RESUMO
BACKGROUND: Although icariin has a clear role in inducing differentiation of bone marrow stromal cells, no studies have reported on its use in proliferation of periosteal cells in animal experiments. Three-dimensional polycaprolactone scaffold is a new scaffold, and it has the characteristics of biodegradability, proper pore size, and good histocompatibility. OBJECTIVE: To investigate the effect of icariin on the proliferation of periosteal cells, to construct periosteal cells and three-dimensional scaffold composite tissue-engineered bone and study its therapeutic effect on radical defects in rabbits, so as to provide a new way for the treatment of bone defects . METHODS: The mandibular periosteum was obtained from the rabbit and the primary cells were obtained by tissue explant method. The passage 3 cells were planted on 24-well plates. In experimental group, icariin at 10-4, 10-3and 10-2mol/L were added into the corresponding cell culture wells; equal volume of PBS was added to control group; positive control group was treated with 10 μg/L basic fibroblast growth factor. The cell proliferation in each group was measured by MTT method on the 2nd, 4th, 6thand 8thdays, respectively. The three-dimensional polycaprolactone scaffolds,10-2mol/L icariin and 107/L periosteum cells were placed in cell culture plates and cultured in vitro to construct the cell-scaffold complex. After the rabbit radical defect model was established, the model rabbits were randomized into three groups (n=15 per group), followed by implanted with scaffold complex (experimental group), polycaprolactone scaffold (polycaprolactone group) or nothing (control group). Pathological sections were taken at the 2nd, 4thand 8thweeks respectively for hematoxylin-eosin staining to observe the bone and count the number of osteoblasts. The X-ray films of the radius were obtained at the 4thweek. RESULTS AND CONCLUSION: (1) The proliferation of periosteal cells induced by different concentrations of icariin, especially 10-2mol/L icariin, was better than that in the control group (P < 0.05), Therefore, the optimal concentration of icariin was 10-2mol/L. (2) At 4 weeks after transplantation, in the experimental group, there were more osterocytes and less microvessels; in the polycaprolactone group, there were more new vessels, less woven newly born bone and osterocytes; in the control group, the defect region was filled with more granulation tissue and less osterocytes. At the 8thweek, complete healing in the defect region was observed in the cell-scaffold complex group, while partial healing in the polycaprolactone scaffold group. In the control group, however, fibroblasts and scar tissues were visible in the defect region, with presence of closed medullary cavity and nonunion. The number of osteocytes in the experimental group was significantly higher than that in the other two groups (P < 0.05). X-ray examination at the 4thweek showed that the defect region in the experimental group achieved healing;in the polycaprolactone group, the callus formed and defect region connected partly; and there was a clear gap in the defect region of the control group. Our findings indicate that icariin at different concentrations can promote the proliferation of periosteal cells and the concentration of 10-2mol/L has the greatest role. Icariin can induce proliferation of periosteal cells in the three-dimensional polycaprolactone scaffold, which contributes to bone repair.
RESUMO
BACKGROUND: Icariin has a broad prospect for promoting cell proliferation. Differentiation direction of periosteal cells is uncertain, but the cells are easy to be induced by ultrasound, oxygen or bone morphogenetic protein 7 (BMP7). Periosteal cells have been applied in bone tissue engineering; however, icariin effects on the proliferation and differentiation of periosteal cells is little reported.OBJECTIVE: To investigate the effect of icariin on the proliferation and differentiation of human periosteal cells, thus providing theoretical basis for icariin applied in bone tissue engineering. METHODS:The human periosteum was obtained and the primary cells were isolated in vitro.After culture and expansion,periosteal cells were cultured in 24-well plates, and induced by 0.001, 0.01 and 0.1 mg/L icariin and 50 μg/L BMP7, respectively. The corresponding avsorbance values of different groups were detected. The levels of alkaline phosphatase and calcium nodules in periosteal cells were measured at 1, 3, 5 and 7 days, and the mRNA levels of osteocalcin, osteopontin and Runx2 were detected at 3, 5 and 7 days. RESULTS AND CONCLUSION: The periosteal cells proliferated well after induction with icariin, and could proliferate well in different concentrations of icariin and the positive control group (P < 0.05). Compared with the control group, the periosteal cells induced by icariin were able to produce more alkaline phosphatase and calcium nodules (P < 0.05). The mRNA expression of osteocalcin, osteopontin and Runx2 in periosteal cells could be up-regulated by icariin (P < 0.05). These findings imply that icariin can promote proliferation and differentiate of periosteal cells into osteoblasts, and it can be used as an inducer for the preparation of seed cells in bone tissue engineering.