| BackgroundIt's a major problem for orthopedic surgeons to treat large segmental bone defects.At present,The primary methods treat it including autologous or allogeneic bone grafting and bone substitutes.However,the clinical applicable bone replacements have many disadvantages,such as,rapid degradation,poor osteogenic induction ability and insufficient mechanical support.In this experiment,the performance of calcium sulphate was optimized.The rare earth element cerium was doped to the calcium sulphate in order to improve its osteoinductive effect.Furthermore,to increase the mechanical strength and delay the degradation rate of calcium sulfate,hydroxyapatite(HA)is added into the cerium-containing calcium sulfate(Ce-CSH).In short,we expect to provide a new strategy for the clinical treatment of large segmental bone defects.Methods1.The Preparation,physical characterization and in vitro degradation of Ce-a-CSH/HA.Ce-CSH/HA was made according to the coprecipitation and hydrothermal methods.The phase composition and crystal structure of the composite were studied by X-ray diffraction(XRD)and Fourier transform infrared spectroscopy(FTIR).We observed the surface morphology of the prepared material by scanning electron microscopy(SEM).The composites were immersed in the degradation solution in vitro for 8 weeks.the degradation rate,concentration of cerium and calcium ions were measured.2.The effect of Ce-CSH/HA on osteogenic differentiation of bone marrow mesenchymal stem cells(BMSC)and polarization of macrophage.After being treated with different concentrations of Ce-CSH/HA composites'extracts,the expression levels of osteogenic differentiation-related genes(ALP,Runx2)of BMSC were detected.The alizarin red and Von Kossa staining were performed on BMSC after being incubated with material extracts for 14 days and 21 days,respectively.The effect of material on the proliferation of macrophage was detected using CCK8 method.Additionally,the phenotypic genes(CD86,CD206,Argine),inflammatory factors(IL-1,IL-18,TNF-a,IL-1?)and pro-angiogenic genes(PDGF-BB,VEGF)were also detected.3.The repair ability of critical bone defects in rats by Ce-CSH/HA compositesThe 5%Ce-CSH/HA and CSH/HA composites were implanted into the tibia bone defect of rats.Four weeks later,EMCN and CD31 immunofluorescence were performed to understand the angiogenesis at the bone defect sites.HE and Goldner staining were used to observe the formation of new bone after being harvested at 8 weeks postoperation.Results1.We successfully synthesized the new composite materials(Ce-CSH/HA).The XRD and FTIR results confirmed that the incorporation of Ce and HA did not affect the crystal structure of calcium sulfate.However,we observed that the particle diameter of Ce-CSH/HA composites became smaller than CSH/HA using SEM.The degradation rate of CSH/HA was significantly faster than that of Ce-CSH/HA.After being immersed in SBF in vitro for about 4 weeks,CSH was mostly degraded.At the same time,the concentration of cerium and calcium ions released from composites also reached the maximum.2.Compared with CSH/HA group,Ce-CSH/HA can statistically significantly promote the expression of ALP,Runx2 and OCN genes.Furthermore,the latter's promotion on mineralization of BMSC is more obvious.Ce-CSH/HA has no promotion effect on the proliferation of macrophages.But it can induce macrophage polarizing to M2 type,inhibit the expression of inflammatory factors,and promote the expression of pro-angiogenic factor.3.Histological staining confirmed that the in vivo degradation rate of Ce-CSH/HA was delayed,and the effect of it's promotion on new bone formation was more obvious.At the same time,bone healing was accelerated owing to the newly formation of H-type blood vessels.Conclusion1.The degradation rate of the novel Ce-CSH/HA is delayed,promoting the new bone formation by the Ce ions released from the composites.2.Ce-CSH/HA promote the expression the osteogenic differentiation-related genes of BMSC and improve their mineralization.Additionally,the prepared composites have a regulatory effect on macrophages,such as,inducing macrophage polarization toward to M2,inhibiting inflammatory factors and promoting the expression of pro-angiogcnic factors.3.Ce-CSH/HA composites accelerate the healing of bone defects in vivo.At the same time,the new blood vessels the in the defect promoted by Ce-CSH/HA can also have a positive effect on bone repair. |
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