Evaluation Of BMMSCs/EPCs Sheet Repairing Alveolar Bone Defect In Ovariectomized Rats

Background: At present, many kinds of diseases lead to alveolar bone defect such as periodontal diseases, oral tumor and oral trauma. So alveolar bone defect is an important research across the globe and attracts much attention from researchers. The result of the repair of bone defect relates with the wound size. The repair of defect will be the fibrous connective tissue instead of bone if the size of defect is too large to repair by itself. Autotransplantation and allotransplantation are two main methods on the repair of bone defect, however, they’re limited by postoperative complication. In recent years,with the development of tissue engineering technology,a new method of the regeneration appeared. In this study, through the technology of cell sheet construction, micro-CT and chemical staining, we repaired the alveolar bone defect with BMMSCs and EPCs which constructed the composite cell sheet and evaluated the effect of repair of defect. We enhance the BMMSCs proliferation and differentiation to bone cell and the effect of the repair of defect. This study established the theoretical basis for the repair of alveolar bone defect, and found a new strategy in repair of defect.Objective: To evaluate the ability of the BMMSCs and EPCs which construct the composite cell sheet in the repair of the alveolar bone defect in ovariectomized rats, and to reveal a new method for bone repair.Contents:(1) To isolate the BMMSCs and EPCs, the density gradient centrifugation and adherent culture method was used. The cellular morphology and memebrane antigen of cultured cells were tested by the method of microscope and flow cytometry. The multi-differentiation potential of cultured cells were detected by osteogenic and adiopogenic induction.(2) After ten days of co-culture of BMMSCs and EPCs, BMMSCs/EPCs composite cell sheet were constructed by the induction solution of vitamin C. After fixed, cell sheet were blocked with paraffin, and sections were carried out for H&E and ALP staining, observing by microscope.(3) After established the rat alveolar bone defect model, we implanted the cell sheet into defect or not as control. Using Micro-CT to evaluate the repair of defect.Methods: In the study, ovariectomized rats were frequently used as experimental model of osteoporosis in this experiment. BMMSCs/EPCs, EPCs and BMMSCs were isolated respectively according the proportion dropping into 6-well plates from normal female rats and cultured in α-MEM medium for 24 hours, and the medium was instead of sheet induction medium. Then these three kinds of cells formed the cell sheet what cultured in cell sheet-inducing medium after 10 days later. The cell membrane was implanted into the defect of the alveolar bone in ovariectomized rat respectively, or not implanted for comparison. The result of the repair of the alveolar bone was assessed by histological observation and Micro-CT scanning in 2-, 4- and 8-weeks after operation.Results: We get the results as the following: After inducing the cells ten days, BMMSCs and EPCs could construct composite cell sheet. The expression of calcification of BMMSCs/EPCs composite cell sheet group is more than BMMSCs sheet group and EPCs sheet group. BMMSCs and EPCs sheet group have higher ability in repair of alveolar bone defect than BMMSCs sheet group and EPCs sheet group.Conclusion: BMMSCs/EPCs sheet group from normal rats was better in the osteogenesis activity and the capacity of bone repairing than BMMSCs sheet group and EPCs sheet group.