Strontium-Substituted Nanohydroxyapatite Incorporated In Situ Gelling Hydrogels As Scaffolds For Bone Tissue Engineering

Objective:Biomaterial-based bone tissue engineering promotes the bone regeneration by providing osteogenic scaffolds,cells and growth factors.Hydrogels are often used as temporary scaffolds for bone tissue engineering because of their good biocompatibility.However,the low mechanical strength of hydrogels often limits its clinical application.In order to solve this problem,many studies focus on adding a variety of mineral particles to the hydrogels.The purpose of this study was to synthesize a novel in situ hydrogel by incorporating strontium-substituted nanohydroxyapatite to enhance the mechanical property of the hydrogel while improve the osteogenic property of the material.Method:1.The strontium substituted hydroxyapatite(Sr-nHA)with different strontium substitution rate(0%3 50%,100%)was synthesized by co-precipitation method,which were termed as:nHA,Sr50nHA,Sr100nHA.The pure hydrogel and the hydrogels containing different Sr-nHA were synthesized by Schiff-base reaction,which were called as:CDH,nHA/CDH,Sr50nHA/CDH,Sr100nHA/CDH.2.Fourier transform infrared(FTIR),x-ray diffractometer(XRD)and scanning electron microscopy(SEM)were used to verify the successful synthesis of Sr-nHA and incorporation into the hydrogels.3.Observe the physical properties of the newly synthesized hydrogels,including swelling properties,degradation,gelling time and rheological properties.4.Determine whether the material could promote cell proliferation and osteogenic differentiation by co-culturing MC3T3-E1 with the material.5.Establish a rat calvarial defect model and evaluate the in vivo osteogenic effect of the material by micro CT and HE staining analysis.Results:1.It can be seen from the FTIR images that the peaks of Sr-nHA were similar to those of Sr-nHA/CDH,the appearance of phosphate characteristic peaks in Sr-nHA/CDH suggested the successful incorporation of Sr-nHA.XRD images confirmed that the synthesized material was Sr-nHA with good crystalline.Among them,S100nHA/CDH had the best crystallinity,while the Sr50nHA/CDH had the poorest crystallinity.2.Scanning electron microscopy images showed that the composite scaffold had a typical three-dimensional porous structure.The surface of the CDH pore wall was smooth;the surface of the Sr-nHA/CDH pore wall was rough,and nanoparticles were dispersed on the pore surface with a small degree of agglomeration.3.Swelling experiments and degradation experiments showed that Sr-nHA/CDH had better swelling performance and degradation properties than CDH.However,there was no significant difference between the three Sr-nHA/CDH hydrogels.The gelling time measurement results indicated that the incorporation of nanoparticles significantly decreased the gelling time;the rheological results indicated that Sr-nHA/CDH had better mechanical properties than CDH.4.In vitro cell co-culture results suggested that Sr-nHA could promote the proliferation of MC3T3-E1 and enhance ALP activity in early osteogenic differentiation.The results of osteogenic gene test confirmed that Sr-nHA/CDH promoted MC3T3-E1 differentiation both in the early and late differentiation stage.Sr100nHA/CDH displayed excellent osteogenesis especially in the late stage.5.Micro CT and HE staining results confirmed that Sr100nHA/CDH had better bone regeneration compared with CDH,nHA/CDH,Sr50nHA/CDH at 4 and 8 weeks after surgery in vivo.Conclusion:The Sr-nHA/CDH synthesized in this experiment is an in situ hydrogel with good biocompatibility,biodegradability and biomineralization.The results of material characterization,physical property tests,in vitro and in vivo osteogenesis tests indicate that,the Sr100nHAP/CDH is considered to be a better bone substitute material.