The aim of the present study was to investigate the role of Semaphorin 3A (Sema3A) in the osteogenic differentiation of human alveolar bone marrow mesenchymal stem cells (hABMMSCs). To investigate whether Sema3A affects hABMMSC proliferation and osteogenic differentiation， a stable Sema3A-overexpression cell line was generated by infection with the pAdCMV-SEMA3A-MCS-EGFP vector. Cell counting kit-8 and clone formation assays were performed to determine the proliferation ability of hABMMSCs， while cell osteogenic differentiation was assayed using Alizarin Red S staining. In addition， reverse transcription-quantitative polymerase chain reaction was employed to detect the mRNA expression level of osteogenesis-associated genes， Runt-related transcription factor 2 (Runx2)， osteopontin (Opn) and osteocalcin (Ocn)， during the osteogenic differentiation. The results revealed that， compared with the normal control group， the cell morphology of the infected cells was stable and no significant alterations were observed. Overexpression of Sema3A in hABMMSCs significantly increased the cell proliferation ability compared with the control group. Furthermore， the Alizarin Red S staining assay results indicated that the ossification process of hABMMSCs overexpressing Sema3A was evidently faster in comparison with that of the control group cells. Overexpression of Sema3A by pAdCMV-SEMA3A-MCS-EGFP infection also significantly increased the mRNA expression levels of the osteogenic marker genes Runx2， Opn and Ocn. In conclusion， Sema3A was observed to be a key positive regulator in hABMMSC osteogenic differentiation.