ObjectiveBone incision leads to interrupted and sluggish blood flow in the process of distraction osteogenesis (DO), creating a hypoxia (0–2% oxygen tension) at the center of the bone callus. This hypoxia is critical in the coupling of osteogenesis and angiogenesis during DO. This study aimed to investigate the effect of Uniaxial Static Strain (USS) on osteogenesis in osteoblasts under hypoxic conditions, with a focus on the expression of osteogenic markers and angiogenic factors.MethodsThe USS was made by a multi-unit tension compression device.Osteoblasts were subjected to 10% USS made under hypoxic conditions to mimic the process of DO in vitro. The cell proliferation, alkaline phosphatase (ALP) activity, mineralized nodule formation, and expression of osteogenic and angiogenic markers were evaluated by using a CCK-8 assay, alkaline phosphatase (ALP) staining, ALP activity assay, alizarin red S staining, qRT-PCR, Western blotting and ELISA.ResultsHypoxia inhibited osteoblast cell proliferation, ALP activity, mineralized nodule formation, and the expression of runt-related transcription factor 2 (Runx- 2), osteopontin(OPN), osteocalcin (OCN), collagen type I (Col1a1). Conversely, hypoxia upregulated the expression of hypoxia-inducible factor 1-alpha (HIF-1α) and vascular endothelial growth factor (VEGF), which are associated with angiogenesis. However, the application of USS enhanced osteoblasts’ osteogenic capacity and upregulated angiogenic factors under hypoxic conditions.ConclusionUSS can enhance osteogenesis in osteoblasts under hypoxic conditions. Moreover, it may stimulate angiogenesis by promoting the expression of VEGF, which further contributes to bone formation. This finding provides important implications for understanding the mechanisms involved in bone regeneration and may have clinical applications in optimizing the effectiveness of DO techniques.