Combination of low-intensity pulsed ultrasound and C3H10T1/2 cells promotes bone-defect healing.

Abstract

We systematically investigated the effect of combined use of low-intensity pulsed ultrasound (LIPUS) and bone mesenchymal stem cells C3H10T1/2 on bone-defect healing. C3H10T1/2 cells were first induced into a stationary phase by incubation with low fetal bovine serum (5 ml/l) for five days and then sonicated with LIPUS for ten minutes once every day for five consecutive days. The same LIPUS treatment combined with C3H10T1/2 cells, which were incubated in regular fetal bovine serum (10 ml/l) were used to aid femoral fracture healing in Sprague-Dawley rats during four consecutive weeks. C3H10T1/2 cell proliferation activity was detected by MTT assay. Cell-cycle changes were determined, and cell proliferation index was calculated using flow cytometry. Bone reparation was evaluated by X-ray imaging and hematoxylin and eosin (H&E) staining during the healing process. LIPUS promoted C3H10T1/2 cell proliferation, the mechanism of which was possibly the up-regulation of Bmi-1 gene expression. At the end of week two after combined use of LIPUS and C3H10T1/2, the femoral gap was reduced on X-ray images. According to H&E staining results, new bone had homogeneous and similar density compared with normal surrounding bone after combined use of LIPUS and C3H10T1/2. At the end of week four, bone defects could not be observed by X-ray in all four groups and repaired bone substance in all four groups could be observed by H&E staining. LIPUS treatment effectively promotes C3H10T1/2 cells to enter the growth/split phase from the stationary phase. This process enhances cell proliferation, which consequently promotes bone-defect healing.