring their own cellspecific media.

We employed the optimized conditions from Chapter 4 to perform an aging muscle and bone cell co-culture study in Chapter 5, using cells isolated from young and old rat hindlimbs contralateral to the ones used in the muscle function studies in Chapter 3. By characterizing the proliferation, differentiation, and crosstalk between muscle and bone cells, we attempt to determine the source of stiffness changes in aged MTUs. We hypothesized that collagen deposition is altered in old vs. young muscle as a result of insulin-like growth factor 1 (IGF-1) changes that may contribute to the observed aging stiffness changes. However, the results showed little difference in collagen deposition between young and old muscle cells across age. The outcome of this work provides a framework for more cross-disciplinary studies leading to designs of new clinical interventions.