| Background and objective:Osteoporosis hinders bone healing in patients with bone injury.The trace element zinc(Zn)has been shown to stimulate bone formation in humans and many animals.Meanwhile,zinc plays an important role in the occurrence and development of osteoporotic fractures.Although the addition of zinc can improve the osteogenic properties of bone repair materials,the optimal zinc dosage is uncertain and varies with the type of zinc carrier material.The use of scholzite(ZCP)and polymer composites as bone graft substitutes has not been reported to date.The research selected polylactic acid(PLA)/nano-hydroxyapatite(nHA)composite with good application prospects.Different masses of nano-ZCP(nZCP)were mixed into the composite,in order to find the mass fraction of nZCP with better osteogenic effect on osteoporotic bone defects.Methods:PLA/nHA and PLA/nHA/nZCP scaffolds were constructed by freeze-drying technology.The experiments are divided into three parts:material characterization,in vitro cell experiments and in vivo animal experiments.Four experimental groups containing different nZCP mass fractions,PLA/nHA,PLA/nHA/0.1ZCP,PLA/nHA/0.3ZCP,and PLA/nHA/0.5ZCP,were established.The osteogenic activity of PLA/nHA/nZCP scaffolds was assessed by using rat bone marrow stromal cells(rBMSCs)as a cellular model.The bone regeneration ability of the scaffolds was investigated using an ovariectomized(OVX)rat model of critical size femoral condyle defect.In the characterization section,scanning electron microscopy(SEM),energy dispersive X-ray spectroscopy(EDS),X-ray diffraction(XRD),Fourier transform infrared spectroscopy(FTIR)were used to determine the scaffold composition;,determined the compressive strength of the material,as well as the release concentration of calcium and zinc ions.In cell experiments,the biocompatibility of the materials was evaluated by CCK-8,dead-live staining,and the pro-osteogenic properties of the materials were explored by alkaline phosphatase(ALP)staining and activity analysis,alizarin red staining,real-time quantitative PCR(qPCR)and western blotting.In the animal experiment,6 SD female rats in each group were used to make the OVX combined femoral condyle bone defect model.The samples were taken at 6 weeks and 12 weeks,and μCT scanning and histological staining were performed to evaluate the bone repair.Results:The scaffold presented a three-dimensional porous structure with 40μm diameter pores distributed inside,and nHA and nZCP were evenly distributed in the scaffold.The ion release experiments showed that the zinc ions released by the three groups containing nZCP were all within the effective osteogenic concentration.In cell experiments,the results of CCK-8 and dead-live staining experiments showed that the four experimental groups had good biocompatibility.Scaffolds containing different masses of nZCP increased ALP activity and extracellular matrix mineralization to varying degrees,and increased the expression of ALP,Runt-associated transcription factor 2(RUNX2),and osteocalcin(OCN)genes.Compared with zinc-free scaffolds,PLA/nHA/nZCP scaffolds provided a more favorable microenvironment and promoted the growth of new bone.In animal experiments,the 6-week data showed that the incorporation of nZCP improved the osseointegration of the material,and the 12-week data observed more new bone formation in PLA/nHA/0.3nZCP and PLA/nHA/0.5nZCP than the remaining two groups of bone defects.Conclusion:The incorporation of nZCP improves the osteogenic properties and osseointegration ability of PLA/nHA composite scaffolds without affecting the lattice parameters of nHA,and can be uniformly distributed in the scaffolds.In the PLA/nHA group incorporating nZCP,the osteogenic activities of PLA/nHA/0.3ZCP and PLA/nHA/0.5ZCP were better,and the nZCP mass fraction at this time was 4.5%-7.5%,that is,in the In the PLA/nHA system,when the mass fraction of nZCP is 4.5%-7.5%,it has better osteogenic properties. |
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