At present, the identity of the proteins that participate in mechanical stimulation during the fracture healing process is not known. Here, we screened for and identified 299 proteins that are expressed in various stages of mechanically stimulated fracture healing. Among these biomarkers, 54 proteins were differentially expressed during all three recovery time points. Gene ontology (GO) analysis and selected biomarker candidates were used to determine their involvement in biological processes required for mechanically stimulated healing. The levels of cofilin-1 and A2HSG protein were significantly upregulated in the forth week after fracture suggesting that that they play a role in mechanically stimulated fracture healing.