It is quite intriguing that infection, which generally results in bone loss, induces osteogenesis under certain conditions, as seen in chronic osteomyelitis and periapical lesions. The mechanism underlying this phenomenon is largely unknown. Accumulating evidence indicates that the immune and skeletal systems interact with each other through various regulators, therefore greatly affect the osteogenic process. Among these regulators, the bioactive lipid sphingosine-1-phosphate (S1P), along with its receptor sphingosine-1-phosphate receptor 1 (S1PR1), has been identified to modulate osteogenesis and the polarization of immune cells such as macrophage ̶ the key player in the immune response against infection, which also interacts with the osteoblast-lineage cells (known as bone marrow stromal cells, BMSCs) to regulate osteogenesis. This study aimed to investigate the role of S1P-S1PR1 signalling in the interaction between macrophages and BMSCs under infectious conditions. Our in vivo results showed that abnormal calcium deposition was formed in human periapical lesions, which was accompanied with macrophage infiltration and upregulation of the S1P-S1PR1 signalling. The in vitro results showed that the osteogenic markers of BMSCs were significantly up-regulated when the BMSCs were co-cultured with macrophages with the supplementation of LPS (to simulate infection); the ALP activities and the formation of mineralization nodules were induced accordingly. Furthermore, the co-cultured macrophages were found to shift from the pro-inflammatory M1 phenotype towards the tissue-regenerative M2 phenotype. Further research revealed that under infectious conditions, macrophages interacted with BMSCs and resulted in over-production of S1P, which on one hand activated S1PR1 on BMSCs to induce the osteogenic process; one the other hand, the S1PR1 activation on macrophages converted them towards the M2 phenotype and hence facilitated osteogenesis. Therefore, our study partially explains the mechanisms of the bone formation in infection-related diseases.