The Preparation Of Novel Bone Repair Material And Its Vitro Study

Objective:To fabricate porous PGA/β-Tricalcium Phosphate for tissue engineering scaffolds.The microstructure of the scaffold were investigated by SEM, the porosity and the mechanical properties were tested. Use the Platelet-Rich-Plasma as an autologous source of various growth factors. we study the combination of PRP with osteoblast in composite materials and investigate the influence of PRP on proliferation and differentiation of osteoblast in PGA/β-TCP scaffolds.Hoping to give a preliminary experimental evidence and theoretical basis as bone graft materials.Methods:1. The PGA/β-TCP (in1:land1:3weight ratios) scaffolds for tissue engineering were fabricated by means of solvent-casting particulate leaching. The structure of the scffolds were observed using scanning electron microscopy (SEM). Their physical and mechanical properties were tested.2. Calvariae from24-hour initiate rat were removed aseptically and proliferated by the second enzyme digestion. The osteoblast were maintained in DMEM supplemented with10%FBS. Osteoblasts were purified by differential attachment method. Osteoblast were identified by morphology, alkaline phosphatase detection,and Alizarin red staining.3. PRP was prepared by twice centrifugation method. Osteoblasts were cultured on the surface of the PGA/β-TCP scaffolds, and then PRP was spread on the PGA/β-TCP scaffolds.5%PRP/OB/PGA/β-TCP composite scaffold was the experimental groupA,10%PRP/OB/PGA/β-TCP composite scaffold was the experimental group B, OB/PGA/β-TCP composite scaffold was used as control group. samples were obtained2,4,6days after culture. Cell proliferation was detected by MTT method;Cell alkaline phosphatase(ALP) activities were assayed by ALP kit.Results:1. The SEM pictures showed that both scaffolds were three-dimensional (3D) porous composite scaffolds. the β-TCP particles were uniformly dispersed within the PGA matrix.The different between the two groups was that moreβ-TCP granules were present on the pore wall surface of group B compared to group A. The compressive strength of the (B) group were significantly higher than that of the (A) group (P<0.001). Both A and B groups had a high porosity(>85%). The opening porosities of A group scaffolds is higher than B group.respectively, which were statistically different.2. The results light of microscopy,48hours later,the isolated and cultured osteoblasts were adherent growth, diversification in morphology, and mostly showed polygonal, spindle-shaped, triangular, and the cytoplasm was abundant and the spurt was stretched out. cell fusion were found after culturing6days.alkaline phosphatase staining and mineralized nodules alizarin red staining was positive. Both the cell proliferation of the A,B of the experimental group and the control group increased with time. A,B of the experimental group were obviously higher than that the control group (P<0.01). B of the experimental group was the highest OD at the time. After2days culture ALP activity of three groups were very low. After4days and6days of culture, ALP activity of three groups increased with time, ALP activity of B group. There was a significant increase in ALP activity in the experimental group compared to the control group.(P<0.05).Conclusion:1. Both A and B groups had a high porosity(>85%).The compressive strength of the (B) group were significantly higher than that of the (A) group (P<0.001). The PGA/β-TCP scaffolds material (weight ratio=1:3) might be a promising material for bone tissue scaffolds engineering.2. PRP could obviously promote the proliferation of rat osteoblast in PGA/β-TCP scaffolds,and improve the ALP activities of the osteoblast in PGA/β-TCP scaffolds. Besides,10%PRP/OB/PGA/β-TCP composite scaffold,which the proliferation and the ALP activities of the osteoblast is better than5%PRP/OB/PGA/β-TCP group.