Osteosarcoma, is a kind of malignant tumor derived from malignant interstitial cells. The pathogenesis of osteosarcoma remains unclear and despite use of chemotherapy drugs, resistance to these drugs affects the success of treatment. The present study was conducted to investigate the effects of icariin (ICA) on osteosarcoma cell proliferation and to investigate the role of the Wnt/β-catenin signaling pathway in the inhibition process of ICA on osteosarcoma cell proliferation. Different concentrations of ICA were selected to treat the osteosarcoma cell line 143B for 24 h, and then the onset concentration of ICA when it inhibited the growth of osteosarcoma cancer cell line 143B was detected via an MTT assay. The effect of ICA on the apoptosis of colon cancer cell line 143B under this concentration was detected using a flow cytometer. RNA in osteosarcoma cell line 143B was extracted, followed by reverse transcription. The expression levels of related and apoptotic proteins in the Wnt/β-catenin signaling pathway using ICA were detected by semi-quantitative PCR and western blot analysis, respectively. The expression quantities of vascular endothelial growth factor (VEGF) and MMP-9 were detected by ELISA. MTT assay showed that ICA inhibited the growth of 143B when its concentration was 5 µM (p<0.01). Flow cytometry showed that the number of apoptotic cells after ICA treatment was significantly higher than that in control group (p<0.01). RNA in osteosarcoma cell line 143B was extracted, followed by reverse transcription. Semi-quantitative PCR and western blot analysis revealed that the expression levels of p-GSK3β, β-catenin, c-Myc and cyclin D1 in cells after ICA treatment were significantly downregulated (p<0.01), while the expression level of caspase-3 was significantly increased (p<0.01). ELISA showed that the expression quantities of VEGF and MMP-9 were significantly decreased (p<0.01). Thus, ICA can significantly inhibit osteosarcoma cell proliferation and promote osteosarcoma cell apoptosis, which may be realized by affecting the expression of the Wnt/β-catenin signaling pathway and blocking the expression of related proteins.