Botox induced muscle paralysis rapidly degrades bone

Abstract

The means by which muscle function modulates bone homeostasis is poorly understood. To begin to address this issue, we have developed a novel murine model of unilateral transient hindlimb muscle paralysis using botulinum toxin A (Botox). Female C57BL/6 mice (16 weeks) received IM injections of either saline or Botox (n = 10 each) in both the quadriceps and calf muscles of the right hindleg. Gait dysfunction was assessed by multi-observer inventory, muscle alterations were determined by wet mass, and bone alterations were assessed by micro-CT imaging at the distal femur, proximal tibia, and tibia mid-diaphysis. Profound degradation of both muscle and bone was observed within 21 days despite significant restoration of weight bearing function by 14 days. The muscle mass of the injected quadriceps and calf muscles was diminished -47.3% and -59.7%, respectively, vs. saline mice (both P < 0.001). The ratio of bone volume to tissue volume (BV/TV) within the distal femoral epiphysis and proximal tibial metaphysis of Botox injected limbs was reduced -43.2% and -54.3%, respectively, while tibia cortical bone volume was reduced -14.6% (all P < 0.001). Comparison of the contralateral non-injected limbs indicated the presence of moderate systemic effects in the model that were most probably associated with diminished activity following muscle paralysis. Taken as a whole, the micro-CT data implied that trabecular and cortical bone loss was primarily achieved by bone resorption. These data confirm the decisive role of neuromuscular function in mediating bone homeostasis and establish a model with unique potential to explore the mechanisms underlying this relation. Given the rapidly expanding use of neuromuscular inhibitors for indications such as pain reduction, these data also raise the critical need to monitor bone loss in these patients.

Fig. 1

Botox and saline group body mass variations. Mean body mass (±SE) in the saline group was significantly elevated following day 0 (*P < 0.05). Body mass transiently decreased by 3 days in Botox treated mice, but returned to initial levels by 21 days (†P < 0.05 vs. day 0). As a result, Botox mice weighed significantly less than saline mice from 1 day through 21 days (‡P < 0.05).

Fig. 2

Gait function was diminished by Botox treatment. All Botox mice demonstrated significantly reduced function compared to saline mice (*P < 0.01). The observed disability due to Botox treatment was maximal by 3 days and was significantly improved by 14 and 21 days (†P < 0.05 vs. 1, 3, and 7 days).

Fig. 3

Micro-CT images (3-D) of the right distal femoral epiphysis, right proximal tibia metaphysis, and right tibia mid-diaphysis from a representative saline injected (top) and Botox injected mouse (bottom). Across groups, trabecular BV/TV (%) was reduced −43.2% and −54.3% at the distal femur and proximal tibia, respectively, and −14.6% at the tibia mid-diaphysis (all P < 0.001). The cortical shell periosteal volume at the distal femur and proximal tibia and the periosteal volume of the tibia mid-diaphysis did not differ between saline and Botox groups.