THE EFFECT OF ENDOGENOUS OPIOIDS ON FRACTURE HEALING

Participants: T. Cooney, E.P. Frankenburg, E. Green, S.A. Goldstein

Keywords: opioids, fracture repair, animal model

Introduction

Humoral factors, in concert with bone-derived factors, modulate fracture healing. Perturbation in the balance between both sources of regulatory factors may cause an aberrant response. In the case of head or spinal cord injury, humoral mediators may be partly or wholly responsible for ectopic bone formation. Abundant literature suggests an association between neurologic insult and HO formation. Further, the sera from such patients is a potent mitogen for cultured osteoblasts. A potential mediator for these effects is endogenous opioids. Acutely elevated levels of beta endorphin have been detected in the CSF and sera or head-injured patients. Endogenous opioids have also been shown to modulate body growth in rat pups. Identification of proenkaphalin transcripts in preosteoblast cell lines provides further evidence for their involvement in bone formation. As endochondral ossification is a recapitulation of ontogeny, we hypothesized a regulatory role for endogenous opioids in fracture healing. Specifically, we posited that opioids would impair healing. Varied doses of an opioid antagonist would foster exuberant callus formation when given in high dose or impede healing in low doses. The study protocol to test this hypothesis was formalized, submitted, and approved by our institutional animal care and use committee.

Materials and Methods

Forty-two juvenile, male Sprague-Dawley rats were randomized into one of three treatment groups: high dose naltrexone (20 mg/kg, n=15), low dose naltrexone (1 mg/kg n=14), or sham/saline (n=13). Injections were administered subcutaneously 24 hours before fracture and then daily for three weeks thereafter. Closed fractures were produced by the method of Bonnarens and Einhorn under general anesthesia. At two weeks of convalescence, a small subset of animals (n=2/group) were euthanized and the femurs harvested to presumptively assess early healing responses. Consistent with previous studies, the majority of animals were sacrificed at four weeks. Mechanical testing was performed to determine load-to-failure and stiffness parameters. Representative fracture calluses were then subjected to sectioning and staining with Masson’s trichrome and then rated for healing maturity with a published rating scale. Differences in the material strength properties of four-week specimens were subjected to a one-way ANOVA to ascertain statistical significance using 0.05 as the threshold.

Results

Load-to-failure values ranged from 10-15 lbs. at two weeks. By four weeks, failure occurred in the range of 28 to 31 lbs. The latter represented 72% to 80% of contralateral values. Stiffness estimates at two weeks suggested a three-fold difference between saline and naltrexone-treated specimens. High variability and low sample size precludes definitive interpretation of this difference. At four weeks, values were similar, representing 2% to 70% of contralateral values. Treatment effects at four weeks were not significant for either material strength parameter. Histologically based ratings of fracture maturity revealed an advanced stage of healing and failed to disclose differences between treatment groups for either healing time.

This study failed to show conclusive evidence of an impact by naltrexone on the strength of fracture healing in rats. As naltrexone antagonizes opioid effects, these results suggest that endogenous opioids do not exert an influence on more advanced stages of fracture healing. However, presumptive two-week stiffness data are suggestive of a role for early fracture stabilization. This observation may be aligned with predecessor studies that convey a role for endogenous opioids during ontogeny. Alternatively, the developmental window for opioid effects may be confined to the early neonatal period. The juvenile animals used in this study may have been too old to discern any impact. Additional testing would be required to confirm or refute these speculations.