To investigate the biological significance of abundant microRNA-23a (miR-23a) expression in osteosarcoma and its correlation with PTEN in the pathogenesis of osteosarcoma migration and invasion. The human osteosarcoma cell lines MG63, HOS58 and SaoS-2, and the human normal osteoblasts (hFOB1.19) were grown in RPMI 1640 medium supplemented with 10% fetal bovine serum. Gene and protein levels of miR-23a and PTEN were examined to determine the molecular relationship between them in the pathogenesis of osteosarcoma. Inhibition of miR-23a effectively reduced migration and invasion of osteosarcoma cell lines. Bioinformatics and luciferase-reporter assay revealed that miR-23a specifically targeted the 3'-untranslational region of PTEN and regulated its expression. Downregulation of PTEN enhanced migration and invasion of osteosarcoma cell lines. Furthermore, in tumor tissues obtained from osteosarcoma patients, the expression of miR-23a was negatively correlated with PTEN and the high expression of miR-23a combined with low expression of PTEN might serve as a risk factor for cancer patients. Besides, miR-23a-mediated suppression of PTEN led to activation of AKT/ERK pathways and epithelial-mesenchymal transition (EMT) in osteosarcoma cells, and finally enhanced the activity of osteosarcoma cell proliferation and movement and promoted osteosarcoma xenograft tumor growth in mouse models. Our study showed that miR-23a, by downregulation of PTEN, enhanced migration and invasion in osteosarcoma cells.