Impact of Gefitinib on Traumatic Fracture Healing in Rats via the OPG/RANKL/RANK Signaling Pathway

Abstract

This study investigates how gefitinib (Gef), an epidermal growth factor receptor (EGFR) inhibitor, affects traumatic fracture healing in rats, particularly through the OPG/RANKL/RANK signaling pathway, which regulates osteoclast and osteoblast activity. Sixty Sprague-Dawley rats were divided into different treatment groups, including fracture only, fracture plus gefitinib, fracture plus functional OPG antagonist (i.e., recombinant mouse RANKL protein), and combinations of both. After 28 days of treatment, bone healing was evaluated using techniques such as micro- CT, histological staining, and molecular analysis. Additionally, in vitro experiments on bone marrow mesenchymal stem cells (BMSCs) were conducted. The results showed that rats treated with gefitinib had significantly higher bone density and better bone structure compared to the fracture-only group. The effects were diminished when the functional OPG antagonist was added. Histological analysis revealed that gefitinib promoted better callus formation and more organized trabecular bone, effects that were reversed by the functional OPG antagonist. Further analysis indicated that gefitinib inhibited osteoclast activity and enhanced osteoblast function. At the molecular level, gefitinib increased the expression of OPG and bone-related proteins while reducing the levels of RANKL and osteoclast markers. In contrast, the functional OPG antagonist had the opposite effect.In vitro, gefitinib increased osteogenic activity and gene expression in BMSCs, but this was blocked by RANKL. The findings suggest that gefitinib promotes fracture healing by upregulating OPG and suppressing RANKL/RANK signaling, reducing inflammation, and this effect can be completely negated by exogenous RANKL.