A total of 26 patients comprising 14 with delayed fracture healing and 12 with bone nonunion were treated via subperiosteum and fractured spatial injection of osteoblasts. Bone marrow was extracted from posterior superior lilac spine to obtain osteoblasts following in vitro induction, culture and amplification. Subsequently, 5-8 mL osteoblasts at a density of 1 ×105 cells/mL was sterilely injected into subperiosteum and fractured interspace in injured region using X-ray positioning. At 4, 6, 10 and 14 weeks after injection, callus formation was checked using X-ray. All cases followed up for 3-12 months, with the means of 5.3 months. Callus was formed after 4 weeks, fractured ends were wrapped up by callus after 6 weeks, and fracture line was unclear after 10 weeks and disappeared after 14 weeks. Bone fracture was healed, with the mean healing time of 12.1 weeks, suggesting that percutaneous subperiosteum injection of osteoblasts is an effective method to treat delayed fracture healing and bone nonunion.