Development Of Biomimetic Bone Unit Structure Bioglass Material And Its Application In Bone Tissue Engineering

Background and Objective:The reconstruction of bone defects caused by trauma,infection,tumour,destruction caused by implants or other diseases with extensive bone loss is a challenging problem in clinical practice.For large bone-defect,or improper treatment of bone defect will lead to a high incidence of bone nonunion and osteonecrosis,and ultimately only amputation can be performed.At present,the treatment of bone defect includes autogenous bone graft and allogeneic bone graft,however,the shortage of bone source and the existence of rejection reaction make it very difficult.Bioglass(BG)is a kind of biomaterial which can promote the healing process of bone tissue defects and achieve its physiological function.Because of its good osteogenic activity and biocompatibility,BG is expected to be a new substitute material for bone defect repair.Cobalt(CO)ions can produce hypoxia like reactions and stimulate the production of a variety of angiogenic factors.In our study,a biological scaffold(lotus sprout scaffold,LSS)with porous structure was synthesized from lotus root,and BG and Co were loaded on it.Finally,we designed a biomimetic osteon structure bioglass material(CoBG@LSS)which have a good advantages of great osteogenic activity and promoting vascularization and neurogenesis in bone tissue.Eventually,we expected this bioglass provide a more effective bone substitute material for the treatment of large bone defects in the clinical.Method:1 Synthetic lotus sprout scaffold: The scaffold with porous structure was obtained by calcining in reaction kettle and drying in vacuum dryer of the lotus root.2 Synthetic bioglass and Cobalt-containing bioglass: Bioglass was synthesized by sol-gel method,and cobalt-containing bioglass was prepared by adding cobalt into bioglass.3 Synthetic lotus sprout scaffold cobalt-containing bioglass: We put the prepared lotus sprout scaffold into the biological glass solution,after the solution was formed into gel,after cutting,shaping and calcining,we obtained the lotus sprout scaffold bioglass and the lotus sprout scaffold cobalt-containing bioglass.4 Observe the structure of bioglass: The structure of lotus sprout scaffold,bioglass,cobalt-containing bioglass and lotus sprout scaffold cobalt-containing bioglass was observed by scanning electron microscope(SEM)to verify the porous structure was exist in modified bioglass.Energy dispersive spectroscopy(EDS)was used to detected the composition of its elements.5 Biocompatibility experiment and cell adhesion experiment: The bioglass with different boron content was immersed in artificial body fluid(SBF)and its quality and pH changes were detected.The bioglass material with lotus root scaffold was co-cultured with rat bone marrow derived mesenchymal stem cells(BMSCs).The adhesion of cells on the surface of the material was observed under scanning electron microscope.6 Measure cytotoxicity: The lotus sprout scaffold cobalt-containing bioglass was co-cultured with BMSCs.Cell survival was determined by CCK8 and fluorescence staining of living and dead cells to verify cytotoxicity of bioglass materials.7 Measure osteogenic activity and promoting vascularization and neurogenesis activity: Bioglass,cobalt-containing bioglass,lotus sprout scaffold bioglass,lotus sprout scaffold cobalt-containing bioglass and blank control group was implanted into rat models of skull bone defect,and the degree of skull healing was observed by MicroCT after 28 days.Eventually,Western blot(WB)was used to detect the expression levels of COL-1,OCN,VEGF and GFAP.Result:1 The results of SEM observation and XRD indicated that the bioglass with porous structure was successfully synthesized.2 Cell adhesion experiment: Rat BMSC adhesion was observed on the surface of the bioglass which co-cultured with rat BMSCs under SEM.3 Measure cytotoxicity:The results of CCK8 and fluorescent staining of living and dead cells showed that BMSCs had a good cell survival rate in lotus sprout scaffold cobalt-containing bioglass,and the low concentration of material infusion had little effect on the cell survival rate.4 Animal experiments : MicroCT results showed that compared with the blank control group,the skull defect area of the group implanted with bioglass,Cobalt-containing bioglass,lotus sprout scaffold bioglass and lotus sprout scaffold cobalt-containing bioglass was smaller after 28 days of construction,and the skull defects in the lotus sprout scaffold bioglass group and the lotus sprout scaffold cobalt-containing bioglass group were nearly completely healed.5 Western blot: The results of WB showed that bioglass,Cobalt-containing bioglass,lotus sprout scaffold bioglass and lotus sprout scaffold cobalt-containing bioglass all have a high expression level of COL-1,OCN,VEGF and GFAP.Among them,the expression levels of 4 proteins in the lotus sprout scaffold cobalt-containing bioglass group were higher than those in the other group.Conclusion1 Bioglass and cobalt-containing bioglass were successfully prepared by sol-gel method.In addition,the porous structure of lotus sprout scaffold was obtained by imitating biological method,and the bioglass was successfully load onto it,so we can integrate the advantages of both.2 The implant with porous structure not only has better biological compatibility,but also this porous structure can promote bone healing more quickly.In the process of bone formation,both blood vessels and nerves grow in,which simulates the havers system structure of human body.