In order to investigate the effect of fluid shear stress on the proliferation of osteoblasts and the regulatory role of the Wnt/β-catenin signaling pathway in cell proliferation, a new method based on endoplasmic reticulum stress and Wnt/β-catenin signaling pathway stress-mediated was proposed. Taking MG63 osteoblasts as the research object, they were inoculated on glass slides (G group), polished titanium sheets (P group), and sandblasted acid-base treated pure titanium sheets (S group). In addition, FSS of 0 dunes/cm2 (static group) and 12 dunes/cm2 (stress group) were given, respectively. Then, quantitative reverse transcription-PCR (RT-qPCR) and western blot were used to detect the mRNA and protein expressions of low-density lipoprotein receptor-related protein 5 (LRP5) and β-catenin in MG63 cells. The results showed that the expression levels of β-catenin mRNA and protein in cells in the stress group were significantly increased ( P < 0.05 ), and the protein expression level of LRP5 was significantly decreased ( P < 0.05 ). The expression level of LRP5 in group S was greatly inhibited, while the expression level of β-catenin was significantly upregulated. Therefore, FSS can stimulate the expression of LRP5 and β-catenin in osteoblasts. Fluid shear stress can promote osteoblast proliferation in vitro; the Wnt/β-catenin signaling pathway is involved in regulating fluid shear stress to promote osteoblast proliferation.