Various studies have reported that SMFs with different parameters can affect bone tissue and cells in different ways[11]. In this study, MMF exposure accelerated the repair of microstructure, mechanical properties and morphology during fracture healing. Meanwhile, MMF exposure regulated the distribution of osteoblasts and osteoclast in callus, and influenced the levels the bone metabolic markers in serum. Furthermore, MMF exposure produced a regulatory effect the iron content in the bone callus and the systemic iron metabolism during the fracture healing.
Routine blood parameters can reflect the basic physiological status of organism, such as the level of inflammation[22]. MMF exposure has no significant effect on the content of RBC, HGB, MCH, and PLT during fracture healing. Inflammatory response in the early stages of fractures is beneficial to fracture healing, but dysregulated chronic inflammation is not conducive to fracture healing[23, 24]. WBC is an important indicator for the inflammation, and MMF exposure was decreased the WBC on day 14 postfracture. It is suggested that MMF exposure could alleviate the inflammatory response during fracture healing.
The callus been connection by newly formed bone tissue is the typical physiological feature of fracture healing. In this study, MMF exposure significantly improved BMD and BV/TV, and accelerated the deposition of mineralized bone matrix in callus during fracture healing. In the final stage of fracture healing, the mechanical properties gradually returned with the maturation and order degree of microstructure continues improving[25]. In this study, the results showed that the MMF exposure significantly improved both of structural parameters and material properties, and the improvement effect of mechanical properties such as maximum load exceeds the change ratio of its microstructure. Therefore, orderliness of newly formed bone matrix under MMF exposure is worthy for further research.
The bone metabolism during fracture healing can be divided into two major biological stages: endochondral ossification and coupled remodeling[4]. SMFs exposure could promote the chondrogenic and osteogenic differentiation of mesenchymal stem cells (MSCs)[26–30]. Endochondral ossification involves the cartilage formation and the transition from cartilage callus to hard callus[31]. In this study, the MMF exposure increased the mineralized area of the callus on the 14th and 28th day of fracture healing, accelerating the transition from cartilage callus to hard callus. Meanwhile, the MMF exposure increased the number of osteoblasts at the callus site after 14 days of fracture healing. It is suggested that the static magnetic field promoting fracture healing perhaps related to the regulation of endochondral ossification. In the stage of coupled remodeling, bone matrix is continuously updated through osteoblast mediated bone formation and osteoclast mediated bone absorption[32, 33]. Result showed that, there was no significant change in the number of osteoblasts under MMF exposure, and the concentration of bone formation markers OCN and PINP have been increased on day 28 postfracture. It is suggested that MMF exposure could improving the bone matrix formation ability of osteoblasts.
Osteoclast participate in multiple stages of fracture healing, and is indispensable in the coupled remodeling in the final stage of fracture healing[33]. Meanwhile, Osteoclast mediated bone resorption may have a destructive effect on the formation of mineralized bone matrix. In this study, the TRAP-5b, β-CTx in serum, and the number and area of osteoclast on the surface of trabecular bone in callus were decreased by MMF exposure for 28 days. Therefore, the effect of MMF exposure on the stage of coupled remodeling during fracture healing need further exploration.
Iron metabolism is closely related to bone health[34]. SMFs can affect bone homeostasis by regulating iron metabolism in various bone tissue cells, including osteoblasts and osteoclasts, etc[20, 35]. In this study, the iron content in liver and serum ferritin have been significant decreased by MMF exposure on day 14 and day 28 postfracture. The iron content of fractured tibia in MMF exposure group was significantly decreased on day 28 postfracture. Furthermore, MMF exposure increased the content of hepcidin in serum on day 14 postfracture, a core marker of systemic iron metabolism. It is suggested that MMF exposure promoting fracture healing is related to its regulation of iron metabolism. Changes in systemic iron metabolism may cause symptoms such as anemia[36]. Meanwhile, fractures may cause a large amount of blood loss and anemia in patients, which can lead to various diseases[37]. This study found that MMF exposure had no significant impact on the HGT and MCH, which mean that not exacerbate anemia symptoms caused by fractures.
Previous found that 0.2 T- 0.4 T SMF has no significant effect on the microstructure and mechanical properties of bones in normal mice, while could suppressed ovariectomize (OVX) induced bone loss and promotes the recovery of HLU induced bone loss in mice[15, 16, 38]. In this study, the magnetic field intensity at the location of the fracture site was between 0.05 T and 0.5 T, and increased bone formation and inhibited bone resorption in bone callus. This means that the static magnetic field has a more significant impact on the pathological process of non equilibrium in the skeletal system, but its impact on healthy bones in a state of bone remodeling balance is relatively weak, and the reasons for this difference need further research.
In summary, this study indicated that MMF could promote fracture healing in mice by regulating the process of endochondral osteogenesis and bone remodeling. This provides new insights into the biological mechanism of the effect of MMF on fracture healing and provides a theoretical basis for its application in clinical bone fractures therapy.