The Study On Magnesium-Containing Microspheres Cement And Its Immuneregulation For Osteogenesis

Injectable bone cement is often used in minimally invasive bone grafting surgery due to its good injectability and osteogenic activity.Currently,the development of injectable bone cement has gradually transformed from bio-inert polymethylmethacrylate bone cement（PMMA）to bio-active calcium phosphate bone cement（CPC）and magnisium phosphate cement（MPC）.However,there is still some issues that limited the clinical applications,including the compact structure and unsatisfactory biodegradability of CPC limited the tissue ingrowth and osteogenesis,and the fast curing reaction in MPC causes short setting time which is not conducive to clinical application.Noval injectable bone cement prepared by using microspheres containing magnesium to replace the powder in MPC can not only retain the biodegradability of MPC,but also have good injectability and mild curing reaction,which makes it more suitable for clinical applications.More importantly,the 3D porous structure constructed by the microspheres containing magnesium and the immunomodulation induced by magnesium ions are also conducive to osteogenesis.Therefore,magnesium containing microspheres（MMSs）were designed and fabricated in this thesis and their surface morphology,pore size distribution and phase composition were characterized.Subsequently,MMSs cement（MMSC）based on MMSs was prepared and its curing mechanism was studied.Then the physiochemical properties of MMSC including rheological properties,heat release,curing time,mechanical strength and ions release were explored.The effects of MMSC on the inflammatory response of RAW264.7 cells and the expression of osteogenic-related genes in rat bone marrow derived mesenchymal stem cells（r BMSCs）were studied in vitro.Finally,the inflammatory response and osteogenesis of MMSC were evaluated in vivo.The main contents of this thesis were as the following:1.The preparation and characterization of MMSs.MMSs were prepared though the droplet condensation method by using gelatin,magnesium carbonate basic and silicon-doped tricalcium phosphate as raw materials.The characterizations of SEM,element mapping,BET showed that MMSs had a good microspherical morphology and a large number of micropores with a diameter of less than 100 nm on its surface and internal.The phase analysis subsequently displayed that the phase composite of MMSs was Mg O,Ca7Mg2P6O24 and Mg2Si O4。2.The curing mechanism and physicochemical properties of MMSC.The results of SEM,element mapping and XRD showed that the curing process of MMSC began from the formation of NH₄Mg PO₄·6H₂O crystals between microspheres,and then a 3D porous scaffold was constructed.Subsequently,the characterizations of physichemical properties indicated that MMSC can better meet the clinical requirements of injectable bone cement compared with CPC and MPC.3.The immuneregulation and osteogenic differentiation of MMSC in vitro.The results of q RT-PCR,flow and ELASA showed that MMSC extract can promote the polarization of M2 macrophages and the osteogenic differentiation of r BMSCs by upregulating the anti-inflammatory gene（CD206 and IL-10）and the anti-inflammatory protein IL-10.4.The immuneregulation and osteogenesis of MMSC in vivo.The results of HE staining,Masson staining,immunohistochemical staining and q RT-PCR displayed that MMSC triggered the acute inflammatory response and then the inflammatory response gradually regressed with the M2 microphages polarization.Finally,the osteogenesis of MMSC was studied through the subcutaneous ectopic osteogenesis model and skull defect model and showed that MMSC can induce osteogenesis effectively.In summary,compared with traditional calcium phosphate cement and magnesium phosphate bone cement,injectable bone cements based on MMSs can not only meet clinical requirements,but also promote osteogenesis by the degradability,3D porous structure and immunomodulatory effects triggered by magnesium ions,showing huge application prospects in minimally invasive bone grafting.