Periodontitis is a local inflammatory environment with dysregulation of host responses, which results in destruction of periodontal tissues. Mesenchymal stem cells (MSCs) have been proven to play important roles in tissue regeneration by serving as progenitor cells, but its therapeutic outcomes are yet, evaluated variable and unpredictable because of the influence of local inflammation. Acetylsalicylic acid (ASA) has been reported to benefit for MSCs in terms of inflammation control and tissue regeneration. In this study, we aimed to explore the effect of bone marrow mesenchymal stem cells (BMMSCs) pretreated with ASA (ASA-BMMSCs) on periodontal bone repair in a ligature and bacteria-induced periodontitis model in rats. We show herein that, ASA-BMMSCs treatment reduced inflammatory infiltration and alveolar bone loss in periodontitis rats, reflected by immunohistochemistry staining of OPG/RANK-L and Micro-CT. Levels of TNF-α and IL-17 decreased while IL-10 increased after the treatment of ASA-BMMSCs in periodontitis rats. In addition, less osteoclasts number was detected in ASA-BMMSCs treated group. In vitro study showed that ASA facilitated BMMSCs proliferation and differentiation, which might explain the reduced bone loss in periodontitis. These results together suggest that local application of ASA-BMMSCs in periodontal lesion sites is capable of improving inflammatory microenvironment, promoting alveolar bone regeneration, thus leading to a recovery of periodontal homeostasis. Besides, this study also provides us a new idea that a combined application of ASA and BMMSCs may be a novel approach for periodontitis treatment and periodontal bone regeneration.