The Gene Expression And Function Of Osteogenic Differentiation On Periosteal Cells In Vitro

ObjectiveTo observe the cytobiologic character of human periosteal cells induced by bone factor BMP7,and evaluate the gene expression and function of osteoblasts differentiated from periosteal cell in vitro.MethodsThe periosteal cells from adult tibia were cultured by routine method in vitro, then divided into experiment group and control group, BMP7 and osteogenic adjuvant were added to experiment group and added osteogenic adjuvant only to control group. The proliferative activity was detected by CCK-8. The expression of the osteocalcin gene and typeⅡcollagen were evaluated by RealTime-PCR, the glycosaminoglycan detected by toluidine blue staining. The levels of ALP, osteocalcin and osteopontin in the supernatant were tested by ELISA on day5, day10, day15 and day20.The ALP activity, ALP and calcium nodules were respectively detected by colorimetry, ALP staining and Von Kossa staining.Results1. The expression of osteocalcin and the level of ALP, osteocalcin and osteopontin in supernatant were both increased with significant difference between the experimental group and control (P<0.05).2. The positive rate of ALP and calcium nodules staining increased with significant difference between the experimental group and control (P<0.05).3. The positive rate of toluidine staining and the gene expression of collagenⅡin the experimental group both increased temporarily and then decreased ,but with significant difference between the experimental group and control (P<0.05). Conclusion1,Human periosteal cells induced by BMP7 can differentiate towards osteoblast with typic morphologic characteristics of osteoblast in vitro, obviously express the osteogenic specific gene, and have the ability to synthetize and then secrete osteogenic specific protein.2,Apart from a direct osteogenic differentiation in the progress of ossification of periosteal cells induced by BMP7, there is a phenomenon that some first formed chondrocyte and then calcified into the bone as well.3,A large number of osteoblasts can be available in vitro via human periosteal cells combined with BMP7 pathway, which can be used in bone tissue engineering construction in vitro and clinical application.