Studies On Novel Injectable Calcium Phosphate Cement-bioactive Glass Composite

In China, there are70million people over the age of50suffering from osteoporosis(OP) and1.81million cases of vertebral compression fractures (VCF) occurred annually.Currently, percutaneous vertebroplasty (VP) and balloon kyphoplasty (BKP) have becomethe main surgical treatment of VCF. Polymethylmethacrylate (PMMA) is the mostcommonly used cement in VP and KP. However, there are several disadvantages ofPMMA: nonbiodegradable, no osteoconductive, osteoinductive and osteogenic activityand the high-polymerization isotherm leading to thermal necrosis of the soft tissues at theaugmentation site. Therefore, to design a novel biomaterial with good flowability,osteoconductive, osteoinductive and osteogenic activity as well as good mechanicalproperty and certain degradation rate for bone regeneration has become the urgentproblem in the field of orthopedics. Calcium phosphate cements (CPC) have been widely used as bone substitutematerials in clinic applications because of their self-setting properties, no thermogeniceffect, high biocompatibility and osteoconduction as well as bone replacement capability.However, the currently used CPC has some limitations due to its poor mechanicalproperties and low biodegradation rate. Furthermore, the osteoconductive properties ofCPC are not sufficient to achieve complete bone repairment under critical conditions, suchas poorly vascularized sites and elderly patients with metabolic disorders. Consequently,the enrichment of CPC with osteopromotive or osteoinductive factors is necessary toimprove its biological performance.As a surface active bone substitute, bioactive glass (BG) has recently attracted moreattention due to their good biocompatibility and bioactivity both in bone and in soft tissues.Studies have shown that close ion exchange between BG and soft issue or bone can bedirectly involved in the metabolism of human bone tissue and repair process. Bioactiveglass such as45S5BG bonds strongly to bone and promotes bone growth with formationof a hydroxycarbonate apatite (HCA) layer and the release of Ca, P and Si ions. As far aswe know, it is not until recently, there were a few relevant literatures available regardingCPC-BG applicably in minimally invasive injectable graft.Objective:The aim of this study is to develop a new injectable biocomposite with excellentbiocompatibility, improved bioactivity and biodegradability by adding BG into CPC.Methods:1. Prepare the CPC-BG composite by adding different proportions of BG45S5powder(10wt%,20wt%,30wt%,40wt%) into CPC powder and mixed with potassiumphosphate buffers (pH7.0) at a given P/L ratio of2.0g/ml. Optimize the compositionratio by comparing setting time and injectability among different groups;2. Fabricate the CPC-BG composite with the selected composition ratio and P/L ratio andinvestigate the composition, morphology/microstructure, setting time, injectability,compressive strength, surface reaction layer formation and degradation of CPC-BG; 3. The osteoblasts (OB) were seeded onto the CPC-BG composite disks and the adhesion,proliferation, morphology and differentiation abilities of the OB were observed;4. The CPC-BG and CPC specimens were implanted into femoral condyle defects ofrabbits. After4and12weeks implantation, macroscopic evaluation, histologicalevaluation, and micro-CT analysis were performed.Results:1. The setting times of CPC-BG were prolonged as the content of BG increased,increased from21min to44min, with the weight ratio of BG varied from10%to40%at P/L ratio of2.0g/ml. The injectability of CPC-BG composite paste was significantlyimproved compared with injectability of CPC paste (p <0.05);2. XRD showed that main peaks for HA of hardened CPC-BG composite, were notobviously altered and peaks for Ca2SiO4and Ca3SiO5could be seen in the XRDpatterns of CPC-BG composite with10%and20%BG. The SEM micrographs for thecross section of the CPC and CPC-BG composite specimens (10%,20%) showed thatthe CPC-BG composite specimens closely combined with each other which showedmore compact microstructure than CPC. The compressive strength of CPC-BGcomposites rose with an increase in the weight ratio of BG and there were significantdifferences between CPC and CPC-20%BG composite specimens at day1and day7(p<0.05). After immersing in simulated body fluid (SBF) for7and14days, the amountof apatite aggregates on CPC-BG composite surface were larger and apatite layersgrew more densely on CPC-BG composite surface than amount on CPC surface. EDSindicated that the surface of CPC-BG composite (10%,20%) consisted of a calciumphosphate with a Ca/P ratio of about1.56and1.53and contained Si, while the surfaceof CPC consisted of a calcium phosphate with a Ca/P ratio of about1.67, contained noSi. The degradation rates of all CPC-BG composite specimens were significantlyhigher than degradation rates of CPC specimens (p <0.05);3. The degree of cell attachment, proliferation and differentiation was increased byadding BG into CPC. The MTT assay showed that the OD values of CPC-BG composite specimens (20%) were significantly higher than OD values of CPCspecimens (p <0.05) in a period of8h. OD values of CPC-BG composite specimens(20%) were significantly higher than OD values of CPC specimens at4and7days (p<0.05), indicating that CPC-BG composite specimens promoted cell growth andfacilitated proliferation with no cytotoxic effect on cells compared with CPCspecimens. The ALP activity of cells cultured on CPC-BG composite (20%) wassignificantly higher than ALP activity on CPC and TCPS control (p <0.05) at4and7days;4. With the prolonged implantation period from4to12weeks, the in vivo degradationrate and the amount of newly formed bone of CPC-BG composites and CPC wereincreased. Moreover, the in vivo degradation rate and the amount of newly formedbone of CPC-BG composites were significantly higher that of CPC specimens (p <0.05).Conclusions:1. The setting time and injectablity of CPC-BG composite (10%,20%) conform to thesurgical requirement. CPC-BG（10%，20%） possessed a retarded setting time andmarkedly better injectability and mechanical properties than CPC. Meanwhile,CPC-BG samples showed significantly improved in vitro degradability and bioactivitycompared to CPC in SBF.2. Adding BG into CPC is benefit for OB to adhere, proliferate and differentiate.3. CPC-BG with excellent biocompatibility, improved bioactivity and in vivodegradability enhanced the efficiency of new bone formation in comparison with CPC,which exhibits promising prospects for bone regeneration.