Transient muscle paralysis disrupts bone homeostasis by rapid degradation of bone morphology

Abstract

We have previously shown that transient paralysis of murine hindlimb muscles causes profound degradation of both trabecular and cortical bone in the adjacent skeleton within 3 weeks. Morphologically, the acute loss of bone tissue appeared to arise primarily due to osteoclastic bone resorption. Given that the loss of muscle function in this model is transient, we speculated that the stimulus for osteoclastic activation would be rapid and morphologic evidence of bone resorption would appear before 21 days. We therefore utilized high-resolution in vivo serial micro-CT to assess longitudinal alterations in lower hindlimb muscle volume, proximal tibia trabecular, and tibia mid-diaphysis cortical bone morphology in 16-week-old female C57 mice following transient calf paralysis from a single injection of botulinum toxin A (BtA; 2U/100 g body weight). In an acute study, we evaluated muscle and bone alterations at days 0, 3, 5, and 12 following transient calf paralysis. In a chronic study, following day 0 imaging, we assessed the recovery of these tissues following the maximum observed trabecular degradation (day 12) through day 84 post-paralysis. The time course and degree of recovery of muscle, trabecular, and cortical bone varied substantially. Significant atrophy of lower limb muscle was evident by day 5 of paralysis, maximal at day 28 (-34.1+/-0.9%) and partially recovered by day 84. Trabecular degradation within the proximal tibia metaphysis occurred more rapidly, with significant reduction in BV/TV by day 3, maximal loss at day 12 (-76.8+/-2.9%) with only limited recovery by day 84 (-51.7+/-5.1% vs. day 0). Significant cortical bone volume degradation at the tibia mid-diaphysis was first identified at day 12, was maximal at day 28 (-9.6+/-1.2%), but completely recovered by day 84. The timing, magnitude, and morphology of the observed bone erosion induced by transient muscle paralysis were consistent with a rapid recruitment and prolific activation of osteoclastic resorption. In a broader context, understanding how brief paralysis of a single muscle group can precipitate such rapid and profound bone resorption in an adjacent bone is likely to provide new insight into how normal muscle function modulates bone homeostasis.

Fig 1

Acute alterations in proximal tibia metaphysis trabecular morphology following BtA-induced transient muscle paralysis (mean ± S.E. percentage vs. d 0) at d 3, 5 and 12. Representative serial micro-CT images from the proximal tibia metaphysis of a single mouse visually demonstrate trabecular degradation observed within just a few days following induced paralysis. M.V: lower limb muscle volume, BV/TV: trabecular bone volume, Tb.Sp: trabecular spacing, with *: p<0.05 vs. d 0; †: p< 0.05 vs. d 3; ‡: p< 0.05 vs. d 5. Scale bar = 1 mm.

Fig 2

Acute alterations in tibia mid-diaphysis morphology following BtA-induced transient muscle paralysis (mean percentage ± SE vs. d 0) at d 3, 5 and 12. Within this time period, only cortical bone (Ct.V) differences reached statistical significance. Ct.V: cortical bone volume, Ec.V: endocortical volume, with *: p<0.05; †: p< 0.05 vs. d 3.

Fig 3

Alterations in proximal tibia metaphysis trabecular morphology following BtA-induced transient muscle paralysis (mean percentage ± SE vs. d 0) at d 12, 28, 56 and 84. Representative serial micro-CT images from the proximal tibia metaphysis of a single mouse visually demonstrate profound degradation but only partial restoration of trabecular morphology by d 84. M. V: lower limb muscle volume, BV/TV: trabecular bone volume, Tb.Sp: trabecular spacing, with *: p<0.05 vs. d 0; †: p< 0.05 vs. d 12; ‡: p< 0.05 vs. d 28; §: p < 0.05 vs. d 56. Scale bar = 1 mm.

Fig 4

Tissue changes in the tibia mid-diaphysis following BtA-induced transient muscle paralysis (mean percentage ± SE vs. d 0) at 12, 28, 56 and 84 d. The maximal degradation of cortical bone (Ct.V) was observed at 28 d due to expansion of the endocortical volume (Ec.V). By 84 d, however, this catabolic response was completely mitigated. Ct.V: cortical bone volume, Ec.V: endocortical volume, with *: p<0.05; †: p< 0.001 vs. d 0; ‡: p<0.05; § vs. d 28.