It is widely accepted that mechanical stress is an important factor in bone associated cell differentiation, including that of mesenchymal stem cells, osteoblasts and osteocytes. The present study aimed to determine the effect of mechanical cyclic compressive load on osteoblast differentiation, and whether this was associated with activation of the wingless‑type (Wnt)/β-catenin signaling pathway. Using a 3D scaffold model, MC3T3‑E1 cells were exposed to cyclic compressive loading via the Flexcell‑5000C™ Compression system. Sinusoidal wave magnitudes of 0.33, 0.5 and 1 MPa were applied for 4, 6 and 8 h, at 1 Hz frequency. Expression levels of genes associated with osteoblast differentiation were enhanced following compression, including alkaline phosphatase, osteocalcin, runt‑related transcription factor 2 and osterix. Optimal compression was observed using a magnitude of 0.5 MPa for 6 h, whereas a magnitude of 1 MPa had no effect on osteoblast differentiation, and had a negative effect when applied for prolonged time periods. Compressive loading additionally enhanced the mRNA expression levels of the Wnt/β‑catenin signaling pathway component, low density lipoprotein receptor‑related protein 5, and the protein expression levels of Wnt1, disheveled segment polarity protein‑2 (DVL2) and β-catenin. By contrast, mRNA expression levels of sclerostin and the inactive form of β-catenin (phosphorylated at Ser33/37/Thr41) were reduced following compressive loading. Following compressive loading of cells, dickkopf-related protein 1 (DKK‑1), an inhibitor of the Wnt signaling pathway, increased protein expression levels of the inactive form of the Wnt‑associated protein, phosphorylated‑β‑catenin, compared with compression alone. However, DVL2 and Wnt1 protein expression levels were unaffected, suggesting that the loading‑induced activation of Wnt/β‑catenin signaling decreased however, it was not prevented by DKK‑1 treatment. In conclusion, the present study demonstrated that cyclic compressive load promoted osteoblast differentiation and may be dependent on the Wnt/β-catenin signaling pathway in regard to magnitude and duration.