Osteoking exerts pro‑osteogenic and anti‑adipogenic effects in promoting bone fracture healing via EGF/EGFR/HDAC1/Wnt/β‑catenin signaling.

Abstract

Bone fractures, as a global public health issue, lead to disability and reduce the quality of life for patients. Chinese patent drug Osteoking has efficacy in bone fracture therapy. However, its therapeutic properties and underlying mechanisms remain unclear. In the present study, a rat model of bone fracture was established to evaluate the pharmacological effects of Osteoking by behavioral feature detection including mechanical pain threshold measurement, inclined plate and hindlimb weight‑bearing test and CatWalk XT gait analysis, as well as X‑ray scanning and micro‑computed tomography 3D reconstruction. Transcriptomics profiling, network analysis and in vivo western blotting, immunohistochemistry and immunofluorescence assessment were performed to determine the potential targets of Osteoking in promoting bone fracture healing. Osteoking effectively shortened the fracture healing time primarily by accelerating the process of endochondral ossification, decreasing the number of osteoclasts, increasing the levels of bone growth factors and bone formation biomarkers, and decreasing the level of bone resorption biomarkers. Following construction and analysis of the disease gene‑drug target network, it was hypothesized that EGF‑EGFR‑histone deacetylase 1 (HDAC1)‑Wnt/β‑catenin axis‑mediated adipogenesis‑angiogenesis‑osteogenesis crosstalk may be a candidate target of Osteoking in bone fracture. Osteoking significantly decreased expression levels of EGF, phosphorylated‑EGFR and HDAC1 protein and activated Wnt/β‑catenin signaling, which subsequently elevated the expression of VEGFA, Osterix (OSX) and CD31 proteins, increased the RUNX2/PPARγ ratio, decreased the receptor activator of nuclear factor κB ligand/osteoprotegerin ratio and reduced the serum levels of total cholesterol (TC), low‑density lipoprotein cholesterol (LDL‑C) and high‑density lipoprotein cholesterol (HDL‑C). There was a negative association between VEGFA, OSX, TC and LDL‑C levels. In conclusion, Osteoking may effectively reverse the disturbance of adipogenesis‑angiogenesis‑osteogenesis homeostasis and promote the fracture healing by regulating the EGF‑EGFR‑HDAC1‑Wnt/β‑catenin axis. These findings may offer guidance for the clinical application of Osteoking in bone fracture therapy.