Experimental Study On Expression Of Alkaline Phosphatase Activity Of Bone Marrow Stromal Cells Induced By Platelet-rich Plasma

Purpose To study the changes of expression of alkaline phosphatase activity (ALP) and mRNA of bone marrow stromal cells induced by platelet-rich plasma (PRP) in vitro and to evaluate the effects of PRP on osteoblastic differentiation in an early state or pre-mature osteoblasts.Method PRP and platelet-poor plasma (PPP) were adopted from Appel and modified for the use of dog blood. Bone marrow was harvested from the femors of dog. Bone marrow cells were cultured in DMEM/F12 supplemented with 15% foetal bovine serum. Bone marrow stromal cells (BMSCs) were purified by culture-expanded as undifferentitated cells in culture for more than 5 passages. After treated with PPP, bone morphogenetic protein (BMP), and different concentrations of PRP, RT-PCR was performed to assay changes of mRNA expression of alkaline phosphate on 6th day. Kinetic method assay was used to measure the alkaline phosphate activity in 3-day intervals during a period of 12 days. Cultures without additional supplements were used as controls. Mineralization was detected by Von Kossa staining techniques.Results Adherent cells were observed after bone marrow cells were cultured for 24h, and fibroblast-like cells were formed to connect with adjoining cells in 12 days. After several culture-expanded, BMSCs with various morphologic features were acquired. The level of expression of ALP mRNA in the group treated by low-concentrations of PRP was significantly higher than that in the control group. During the 12 culture days, ALP activities of all groups increased steadily (P<0.05). After the 9th day, ALP activities tended to decline. Mineralization was detected by Von Kossa staining techniques. Conclusion Low-concentrations of PRP increases the expression of ALP mRNA and promotes alkaline phosphate activity in BMSCs. It suggests that low-concentration of PRP could induce differentiation of the BMSCs into premature osteoblasts. The marrow stromal cells from dogs could be promoted to the osteoblastic phenotype in vitro by low-concentrations of PRP.