Preparation Of Size-controlled NHA/PLGA Nanocomposite Microspheres And Its Application In Bone Defect Regeneration

Various diseases like cancer,trauma and inflammation not only affect the physical and mental health of patients,but also lead to increasing clinical demand for bone substitute materials.Owing to special oral anatomic structure,oral bone defect caused by oral diseases always show irregular appearance resulting in the high demand for bone substitute materials.However,currently most of the clinical bone implant materials are imported,which are expensive,need to be further enhanced in bone guidance,and difficult to shape.Therefore,the development of new bone implant materials with better bone guidance and easy to shape and operate has become an important subject in the field of stomatology.Recently,due to the rapid development of tissue engineering,microspheres have been investigated to be a great substitute for traditional tissue engineering scaffolds.Currently,several techniques were used to make microspheres.However,each techniques show more or less defect and still need to be improved.Herein,a novel airflow shearing method was successfully invented to prepare size-controlled microspheres rapidly.The optimum process parameters of PLGA(Poly(glycolide-co-lactide))microspheres was investigated.Based on these parameters,different contents of nano hydroxyapatite(nHA)nHA/PLGA microspheres was prepared.The biocompatibility and osteogenesis effect were investigated through the in vitro and in vivo experiments.The optimal concentration of nHA to induce osteogenesis of nHA/PLGA microspheres was also found.It can provide the basis for the research and development of novel microspheres.1.Investigation of the preparation of nHA/PLGA microspheres by using airflow shearing method;1).The optimization of fabrication parameters of PLGA microspheres.The results show that with the parameters of 8% PLGA concentration,30 G nozzle size and10 L/min airflow strength,the sphericity and homogeneity of the PLGA microspheres were favorable with a diameter of 200-350 ?m.2).Based on PLGA microsphere analysis,gradient concentration of HA nanoparticles(5 wt%,10 wt%,20 wt%,40 wt%)HA/PLGA microspheres were successfully prepared under similar parameters.By physical and chemical characterization investigation,airflow shearing method showed great significance to preserve the bioactivity of microspheres.The airflow shearing method was also proved to have a great control of microsphere formation.2.In vitro cytotoxicity experiment and the ability to promote bone differentiation were carried out by mouse pre-osteoblast MC3T3-E1 cells.Cell proliferation test showed that the cell proliferation rate was promoted by the increasing of nHA content in microspheres from 5 wt% to 20 wt%.These microspheres can achieve cell attachment efficiently as well as cell proliferation by cell adhesion staining.Calcium deposition quantity analysis showed that within 0-40wt%,osteogenesis ability was improved with the increasing of HA proportion.3.The preparation of rat cranial critical sized bone defect model,with commercialized bone powder Bio-Oss as a control group in order to investigate the bone defect regeneration effect.By scanning the specimen of rat cranial bone 8 weeks postoperation by using micro CT instrument,it was found that within 0-20 wt%,bone defect regeneration ability was improved with the increasing of HA proportion.When HA proportion increased to 40 wt%,microspheres showed a poor bone regeneration ability.Therefore,bone defect regeneration effect was promoted with the increasing of HA proportion in some scope.20 wt% HA was the optimal proportion in this study.Conclusion:1.Airflow shearing method was a novel method to prepare uniformed,size-controlled biocompatible microspheres.2.The optimal parameters to prepare PLGA microspheres by airflow shearing method:8% PLGA concentration,30 G nozzle size and 8 L/min airflow strength,with a diameter of 200-350 ?m.3.In vitro and in vivo experiments showed that the novel nHA/PLGA microspheres were biocompatible.Bone defect regeneration effect was improved by increasing nHA proportion within 0-20 wt%.20 wt% nHA/PLGA microspheres were optimal for bone defect regeneration and can provide reference for further investigation and clinical applications.