Introduction: The clinically known importance of patient sex as a major risk factor for compromised bone healing is poorly reflected in animal models. Consequently, the underlying cellular mechanisms remain elusive. Because mesenchymal stem cells (MSCs) are postulated to regulate tissue regeneration and give rise to essential differentiated cell types, they may contribute to sex-specific differences in bone healing outcomes.
Methods: We investigated sex-specific variations in bone healing and associated differences in MSC populations. A 1.5 mm osteotomy gap in the femora of 8 male and 8 female 12-month-old Sprague-Dawley rats was stabilized by an external fixator. Healing was analyzed in terms of biomechanical testing, bridging and callus size over time (radiography at 2, 4, and 6 weeks after surgery), and callus volume and geometry by microCT at final follow-up. MSCs were obtained from bone marrow samples of an age-matched group of 12 animals (6 per gender) and analyzed for numbers of colony-forming units (CFUs) and their capacity to differentiate and proliferate. The proportion of senescent cells was determined by beta-galactosidase staining.
Results: Sex-specific differences were indicated by a compromised mechanical competence of the callus in females compared with males (maximum torque at failure, p=0.028). Throughout the follow-up, the cross-sectional area of callus relative to bone was reduced in females (p< or =0.01), and the bridging of callus was delayed (p(2weeks)=0.041). microCT revealed a reduced callus size (p=0.003), mineralization (p=0.003) and polar moment of inertia (p=0.003) in female animals. The female bone marrow contained significantly fewer MSCs, represented by low CFU numbers in both femora and tibiae (p(femur)=0.017, p(tibia)=0.010). Functional characteristics of male and female MSCs were similar.
Conclusion: Biomechanically compromised and radiographically delayed bone formation were distinctive in female rats. These differences were concomitant with a reduced number of MSCs, which may be causative for the suboptimal bone healing.