Preparation And Characterization Of Whitlockite/Calcium Phosphate Composite Scaffold And Studiea On Its Osteogenesis

Natural bone tissue is composed of collagen nanofibers coated with hydroxyapatite(HA)and whitlockite(WH)nanoparticles.As a commonly used bone repair material in bone tissue engineering,HA provides mechanical strength,and has good biocompatibility and osteoconductivity.However,WH as the second largest inorganic calcium phosphate of bone mineral material has been rarely studied due to its harsh synthesis conditions and other reasons.As the second largest inorganic component of bone minerals,WH is negatively charged on its surface and can adsorb proteins,thereby having a stronger ability to induce bone formation,and WH degrades faster in the body,releasing a large amount of Mg2+and PO43-to induce the early formation of bone differentiation.HA/WH inorganic composite scaffolds have been rarely studied because of the difficulties in WH preparation and the inherent brittleness of pure inorganic ceramic scaffolds.Therefore,in this experiment,different preparation methods of WH were studied.By exploring the method of HA/WH composite inorganic scaffold,the preparation method of pure inorganic HA/WH composite scaffold with stable mechanical properties was further explored,and then the artificially synthesized HA/WH in different proportions.The composite scaffold was used to explore the composition structure,mechanical properties and in vitro biological properties.Finally,a series of HA/WH composite scaffolds were used in rats to explore the synergistic osteogenesis ability of HA and WH in vivo.The specific research methods are:(1)To investigate the synthesis methods of HA/WH composites with different proportions,the synthesis methods of HA/DCPD and biphasic calcium phosphate(BCP)before hydrothermal reaction have been individually explored,which were based on the premise that calcium hydrogen phosphate dihydrate(DCPD)hydrothermally generates WH,?-TCP hydrothermally generates WH,and HA sinters to generate BCP.Considering the sintering stability of the scaffold and other factors,?-TCP and HA composite scaffold raw material have been finally chosen.In particular,X-ray diffraction(XRD)results proved that?-TCP hydrothermally generated WH showed the high crystallinity in SBF solution for 2 to 4 days.(2)The specific process of preparing HA/WH composite scaffold by the organic foam template method has been investigated.Based on the premise of ensuring the controllable composition of the scaffold and the mechanical stability before and after hydrothermal,the composition of impregnation slurry composed of HA nanorods and ?-TCP,the specific process of template scaffold impregnation and the conditions of scaffold molding water and heat have been studied.Characterization results such as XRD and scanning electron microscope(SEM)proved that the composition of the composite scaffold was controllable,and its porosity and mechanical properties have met the requirements of bone tissue engineering.(3)The series of successfully prepared HA/WH composite scaffolds were used for in vitro mineralization and co-cultivation with rat bone marrow mesenchymal stem cells(BMSCs).By using the characterization methods such as CCK-8 and Live/Dead staining,it showed that the scaffolds were noncytotoxic and had good biocompatibility.Furthermore,it could induce osteogenic differentiation in vitro and increase the activity of ALP.With the increase in WH content,the differentiation ability could be improved.The scaffolds were implanted in rats to observe their effects on ectopic osteogenesis.Through characterization methods such as micro computed tomography(microCT)and histological staining,it was observed that the WH-containing scaffolds were degraded to varying degrees within 12 weeks.Micro-CT showed the highest the relative increasing volume of ceramics value could be observed in the 3HA1WH group.The staining results showed that the higher the WH content,the faster the scaffold degraded and more bone could be generated.Overall,the HA/WH scaffold prepared by using organic foam templates could successfully tune the proportion of HA/WH ratio by controlling the composition of the slurry.Moreover,the compressive strength and cell osteogenic activity both in vitro and in vivo of the synthesized three-dimensional composite scaffold could be regulated.This composite scaffold showed promising application prospect as a bone tissue engineering scaffold.