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Excerpt
Introduction: It has been shown that LEPUS accelerates fracture healing in clinical studies and animal models. But by which mechanisms, LEPUS accelerates fracture healing is still unclear. Recently, it has been suggested that the interaction between osteoblasts and endothelial cells is involved in the process of bone formation. In the present study, we examined the effects of LEPUS and 1,25(OH)2D3 on cells, using the co-culture system of human osteoblastic cells with endothelial cells.
Materials and Methods: The co-culture system was established with human osteoblastic cells (SaOS-2) and human umbilical vein endothelial cells (HUVEC). SaOS-2 were cultured at a density of 1.5 × 105 cells/well in human type I collagen-coated 6 well culture plate. McCoy's 5A medium supplemented with 10% FCS and antibiotics was employed. One day after plating, HUVEC were cultured at a density of 2 × 105 cells/well onto plated SaOS-2. One day after, medium were changed to the new medium containing 1,25(OH)2D3 (1 × 10−8 M) or vehicle. LEPUS treatment was administered for 20 minutes per day for consecutive 4 days for each well by a ultrasound therapy unit (Sonic Accelerated Fracture Healing System [SAFHS]; Exogen, NJ). LEPUS signal consisted 1.5 MHz, 200 μs burst sine wave repeating pulsation at 1.0 kHz, delivered at an intensity of 30 mW/cm2 spatial and temporal average (SATA). The day after the final treatment, the conditioned medium and cell lysate were collected from each well. The effects of LEPUS and/or 1,25(OH)2D3 were determined by examining cell number and growth factors' concentration in the conditioned medium.
Results: LEPUS treatment increased growth factors (PDGF and bFGF) levels in the conditioned medium. 1,25(OH)2D3 also enhanced the production of these growth factors. The production of PDGF and bFGF in the medium was synergistically enhanced by the combined treatment with LEPUS and 1,25(OH)2D3. These phenomenon were observed when cells were plated on type I collagen-coated well. Interestingly, cells plated on non-coated (plastic) well did not cause these changes. It has been shown that bFGF and PDGF accelerated fracture healing in animal models by its topical injection to the fractured sites.
Conclusion: Low-intensity ultrasound may accelerate fracture healing by inducing various growth factors' secretion from osteoblasts and endothelial cells in vivo.
