Glucocorticoids (GCs) are the most common cause of atraumatic osteonecrosis of the femoral head (ONFH) because their effect compromises the osteogenic capability of bone marrow‑derived mesenchymal stem cells (BMSCs). Valproic acid (VPA) is a widely used anti‑epileptic and anti‑convulsant drug. Previous studies have reported that VPA promotes osteogenic differentiation of MSCs in vitro and osteogenesis in vivo as a histone deacetylase (HDAC) inhibitor. The purpose of the present study was to investigate the efficacy of VPA as a precautionary treatment of ONFH after GC treatment in rats. In vitro, the effect of VPA, dexamethasone or a combination treatment of the two on the proliferation and osteogenic differentiation of human BMSCs was assessed using a Cell Counting Kit‑8 and apoptosis assays, and by measuring the expression of proteins associated with osteogenesis. In vivo, a GC‑induced ONFH model was established in rats and VPA was added during GC treatment to investigate the preventive effect of VPA against ONFH. Rat BMSCs were also extracted to investigate the osteogenic capacity. The results of micro‑computed tomography scanning, angiography of the femoral head and histological and immunohistochemical analyses indicated that 11 of 15 rats induced with methylprednisolone (MP) presented with ONFH, while only 2 of 15 rats treated with a combination of MP and VPA developed ONFH. VPA produced beneficial effects on subchondral bone trabeculae in the femoral head with significant preservation of bone volume and blood supply, as well as improved osteogenic capability of BMSCs compared with those in rats treated with GC alone. In conclusion, VPA attenuated the inhibitory effect of GC on BMSC proliferation and osteogenesis by inhibiting apoptosis and elevating the expression of proteins associated with osteogenesis, which may contribute to the prevention of GC‑induced ONFH in rats.