Short‐term hindlimb immobilization induces rapid loss of bone strength in a mouse model of disuse atrophy

Abstract

Muscle atrophy and bone loss are known to occur in settings of disuse such as prolonged bedrest, spinal cord injury, or exposure to microgravity with spaceflight. It is also well‐established that crosstalk exists between muscle and bone mediated by myokines such as myostatin and irisin, and that changes in these muscle‐derived factors with exercise, aging, or muscle atrophy may contribute to changes in bone formation and resorption. Muscle loss in many cases often precedes bone loss, and can be more rapid relative to detectable changes in bone mass or density. We investigated short‐term (2 weeks) changes in muscle and bone with disuse atrophy utilizing a single hindlimb immobilization model to identify associated between changes in muscle mass and bone strength. Twelve‐month C57/BL6 mice (8 male, 4 female) were subjected to two weeks of hindlimb (left hindlimb) immobilization, using a casting technique that involved wrapping surgical tape from the pad of the foot to the proximal thigh with a final layer of vetwrap covering the surgical tape. Mice were euthanized following IACUC‐approved procedures, and quadriceps muscles weighed and fiber size measured histologically. Femora were snap frozen in liquid nitrogen and later rehydrated in PBS for 1 hour prior to mechanical testing, after which they were loaded to failure in 3‐point bending (10 mm span length, 10 mm/min) on a Bose ELF mechanical testing system. Results indicate that quadriceps mass and fiber size declined significantly in immobilized (left) hindlimb compared to the non‐immobilized hindlimb, decreasing approximately ~20% and 30%, respectively over the two‐week treatment period. Ultimate force of the femur also declined significantly in the mice over this timeframe, decreasing up to 20% in the male mice. Regression analyses demonstrate that quadriceps mass and fiber size were each significantly correlated with femur ultimate force (P=.01), and that correlation coefficients between muscle mass and fiber size (r=.48) were similar to correlations between muscle mass and femur ultimate force (r=.47) and muscle fiber size and femur ultimate force (r=.47). These findings suggest that even in the setting of short‐term disuse declines in muscle mass and bone strength are tightly coupled, and that therapeutic interventions to mitigate loss of muscle and bone should be considered to reduce fracture risk.