The SPHK/S1P signaling axis is involved in modulating osteoblast proliferation and differentiation in microgravity

Space-induced bone loss remains a significant health challenge for long-term human space exploration. Under microgravity conditions, osteoblast-mediated bone formation is suppressed, contributing to bone loss; however, the precise mechanisms are not yet fully understood. Sphingosine-1-phosphate (S1P), a bioactive lipid metabolite derived from cell membranes, is crucial for maintaining bone metabolic homeostasis by regulating osteoblast function. In this study, a simulated microgravity environment created by a random positioning machine (RPM) and real weightlessness conditions aboard the Chinese Space Station (CSS) were utilized. This study examined the proliferation and mineralized nodule formation in the osteoblast cell line MC3T3-E1 and primary osteoblasts (POBs) isolated from mouse calvariae. Concurrently, changes in the expression levels of SPHK/S1P-related signaling molecules and key regulatory factors involved in bone formation were analyzed. The results demonstrated that activation of the S1P signaling pathway using the S1P receptor agonist FTY720 significantly promoted osteoblast proliferation and mineralization, partially reversing the microgravity-induced suppression of osteogenic function. Under RPM-simulated microgravity, FTY720 treatment enhanced alkaline phosphatase (ALP) activity, increased calcium nodule formation, and upregulated osteogenesis-related gene and protein expression. Further analysis showed that microgravity primarily downregulated the expression of S1P receptors (S1P1 and S1P2) and S1P kinases (SPHK1 and SPHK2), accompanied by upregulation of S1P phosphatase (SPP) and S1P lyase (SPL). However, administration of exogenous S1P and FTY720 significantly reversed these effects. Spaceflight experiments aboard the CSS further confirmed that FTY720 significantly enhanced osteoblast proliferation and promoted mineralized nodule formation in POBs. These findings suggest that the SPHK/S1P signaling axis represents a critical gravity-responsive pathway regulating osteoblast proliferation and differentiation under microgravity conditions.