Discrepancy Between Biological Activity and Functional Fracture Healing Following Vitamin K2 Supplementation in an Ovariectomized Rat Model of Osteoporosis

Background: Vitamin K2 (menaquinone) has been studied as a molecule with important effects on bone metabolism and has been proposed as a potential adjuvant in fracture healing, particularly under osteoporotic conditions. However, its functional impact on osteoporotic fracture healing remains largely undefined. The aim of this study was to evaluate the effects of vitamin K2 supplementation, in the form of menaquinone-4 (MK-4) and menaquinone-7 (MK-7), on fracture healing in an ovariectomized rat model of osteoporosis. Methods: Forty Wistar rats were included in this study and allocated to four equal groups: Sham control, ovariectomized control, MK-4, and MK-7. Osteoporosis was induced by bilateral ovariectomy, and 12 weeks after ovariectomy, a femoral fracture was produced and fixed by intramedullary nailing. Starting on postoperative day 2, the MK-4 group received 5 mg/kg/day of MK-4, while the MK-7 group received MK-7 at a dose of 0.05 mg/kg/day. Fracture healing was assessed primarily by biomechanical testing using a three-point bending test and was further analyzed by histological and biochemical parameters, including CTXI, PINP, ucOC, BALP, and ALT. Results: Vitamin K2 supplementation did not improve functional fracture healing. In both treatment groups, fractures showed nonunion-like mechanical behavior, precluding meaningful quantitative biomechanical comparison. Although histological and biochemical findings, particularly in the MK-4 group, showed some degree of biological activity, these changes did not translate into mechanically competent bone union. Both treatment groups showed a tendency toward impaired healing, with progression toward nonunion-like behavior under the present experimental conditions. No significant hepatic toxicity was observed. Conclusions: In this ovariectomized rat femoral fracture model, vitamin K2 supplementation with either MK-4 or MK-7 did not enhance functional fracture healing despite evidence of biological activity of the treatment. These findings suggest a discrepancy between molecular or histological effects and biomechanical outcomes, indicating that, under the conditions tested, vitamin K2 is insufficient to overcome impaired healing in osteoporotic bone and may adversely influence fracture repair under these experimental conditions, although the mechanism remains uncertain.