Fatty infiltration of gastrocnemius-soleus muscle complex: Considerations for myosteatosis rehabilitation.

Although previous studies have reported fatty infiltration of the gastrocnemius-soleus complex, little is known about the volumetric distribution and patterns of fatty infiltration. The purpose of this anatomical study was to document and quantify the frequency, distribution, and pattern of fatty infiltration of the gastrocnemius-soleus complex. One hundred formalin-embalmed specimens (mean age 78.1 ± 12.3 years; 48F/52M) were serially dissected to document the frequency, distribution, and pattern of fatty infiltration in the medial and lateral heads of gastrocnemius and soleus muscles. Fatty infiltration was found in 23% of specimens, 13 unilaterally (8F/5M) and 10 (5M/5F) bilaterally. The fatty infiltration process was observed to begin medially from the medial aspect of the medial head of gastrocnemius and medial margin of soleus and then progressed laterally throughout the medial head of gastrocnemius and the marginal, anterior, and posterior soleus. The lateral head of gastrocnemius remained primarily muscular in all specimens. Microscopically, the pattern of infiltration was demonstrated as intramuscular with intact aponeuroses, and septa. The remaining endo-, peri-, and epimysium preserved the overall contour of the gastrocnemius-soleus complex, even in cases of significant fatty replacement. Since the external contour of the calf is preserved, the presence of fatty infiltration may be underdiagnosed in the clinic without imaging. Myosteatosis is associated with gait and balance challenges in the elderly, which can impact quality of life and result in increased risk of falling. The findings of the study have implications in the rehabilitation management of elderly patients with sarcopenia and myosteatosis.

Three-dimensional morphometric analysis of cranial sutures - A novel approach to quantitative analysis.

Objective: Differences in complexity of cranial suture forms on the endocranial (i.e., deep) and ectocranial (i.e., superficial) skull surfaces have been noted in the literature, indicating through thickness three-dimensional (3D) suture variability depending on the chosen section and necessity for considering the complete 3D structure in many cases. This study aims to evaluate the variability of suture morphology through the skull thickness using a rat model, and to provide more robust metrics and methodologies to analyze suture morphology. Design: X-ray micro-computed tomographic (µCT) imaging methods were utilized in order to provide internal structure information. Methods were developed to isolate and analyze sutures widths and linear interdigitation index (LII) values on each adjacent offset transverse plane of the µCT datasets. LII was defined as the curved path length of the suture divided by the linear length between the ends of the region of interest. Scans were obtained on 15 female rats at ages of 16, 20, and 24 weeks (n = 5/age). Samples were imaged at 18 µm resolutions with 90 kV source voltage, 278 µA source amperage, and 0.7° increments. Suture widths and LII values were compared using a Kruskal-Wallis test. Results: 3D variability in local suture widths within individuals, as well as through thickness variabilities in planar widths and LII was observed. Kruskal-Wallis tests for bulk through thickness averaged suture widths and LII were found to be statistically insignificant, despite clear geometric differences through suture thicknesses. Conclusion: Although the bulk morphometric variability between age groups was found to be statistically insignificant, the 3D variability within individuals point to the importance of analyzing suture form using 3D metrics when studying suture development, response to functional activity, or morphometry in general.

A Preliminary Step Towards a Physical Surrogate of the Human Calvarium to Model Fracture.

A surrogate model of the human calvarium can be used to assess skull-fracture-related head injuries without continuously requiring post-mortem human skulls. Skull simulants developed in the literature often require sophisticated manufacturing procedures and/or materials not always practical when factoring in time or expense considerations. This study's objective was to fabricate three exploratory surrogate models (1. pure epoxy prototype, 2. epoxy-chalk mix prototype, and 3. epoxy-chalk three-layered prototype) of the calvarium to mimic the calvarium's mechanical response at fracture using readily available and cost-effective materials, specifically epoxy and chalk. The surrogates and calvaria were subject to quasi-static and dynamic impact 4-point bending and their mechanical responses were compared statistically. Under quasi-static loading, all three surrogates showed a considerable number of differences in mechanical response variables to calvaria that was deemed significant (p < 0.05). Under dynamic impact loading, there was no sufficient evidence to reject that the average mechanical response variables were equal between the epoxy-chalk three-layered prototype and calvaria (p > 0.05). This included force and bending moment at fracture, tensile strain at fracture, tensile and compressive stress at fracture, tensile modulus, and tensile strain rate. Overall, our study illustrates two main remarks: (1) the three exploratory surrogate models are potential candidates for mimicking the mechanical response of the calvarium at fracture during impact loading and (2) employing epoxy and chalk, which are readily available and cost-effective has the potential to mimic the mechanical response of calvaria in impact loading.

The effect of morphometric and geometric indices of the human calvarium on mechanical response.

BACKGROUND: When developing a surrogate model of the human skull, there is a multitude of morphometric and geometric properties to consider when constructing the model. To simplify this approach, it is important to identify only the properties that have a significant influence on the mechanical response of the skull. The objective of this study was to identify which morphometric and geometric properties of the calvarium were significant predictors of mechanical response. METHODS: Calvarium specimens (N = 24) were micro-computed tomography scanned to determine morphometric and geometric properties. The specimens were assumed to be Euler-Bernoulli beams and were subject to 4-point quasi-static bending to determine mechanical response. Univariate linear regressions were performed whereby the morphometric and geometric properties were independent or predictor variables and the mechanical responses were dependent or outcome variables. FINDINGS: Nine significant linear regression models were established (p < 0.05). In the diploë, trabecular bone pattern factor was a significant predictor of force and bending moment at fracture. The inner cortical table had more significant predictors (thickness, tissue mineral density, and porosity) of mechanical response compared to the outer cortical table and diploë. INTERPRETATION: Morphometric and geometric properties had a key influence on the calvarium's biomechanics. Trabecular bone pattern factor and the morphometry and geometry of the cortical tables must be considered when evaluating the mechanical response of the calvarium. These properties can aid the design of surrogate models of the skull that seek to mimic its mechanical response for head impact simulation.

A Direct Comparison of Targeted Muscle Reinnervation and Regenerative Peripheral Nerve Interfaces to Prevent Neuroma Pain.

BACKGROUND AND OBJECTIVES: Targeted muscle reinnervation (TMR) and regenerative peripheral nerve interface (RPNI) surgeries manage neuroma pain; however, there remains considerable discord regarding the best treatment strategy. We provide a direct comparison of TMR and RPNI surgery using a rodent model for the treatment of neuroma pain. METHODS: The tibial nerve of 36 Fischer rats was transected and secured to the dermis to promote neuroma formation. Pain was assessed using mechanical stimulation at the neuroma site (direct pain) and von Frey analysis at the footpad (to assess tactile allodynia from collateral innervation). Once painful neuromas were detected 6 weeks later, animals were randomized to experimental groups: (a) TMR to the motor branch to biceps femoris, (b) RPNI with an extensor digitorum longus graft, (c) neuroma excision, and (d) neuroma in situ. The TMR/RPNIs were harvested to confirm muscle reinnervation, and the sensory ganglia and nerves were harvested to assess markers of regeneration, pain, and inflammation. RESULTS: Ten weeks post-TMR/RPNI surgery, animals had decreased pain scores compared with controls ( P < .001) and they both demonstrated neuromuscular junction reinnervation. Compared with neuroma controls, immunohistochemistry showed that sensory neuronal cell bodies of TMR and RPNI showed a decrease in regeneration markers phosphorylated cyclic AMP receptor binding protein and activation transcription factor 3 and pain markers transient receptor potential vanilloid 1 and neuropeptide Y ( P < .05). The nerve and dorsal root ganglion maintained elevated Iba-1 expression in all cohorts. CONCLUSION: RPNI and TMR improved pain scores after neuroma resection suggesting both may be clinically feasible techniques for improving outcomes for patients with nerve injuries or those undergoing amputation.

The Mechanical Characterization and Comparions of Male and Female Calvaria Under Four-Point Bending Impacts.

The circumstances in which we mechanically test and critically assess human calvarium tissue would find relevance under conditions encompassing real-world head impacts. These conditions include, among other variables, impact velocities, and strain rates. Compared to quasi-static loading on calvaria, there is less reporting on the impact loading of the calvaria and consequently, there are relatively fewer mechanical properties on calvaria at relevant impact loading rates available in the literature. The purpose of this work was to report on the mechanical response of 23 human calvarium specimens subjected to dynamic four-point bending impacts. Impacts were performed using a custom-built four-point impact apparatus at impact velocities of 0.86-0.89 m/s resulting in surface strain rates of 2-3/s-representative of strain rates observed in vehicle collisions and blunt impacts. The study revealed comparable effective bending moduli (11-15 GPa) to the limited work reported on the impact mechanics of calvaria in the literature, however, fracture bending stress (10-47 MPa) was relatively less. As expected, surface strains at fracture (0.21-0.25%) were less compared to studies that performed quasi-static bending. Moreover, the study revealed no significant differences in mechanical response between male and female calvaria. The findings presented in this work are relevant to many areas including validating surrogate skull fracture models in silico or laboratory during impact and optimizing protective devices used by civilians to reduce the risk of a serious head injury.

Muscle attachment sites and behavioral reconstruction: An experimental test of muscle-bone structural response to habitual activity.

OBJECTIVE: Behavioral reconstruction from muscle attachment sites (entheses) is a common practice in anthropology. However, experimental evidence provides mixed support for the assumed association between enthesis size and shape with changes in habitual activity. In this study, a laboratory mouse model was used to experimentally test whether activity level and type alters muscle architecture and the underlying bone cross-sectional geometry of entheses in order to assess the underlying assumption that behavioral changes lead to quantifiable differences in both muscle and enthesis morphology. MATERIALS AND METHODS: Female wild-type mice were separated into one control group and two experimentally increased activity groups (running, climbing) over an 11-week study period. At the start of the experiment, half of the mice were 4 weeks and half were 7 weeks of age. The postmortem deltoideus and biceps brachii muscles were measured for potential force production (physiological cross-sectional area) and potential muscle excursion (fiber length). Bone cross-sectional geometry variables were measured from microCT scans of the humerus and radius at the enthesis and non-enthesis regions of interest across activity groups. RESULTS: Activity level and type altered potential force production and potential muscle excursion of both muscles in the younger cohort. We observed differences in cortical bone geometry in both the humerus enthesis and radius non-enthesis region driven exclusively among the younger wheel-running mice. DISCUSSION: These results indicate that in addition to muscle architectural changes, bone structural properties at the enthesis do show an adaptive response to increased activity, such as running but only during earlier development. However, further research is required in order to apply these findings to the reconstruction of living behavior from anthropological specimens.

Recovering the regenerative potential in chronically injured nerves by using conditioning electrical stimulation.

OBJECTIVE: Chronically injured nerves pose a significant clinical challenge despite surgical management. There is no clinically feasible perioperative technique to upregulate a proregenerative environment in a chronic nerve injury. Conditioning electrical stimulation (CES) significantly improves sensorimotor recovery following acute nerve injury to the tibial and common fibular nerves. The authors' objective was to determine if CES could foster a proregenerative environment following chronically injured nerve reconstruction. METHODS: The tibial nerve of 60 Sprague Dawley rats was cut, and the proximal ends were inserted into the hamstring muscles to prevent spontaneous reinnervation. Eleven weeks postinjury, these chronically injured animals were randomized, and half were treated with CES proximal to the tibial nerve cut site. Three days later, 24 animals were killed to evaluate the effects of CES on the expression of regeneration-associated genes at the cell body (n = 18) and Schwann cell proliferation (n = 6). In the remaining animals, the tibial nerve defect was reconstructed using a 10-mm isograft. Length of nerve regeneration was assessed 3 weeks postgrafting (n = 16), and functional recovery was evaluated weekly between 7 and 19 weeks of regeneration (n = 20). RESULTS: Three weeks after nerve isograft surgery, tibial nerves treated with CES prior to grafting had a significantly longer length of nerve regeneration (p < 0.01). Von Frey analysis identified improved sensory recovery among animals treated with CES (p < 0.01). Motor reinnervation, assessed by kinetics, kinematics, and skilled motor tasks, showed significant recovery (p < 0.05 to p < 0.001). These findings were supported by immunohistochemical quantification of motor endplate reinnervation (p < 0.05). Mechanisms to support the role of CES in reinvigorating the regenerative response were assessed, and it was demonstrated that CES increased the proliferation of Schwann cells in chronically injured nerves (p < 0.05). Furthermore, CES upregulated regeneration-associated gene expression to increase growth-associated protein-43 (GAP-43), phosphorylated cAMP response element binding protein (pCREB) at the neuronal cell bodies, and upregulated glial fibrillary acidic protein expression in the surrounding satellite glial cells (p < 0.05 to p < 0.001). CONCLUSIONS: Regeneration following chronic axotomy is impaired due to downregulation of the proregenerative environment generated following nerve injury. CES delivered to a chronically injured nerve influences the cell body and the nerve to re-upregulate an environment that accelerates axon regeneration, resulting in significant improvements in sensory and motor functional recovery. Percutaneous CES may be a preoperative strategy to significantly improve outcomes for patients undergoing delayed nerve reconstruction.

Cortical and trabecular morphometric properties of the human calvarium.

There is currently a gap in the literature that quantitatively describes the complex bone microarchitecture within the diploë (trabecular bone) and cortical layers of the human calvarium. The purpose of this study was to determine the morphometric properties of the diploë and cortical tables of the human calvarium in which key interacting factors of sex, location on the calvarium, and layers of the sandwich structure were considered. Micro-computed tomography (micro-CT) was utilized to capture images at 18 µm resolution of male (n = 26) and female (n = 24) embalmed calvarium specimens in the frontal and parietal regions (N = 50). All images were post-processed and analyzed using vendor bundled CT-Analyzer software to determine the morphometric properties of the diploë and cortical layers. A two-way mixed (repeated measures) analysis of variance (ANOVA) was used to determine diploë morphometric properties accounting for factors of sex and location. A three-way mixed ANOVA was performed to determine cortical morphometric properties accounting for factors of cortical layer (inner and outer table), sex, and location. The study revealed no two-way interaction effects between sex and location on the diploë morphometry except for fractal dimension. Trabecular thickness and separation in the diploë were significantly greater in the male specimens; however, females showed a greater number of trabeculae and fractal dimension on average. Parietal specimens revealed a greater porosity, trabecular separation, and deviation from an ideal plate structure, but a lesser number of trabeculae and connectivity compared to the frontal location. Additionally, the study observed a lower density and greater porosity in the inner cortical layer than the outer which may be due to clear distinctions between each layer's physiological environment. The study provides valuable insight into the quantitative morphometry of the calvarium in which finite element modelers of the skull can refer to when designing detailed heterogenous or subject-specific skull models to effectively predict injury. Furthermore, this study contributes towards the recent developments on physical surrogate models of the skull which require approximate measures of calvarium bone architecture in order to effectively fabricate a model and then accurately simulate a traumatic head impact event.

Conditioning Electrical Stimulation Accelerates Regeneration in Nerve Transfers.

OBJECTIVE: Compared to the upper limb, lower limb distal nerve transfer (DNT) outcomes are poor, likely due to the longer length of regeneration required. DNT surgery to treat foot drop entails rerouting a tibial nerve branch to the denervated common fibular nerve stump to reinnervate the tibialis anterior muscle for ankle dorsiflexion. Conditioning electrical stimulation (CES) prior to nerve repair surgery accelerates nerve regeneration and promotes sensorimotor recovery. We hypothesize that CES prior to DNT will promote nerve regeneration to restore ankle dorsiflexion. METHODS: One week following common fibular nerve crush, CES was delivered to the tibial nerve in half the animals, and at 2 weeks, all animals received a DNT. To investigate the effects of CES on nerve regeneration, a series of kinetic, kinematic, skilled locomotion, electrophysiologic, and immunohistochemical outcomes were assessed. The effects of CES on the nerve were investigated. RESULTS: CES-treated animals had significantly accelerated nerve regeneration (p < 0.001), increased walking speed, and improved skilled locomotion. The injured limb had greater vertical peak forces, with improved duty factor, near-complete recovery of braking, propulsive forces, and dorsiflexion (p < 0.01). Reinnervation of the tibialis anterior muscle was confirmed with nerve conduction studies and immunohistochemical analysis of the neuromuscular junction. Immunohistochemistry demonstrated that CES does not induce Wallerian degeneration, nor does it cause macrophage infiltration of the distal tibial nerve. INTERPRETATION: Tibial nerve CES prior to DNT significantly improved functional recovery of the common fibular nerve and its muscle targets without inducing injury to the donor nerve. ANN NEUROL 2020;88:363-374.

Characteristics of Vibration that Alter Cardiovascular Parameters in Mice.

We hypothesized that short-term exposure of mice to vibration within a frequency range thought to be near the resonant frequency range of mouse tissue and at an acceleration of 0 to 1 m/s(2) would alter heart rate (HR) and mean arterial pressure (MAP). We used radiotelemetry to evaluate the cardiovascular response to vibration in C57BL/6 and CD1 male mice exposed to vertical vibration of various frequencies and accelerations. MAP was consistently increased above baseline values at an acceleration near 1 m/s(2) and a frequency of 90 Hz in both strains, and HR was increased also in C57BL/6 mice. In addition, MAP increased at 80 Hz in individual mice of both strains. When both strains were analyzed together, mean MAP and HR were increased at 90 Hz at 1 m/s(2), and HR was increased at 80 Hz at 1 m/s(2). No consistent change in MAP or HR occurred when mice were exposed to frequencies below 80 Hz or above 90 Hz. The increase in MAP and HR occurred only when the mice had conscious awareness of the vibration, given that these changes did not occur when anesthetized mice were exposed to vibration. Tested vibration acceleration levels lower than 0.75 m/s(2) did not increase MAP or HR at 80 or 90 Hz, suggesting that a relatively high level of vibration is necessary to increase these parameters. These data are important to establish the harmful frequencies and accelerations of environmental vibration that should be minimized or avoided in mouse facilities.

Locomotor activity influences muscle architecture and bone growth but not muscle attachment site morphology.

The ability to make behavioural inferences from skeletal remains is critical to understanding the lifestyles and activities of past human populations and extinct animals. Muscle attachment site (enthesis) morphology has long been assumed to reflect muscle strength and activity during life, but little experimental evidence exists to directly link activity patterns with muscle development and the morphology of their attachments to the skeleton. We used a mouse model to experimentally test how the level and type of activity influences forelimb muscle architecture of spinodeltoideus, acromiodeltoideus, and superficial pectoralis, bone growth rate and gross morphology of their insertion sites. Over an 11-week period, we collected data on activity levels in one control group and two experimental activity groups (running, climbing) of female wild-type mice. Our results show that both activity type and level increased bone growth rates influenced muscle architecture, including differences in potential muscular excursion (fibre length) and potential force production (physiological cross-sectional area). However, despite significant influences on muscle architecture and bone development, activity had no observable effect on enthesis morphology. These results suggest that the gross morphology of entheses is less reliable than internal bone structure for making inferences about an individual's past behaviour.

Vibrating Frequency Thresholds in Mice and Rats: Implications for the Effects of Vibrations on Animal Health.

Vibrations in research facilities can cause complex animal behavioral and physiological responses that can affect animal health and research outcomes. The goal of this study was to determine the range of frequency values, where animals are unable to attenuate vibrations, and therefore may be most susceptible to their effects. Anesthetized and euthanized adult rats and mice were exposed to vibration frequencies over a wide range (0-600 Hz) and at a constant magnitude of 0.3 m/s(2). Euthanized animals were additionally exposed to vibrations at an acceleration of 1 m/s(2). The data showed that at most frequencies rodents were able to attenuate vibration magnitudes, with values for the back-mounted accelerometer being substantially less than that of the table. At frequencies of 41-60 Hz mice did not attenuate vibration magnitude, but instead the magnitude of the table and animal were equal or amplified. Rats experienced the same pattern of non-attenuation between 31 and 50 Hz. Once euthanized, the mice vibrated at a slightly more elevated frequency (up to 100 Hz). Based on these results, it may be prudent that in laboratory settings, vibrations in the ranges reported here should be accounted for as possible contributors to animal stress and/or biomechanical changes.

Life-long caloric restriction does not alter the severity of age-related osteoarthritis.

Chronic adipose tissue inflammation and its associated adipokines have been linked to the development of osteoarthritis (OA). It has been shown that caloric restriction may decrease body mass index and adiposity. The objectives of this study were to investigate the effect of lifelong caloric restriction on bone morphology, joint inflammation, and spontaneously occurring OA development in aged mice. C57BL/NIA mice were fed either a calorie-restricted (CR) or ad libitum (AL) diet starting at 14 weeks of age. All mice were sacrificed at 24 months of age. Adipose tissue and knee joints were then harvested. Bone parameters of the joints were analyzed by micro-CT. OA and joint synovitis were determined using histology and semiquantitative analysis. Lifelong caloric restriction did not alter the severity of OA development in C57BL/NIA aged mice, and there was no difference in the total joint Mankin score between CR and AL groups (p = 0.99). Mice also exhibited similar levels of synovitis (p = 0.54). The bone mineral density of the femur and the tibia was comparable between the groups with a small increase in cancellous bone volume fraction in the lateral femoral condyle of the CR group compared with the AL group. Lifelong caloric restriction did not alter the incidence of OA or joint synovitis in C57BL/NIA mice, indicating that a reduction of caloric intake alone was not sufficient to prevent spontaneous age-related OA. Nonetheless, early initiation of CR continued throughout a life span did not negatively impact bone structural properties.