12-WEEK PROBIOTIC TREATMENT YIELDS IMPROVED PELVIC BONE MICROSTRUCTURE FOLLOWING FRACTURE IN OSTEOPOROTIC RATS

Osteoporosis causes bones to become thin and brittle, making them more susceptible to fracture. Osteoporotic fractures remain a significant healthcare burden with delayed and/or impaired healing. Current literature has demonstrated an effect of gut microbiota on cytokine pathways, including those involved in bone formation and reabsorption. In this analysis we quantified the impact of daily oral probiotic treatment pre- and post- unilateral pelvic fracture on the microstructural parameters of both the fractured and the contralateral intact hemi-pelvis. Eighteen six-month-old female Sprague Dawley rats were bilaterally ovariectomized and housed for three months to establish the osteoporotic phenotype. Rats were randomly separated into control (PBS was administered for twelve weeks), pre-fracture probiotic treatment (administered for twelve weeks starting six-weeks prior to the creation of the fracture), and post-fracture probiotic treatment (for six weeks following the creation of the fracture) groups. At three months post-ovariectomy, stable unilateral fractures of the pelvis were created via osteotomy of the superior and inferior pubic rami of the left hemi-pelvis using a Gigli saw. All rats were sacrificed at six-weeks post-fracture. Post-sacrifice, pelvi were excised, µCT imaged (35μm isotropic voxel size), and microstructurally analyzed with respect to total volume (TV), bone volume (BV), bone volume/total volume (BV/TV), bone mineral density (BMD), and tissue mineral density (TMD). An analysis of variance (ANOVA) with post-hoc testing (Tukey HSD test) was completed to determine differences between the treatment groups and the control group on the unfractured hemi-pelvis. For data that did not follow a normal distribution, a Kruskal-Wallis test with post-hoc testing (Dunn test) was completed. The fractured and intact hemi-pelvi of sixteen rats were included in this analysis. The intact hemi-pelvis of the pre-fracture probiotic treatment group (n=5) had significantly higher BV (49.1 vs 40.2mm3, P = 0.045), BV/TV (43.2 vs 40.7%, P = 0.043), BMD (646.9 vs 572.5mgHA/cm3, P = 0.034), and TMD (1094.8 vs 1016.5mgHA/cm3, P = 0.031) when compared to the post-fracture probiotic treatment group (n=7). There were no significant differences between pre- or post-fracture probiotic treatment group and the control group (n=4). However, on the fractured hemi-pelvis, when compared to the control group, the pre-fracture treatment group had increased TV (176.8 vs 119.8mm3, P = 0.031) and BV (78.1 vs 49.1mm3, P = 0.020). Daily oral probiotic treatment may result in improvements in bone microstructural parameters in individuals with osteoporosis, however, the effectiveness of this treatment appears to be closely related to treatment duration. Administration of probiotics for twelve weeks significantly increases bone microstructure in both the healing fracture and intact contralateral hemi-pelvis. This highlights that probiotics can alter the immune response that occurs following a fracture. These findings suggest that there exists a critical threshold that must be met in order for gut microbiota to become optimized and produce changes in osteoporotic bone.