OBJECTIVE: The study investigated in vitro the influences of shear stress preconditioning on human periodontal ligament stem cells (hPDLSCs) under serum deprivation. DESIGN: hPDLSCs were subjected to shear stress at 0.5 and 5 dyn/cm², both with and without serum starvation. Cell viability and apoptosis were assessed using the Resazurin assay and flow cytometry analysis, respectively. Gene and protein expressions were analysed by real-time polymerase chain reaction, immunofluorescent staining, and Western blotting. RESULTS: Our results revealed that shear stress potentially mitigated serum derivation-induced cell death by inducing cell viability, enhancing colony formation, and inhibiting cell apoptosis. The addition of an ERK inhibitor inhibited the shear stress-induced cell apoptosis resistance. Shear stress treatment upregulated cell viability-related gene expression, including SOX2, SOD1 and BIRC5. In particular, shear stress promoted the nuclear translocation of SOX2. Meanwhile, the expression of BIRC5 was not inhibited by cycloheximide. Shear stress-induced SOX2 and BIRC5 expression was attenuated by PI3K and ERK inhibitors, respectively. CONCLUSIONS: Shear stress contributes to promoting SOX2 and BIRC5 expression by hPDLSCs, implicating the promotion of stemness and cell survival under serum starvation.