Fabrication Of Gradient Scaffolds And Observation Of Their Osteogenic Properties

Objective: To make new gradient scaffold,and compare the pore diameter,porosity and compressive strength of gradient scaffolds and homogeneous scaffolds.The biological behavior of osteoblasts on different scaffolds was compared to evaluate the effect of gradient structure on osteoblasts.Methods: Using Hydroxyapatite(HA)as material and wax ball as pore-forming agent,the pore size gradient scaffold was prepared.Specific diameter ball of wax and HA sol mixed compressed into cylindrical.The physical properties of gradient scaffolds were compared with those of uniform scaffolds.Osteoblasts derived from the cranial cap of SD rats were cultured.The proliferation potential of osteoblasts on the scaffolds,ALP activity,OPN and OCN gene expression were detected by flow cytometry to observe the effect of pore size on osteoblasts.Results: The gradient scaffold prepared by the method of wax ball pore-making agent can effectively design and control its internal structure,and the pores are highly interconnected.The phase detection of the sintered scaffold showed that the characteristic peak of HA powder was the same as that of the sintered scaffold grinding powder XRD,and no impurities were added.Sem results showed that the gradient scaffold had interconnected pores,large outer pore diameter,small inner pore diameter and high porosity.The proliferation index(PI)of osteoblasts was measured by flow cytometry,and it was found that the gradient structure could promote cell proliferation,and the optimal pore size to promote osteogenesis was between 350?m~400?m.The ALP activity,OCN and OPN gene expression levels of osteoblasts were significantly increased in the D300 group,suggesting that small pore size could promote the differentiation of osteoblasts.Cytoskeleton staining showed that cells in the d-gradient group had the largest amount of cell adhesion and clearer actin fibers on the scaffold surface,and the gradient structure could promote cell migration and improve the distribution of cells in the scaffold.Conclusion: Gradient scaffold porosity >70%,to ensure nutrition and oxygen flow and to ensure the physical properties of the scaffolds;Gradient scaffolds promote the proliferation,adhesion and migration of osteoblasts on the surface of scaffolds,providing important reference value for the design of bone tissue engineering scaffolds.