Study On Fabrication And Properties Of A Novel Osteoinductive Self-setting Nacre/Orthophosphate Composite As Bone Substitute

[Background and Objectives]Bone defects are commonly found in orthopedic and dental conditions,which lead to a high morbidity arid severe harm for patient.The disease is one of the challenges for contemporary surgery.Nowadays,utilizing bone substitute biomaterials to treat bone defects is gradually being practiced in clinic,and probably become the mainstream treatment in the future.Orthophosphate biomaterial is one of the most commonly used bone substitute,by attributing to the similarity between its chemical composition and the inorganic chemistry of natural bone,the excellent osteoconductivity and bone replaceability.However,because of the material lacks of osteoinductivity and the slow resorption time,the clinical results are limited.Thus to find a combined material,which meets economical efficiency,good operability and safety,to improve the bioactivities of orthophosphate biomaterial is the focus of current researches.Nacre forms the hierarchically structured internal shell of mollusks.It is a natural organic-inorganic composite.Plenty of In vivo and In vitro studies had proven that nacre possesses potent biocompatibility,osteoinductivity and osteoconductivity,and it is a promisingly ideal bone substitute.However,being restricted by the configuration of nacre,for the studies,the nacre was directly employed by the forms of powder or bulk,otherwise was combined with polymers.So far,to the best of our knowledge,there is no detailed studies have investigated the incorporation of nacre powder with osteophosphate.Thus,our goal of this research is under the premise of keeping the bioactivity of nacre,to incorporate it with osteophosphate.By taking the advantages of both,namely,exerting the osteoinductivity of nacre,meanwhile,using osteophosphate as a malleable carrier,to fabricate a novel osteoinductive bone biomaterial.[Method]The study has been divided into three parts.1.Physical,chemical and biological influence for calcium phosphate cement by incorporating with nacre powder.The nacre powder and the powder phase of calcium phosphate cement(CPC)were homogenized.Then,nacre/CPC composites were prepared by a self-setting process.The CPC and inorganic calcium carbonate/CPC composites were set as control.The characterization,cytocompatibility and cell osteoinductivity were tested.2.The preparation of the novel nacre/orthophosphate composite and the studies on characterization,cytocompatibility,cell osteoinductivity and in vivo degradation rate of the material.An innovational two-step method was adopted to prepare the nacre/orthophosphate self-setting composite.Specifically,for the first step,by using nacre powder and Monocalcium phosphate monohydrate(MCPM)to prepare the brushite(Dicalcium phosphate dihydrate,DCPD)that contains the trace elements from nacre powder.Then,for the second step,by employing the mixture of nacre powder and the product of the first step as the powder phase of the self-setting material,and adopting 2.5%disodium hydrogen phosphate solution as liquid phase.Finally,the nacre/orthophosphate composites were prepared through the processes of mixing of liquid phase and powder phase,molding and self-setting.According to the different mass ratio of nacre powder that was participated in the second step,the final products were divided into five groups.The characterization,cytocompatibility of Hela cell and MLO-y4 cell and cell differentiation of MC3T3-el cell and in vivo degradation rate of the samples from different groups were examined.3.Repair of cortical bone defect on rabbit distal femur with the novel nacre/orthophosphate composite.We established the animal model of the cortical bone defect on rabbit distal and lateral right femur.The grouping of the animals corresponded to the different implants,which were adopted to infill the defect site(Blank control group,nacre/orthophosphate group,hydroxyapatite group).The animals were sacrificed in the time points of 4 weeks and 8weeks after surgery.The femur was retrieved after sacrifice.Then the defect site was analyzed by Micro-CT.The new bone formation,the osseointegration were analyzed.[Result]The first part:?The main inorganic chemical composition of CPC is converted to aragonite,hydroxyapatite and carbonate apatite,from hydroxyapatite,after the incorporation of nacre powder.?The setting time is lengthened,the compressive strength is weakened,and the porosity is increased after incorporating nacre powder into CPC powder phase.?The in vivo degradation rate of CPC is fastened by incorporating nacre powder into CPC powder phase.?The incorporation of nacre powder could promote the cell attachment,cell spreading,cell proliferation and cell differentiation of osteoblasts grown on CPC material surface.The second part:?The final products that prepared by our innovational method are mainly composed by aragonite,brushite,hydroxyapatite and carbonate apatite.?The setting time of nacre/orthophosphate composites are positive correlated with the mass ratio of nacre powder,which participated in the setting reactions.?The compressive strength of nacre/orthophosphate composite is stronger than nacre/CPC composite.?The nacre particles with integrated structure could be detected on the surface of nacre/orthophosphate composites.?Nacre/orthophosphate composites possessed good cytocompatibility,and could promote the growth of dendritic network of osteocytes compared with CPC.?To compare with CPC,nacre/orthophosphate composites could promote the proliferation and differentiation of osteoblasts grown on material surface.?Nacre/orthophosphate composites display a faster in vivo degradation rate than CPC.The third part:?The implant of nacre/orthophosphate composite does not give rise to infection,rejection or other adverse reactions.?Nacre/orthophosphate composite could stimulate the formation of new trabecular bone in the early stage of bone repair.?To compare with hydroxyapatite,the implant of nacre/orthophosphate could regenerate more new bone tissue in bone defect site.[Conclusion]1.Nacre powder is an addictive,which could efficiently promote the osteoinductivity of CPC material2.To adopt the method of self-setting to incorporate nacre with orthophosphate under the circumstances of the reaction conditions,such as temperature and PH value,are moderate could be a feasible means to improve the biological properties of this kind of biocomposite.3.To incorporate with nacre powder is an effective method to promote the degradation rate of orthophosphate material.4.The novel self-setting nacre/orthophosphate composite,which is fabricated by the innovational two-step method,possesses good biocompatibility,osteoinductivity.The material is an ideal bone substitute.Meanwhile,the method is easy-to-operate and repeatable,which could offer a new opportunity to translate nacre into clinical practice.