Study Of Yunnan Baiyao On The Differentiation Of Human Periodontal Ligament Fibroblasts On Bio-Oss Collagen Scaffold In Vivo

[Objective] The aim of this study was to investigate whether Yunnan Baiyao can induce HPDLFs into osteoblasts in vivo.[Methods] HPDLFs were obtained from explant cultures of healthy human periodontal ligaments taken from premolars that had been extracted for orthodontic purposes. HPDLFs, at passage four, were used in all experiment. HPDLFs were seeded into Bio-Oss Collagen in vitro which are divided into four groups to culture. There were two different concentrations of Yunnan Baiyao medium in50μg/ml and100μg/ml which were served as experimental groups. Medium with100ng/ml rhBMP-2served as a positive control and1%fetal bovine serum(FBS) medium were used as a negative control. HPDLFs-Bio-Oss Collagens were culture in four groups for seven days. Then they were implanted subcutaneously in BALB/c-nu mice. At the8th weeks after implantation, the implants were harvested and the harvests were detected by HE staining and immunohistochemistry to study about the effects of Yunnan Baiyao on HPDLFs-Bio-Oss Collagen cultured in vivo.[Results]1. The compounds complexed HPDLFs with Bio-Oss Collagen scaffold which transplanted subcutaneously in nude mice, showed the results of HE staining and IHC that a lot of long spindle cells attached and radially growed at the edge and interior of Bio-Oss Collagen, and there were some degradation of the scaffold.2. HE staining showed that there were new capillaries and osteoblasts on the bone scaffold, and the statistics of new blood capillaries and osteoblasts in HE staing found that compounds cultured in100μg/ml Yunnan Baiyao medium produce new capillaries and osteoblasts which was significantly different compared with compunds cultured in50μg/ml Baiyao solution, medium with100μg/ml rhBMP-2and the control group, P<0.001.3. IHC slice showed HPDLFs cultured in Yunnan Baiyao differentiated into BSP, OC, OPN and CON-I in animal experiment. Statistics showed that there were significant differences in the expression of intensity of BSP, OC, OPN and CON-I between the experimental group and the control group. Besides, there were significant differences compared the expression of four kinds of protein in100μg/ml Baiyao group in the positive area, the average optical density and integral optical density with50μg/ml Baiyao group, positive group and nagetive group, P<0.001.[Conclusion]1. HPDLFs seeded in Bio-Oss Collagen scaffold implanted into animal body to find that Baiyao has a tendency to differentiate HPDLFs into osteoblasts, the effective concentration was100μg/ml concentration Baiyao medium.2. Baiyao has some potentials in promoting periodontal tissue regeneration, which was served as a cell growing factor for periodontal tissue engineering which are worthy of further in-depth study.