Low-Magnitude Mechanical Signals Combined with Zoledronic Acid Reduce Musculoskeletal Weakness and Adiposity in Estrogen-Deprived Mice

AUTHORS

Gabriel M. Pagnotti, Trupti Trivedi, Laura E. Wright, Sutha K. John, Sreemala Murthy, Ryan R. Pattyn, Monte S. Willis, Yun She, Sukanya Suresh, William R. Thompson, Clinton T. Rubin, Khalid S. Mohammad, Theresa A. Guise

ABSTRACT

Combination treatment of Low-Intensity Vibration (LIV) with zoledronic acid (ZA) was hypothesized to preserve bone mass and muscle strength while reducing adipose tissue accrual associated with complete estrogen (E2)-deprivation in young and skeletally mature mice. Complete E2-deprivation (surgical-ovariectomy (OVX) and daily injection of aromatase inhibitor (AI) letrozole) were performed on 8-week-old C57BL/6 female mice for 4 weeks following commencement of LIV administration or control (no LIV), for 28 weeks. Additionally, 16-week-old C57BL/6 female E2-deprived mice were administered ±LIV twice daily and supplemented with ±ZA (2.5 ng/kg/week). By week 28, lean tissue mass quantified by dual-energy X-ray absorptiometry was increased in younger OVX/AI+LIV(y) mice, with increased myofiber cross-sectional area of quadratus femorii. Grip strength was greater in OVX/AI+LIV(y) mice than OVX/AI(y) mice. Fat mass remained lower in OVX/AI+LIV(y) mice throughout the experiment compared with OVX/AI(y) mice. OVX/AI+LIV(y) mice exhibited increased glucose tolerance and reduced leptin and free fatty acids than OVX/AI(y) mice. Trabecular bone volume fraction and connectivity density increased in the vertebrae of OVX/AI+LIV(y) mice compared to OVX/AI(y) mice; however, this effect was attenuated in the older cohort of E2-deprived mice, specifically in OVX/AI+ZA mice, requiring combined LIV with ZA to increase trabecular bone volume and strength. Similar improvements in cortical bone thickness and cross-sectional area of the femoral mid-diaphysis were observed in OVX/AI+LIV+ZA mice, resulting in greater fracture resistance. Our findings demonstrate that the combination of mechanical signals in the form of LIV and anti-resorptive therapy via ZA improve vertebral trabecular bone and femoral cortical bone, increase lean mass, and reduce adiposity in mice undergoing complete E2-deprivation.

One Sentence Summary: Low-magnitude mechanical signals with zoledronic acid suppressed bone and muscle loss and adiposity in mice undergoing complete estrogen deprivation.

Translational Relevance Postmenopausal patients with estrogen receptor-positive breast cancer treated with aromatase inhibitors to reduce tumor progression experience deleterious effects to bone and muscle subsequently develop muscle weakness, bone fragility, and adipose tissue accrual. Bisphosphonates (i.e., zoledronic acid) prescribed to inhibit osteoclast-mediated bone resorption are effective in preventing bone loss but may not address the non-skeletal effects of muscle weakness and fat accumulation that contribute to patient morbidity. Mechanical signals, typically delivered to the musculoskeletal system during exercise/physical activity, are integral for maintaining bone and muscle health; however, patients undergoing treatments for breast cancer often experience decreased physical activity which further accelerates musculoskeletal degeneration. Low-magnitude mechanical signals, in the form of low-intensity vibrations, generate dynamic loading forces similar to those derived from skeletal muscle contractility. As an adjuvant to existing treatment strategies, low-intensity vibrations may preserve or rescue diminished bone and muscle degraded by breast cancer treatment.

Inhibition of MicroRNA-122-5p Relieves Myocardial Ischemia-Reperfusion Injury via SOCS1

AUTHORS

Jun Zhang , Li Fu , Jing Zhang , Bo Zhou , Yanrong Tang , Zhenzhen Zhang , Tongqing Gu

ABSTRACT

Objective 

Evidence has shown that microRNA (miR)-122–5p is a diagnostic biomarker of acute myocardial infarction. Here, we aimed to uncover the functions of miR-122–5p in the pathological process of myocardial ischemia-reperfusion injury (MI/RI).

Methods 

An MI/RI model was established by left anterior descending coronary artery ligation in mice. The levels of miR-122–5p, suppressor of cytokine signaling-1 (SOCS1), phosphorylation of Janus kinase 2 (p-JAK2), and signal transducers and activators of transcription (p-STAT3) in the myocardial tissues of mice were measured. Downregulated miR-122–5p or upregulated SOCS1 recombinant adenovirus vectors were injected into mice before MI/RI modeling. The cardiac function, inflammatory response, myocardial infarction area, pathological damage, and cardiomyocyte apoptosis in the myocardial tissues of mice were evaluated. Cardiomyocytes were subjected to hypoxia/reoxygenation (H/R) injury and cardiomyocyte biological function was tested upon transfection of miR-122–5p inhibitor. The target relation between miR-122–5p and SOCS1 was evaluated.

Results 

miR-122–5p expression and p-JAK2 and p-STAT3 expression were high, and SOCS1 expression was low in the myocardial tissues of MI/RI mice. Decreasing miR-122–5p or increasing SOCS1 expression inactivated the JAK2/STAT3 pathway to alleviate MI/RI by improving cardiac function and reducing inflammatory reaction, myocardial infarction area, pathological damage, and cardiomyocyte apoptosis in mice. Silencing of SOCS1 reversed depleted miR-122–5p-induced cardioprotection for MI/RI mice. In vitro experiments revealed that the downregulation of miR-122–5p induced proliferative, migratory, and invasive capabilities of H/R cardiomyocytes while inhibiting apoptosis. Mechanically, SOCS1 was a target gene of miR-122–5p.

Conclusion 

Our study summarizes that inhibition of miR-122–5p induces SOCS1 expression, thereby relieving MI/RI in mice.

Development of Nystagmus With the Absence of MYOD Expression in the Extraocular Muscles

AUTHORS

Laura L. Johnson; Rachel B. Kueppers; Erin Y. Shen; Jolene C. Rudell; Linda K. McLoon

ABSTRACT

Purpose: Myoblast determination protein 1 (MYOD) is a critical myogenic regulatory factor in muscle development, differentiation, myofiber repair, and regeneration. As the extraocular muscles significantly remodel their myofibers throughout life compared with limb skeletal muscles, we hypothesized that the absence of MYOD would result in their abnormal structure and function. To assess structural and functional changes in the extraocular muscles in MyoD−/− mice, fiber size and number and optokinetic nystagmus reflex (OKN) responses were examined.

Methods: OKN was measured in MyoD−/− mice and littermate wild-type controls at 3, 6, and 12 months. The extraocular muscles were examined histologically for changes in mean myofiber cross-sectional area, total myofiber number, and nuclei immunostained for PAX7 and PITX2, markers of myogenic precursor cells.

Results: The MyoD−/− mice developed nystagmus, with both jerk and pendular waveforms, in the absence and in the presence of moving visual stimulation. At 12 months, there were significant losses in mean myofiber cross-sectional area and in total number of orbital layer fibers in all rectus muscles, as well as in global layer fibers in the superior and inferior rectus muscles. Haploinsufficient mice showed abnormal OKN responses. PITX2-positive cell entry into myofibers of the MyoD−/− mice was significantly reduced.

Conclusions: This study is the first demonstration of the development of nystagmus in the constitutive absence of expression of the muscle-specific transcription factor MYOD. We hypothesize that myofiber loss over time may alter anterograde and/or retrograde communication between the motor nerves and extraocular muscles that are critical for maintaining normalcy of extraocular muscle function.

Adipogenic Differentiation Alters Properties of Vascularized Tissue-Engineered Skeletal Muscle

AUTHORS

Francisca M. Acosta, Kennedy K. Howland, Katerina Stojkova, Elizabeth Hernandez, Eric M. Brey, and Christopher R. Rathbone

ABSTRACT

Advances in the engineering of comprehensive skeletal muscle models in vitro will improve drug screening platforms and can lead to better therapeutic approaches for the treatment of skeletal muscle injuries. To this end, a vascularized tissue-engineered skeletal muscle (TE-SkM) model that includes adipocytes was developed to better emulate the intramuscular adipose tissue that is observed in skeletal muscles of patients with diseases such as diabetes. Muscle precursor cells cultured with and without microvessels derived from adipose tissue (microvascular fragments) were used to generate TE-SkM constructs, with and without a microvasculature, respectively. TE-SkM constructs were treated with adipogenic induction media to induce varying levels of adipogenesis. With a delayed addition of induction media to allow for angiogenesis, a robust microvasculature in conjunction with an increased content of adipocytes was achieved. The augmentation of vascularized TE-SkM constructs with adipocytes caused a reduction in maturation (compaction), mechanical integrity (Young's modulus), and myotube and vessel alignment. An increase in basal glucose uptake was observed in both levels of adipogenic induction, and a diminished insulin-stimulated glucose uptake was associated with the higher level of adipogenic differentiation and the greater number of adipocytes.

Key indicators of repetitive overuse-induced neuromuscular inflammation and fibrosis are prevented by manual therapy in a rat model

AUTHORS

Mary F. Barbe, Michele Y. Harris, Geneva E. Cruz, Mamta Amin, Nathan M. Billett, Jocelynne T. Dorotan, Emily P. Day, Seung Y. Kim & Geoffrey M. Bove

ABSTRACT

Background

We examined the effectiveness of a manual therapy consisting of forearm skin rolling, muscle mobilization, and upper extremity traction as a preventive treatment for rats performing an intensive lever-pulling task. We hypothesized that this treatment would reduce task-induced neuromuscular and tendon inflammation, fibrosis, and sensorimotor declines.

Methods

Sprague-Dawley rats performed a reaching and lever pulling task for a food reward, 2 h/day, 3 days/week, for 12 weeks, while simultaneously receiving the manual therapy treatment 3 times per week for 12 weeks to either the task-involved upper extremities (TASK-Tx), or the lower extremities as an active control group (TASK-Ac). Results were compared to similarly treated control rats (C-Tx and C-Ac).

Results

Median nerves and forearm flexor muscles and tendons of TASK-Ac rats showed higher numbers of inflammatory CD68+ and fibrogenic CD206+ macrophages, particularly in epineurium, endomysium and epitendons than TASK-Tx rats. CD68+ and CD206+ macrophages numbers in TASK-Tx rats were comparable to the non-task control groups. TASK-Ac rats had more extraneural fibrosis in median nerves, pro-collagen type I levels and immunoexpression in flexor digitorum muscles, and fibrogenic changes in flexor digitorum epitendons, than TASK-Tx rats (which showed comparable responses as control groups). TASK-Ac rats showed cold temperature, lower reflexive grip strength, and task avoidance, responses not seen in TASK-Tx rats (which showed comparable responses as the control groups).

Conclusions

Manual therapy of forelimbs involved in performing the reaching and grasping task prevented the development of inflammatory and fibrogenic changes in forearm nerves, muscle, and tendons, and sensorimotor declines.