The Study Of Biodegradable Thermosensitive Hydrogel Composite Calcium-silicon Based Bioactive Bone Cement

Severe osteoporotic low back pain and osteoporotic vertebral compression fractures（OVCF）have become a global disease affecting the lives and health of the elderly.Clinically,this is mainly treated by vertebroplasty（PVP）.Calcium silicate cement（CSC）has excellent properties in biocompatibility,bone conduction and induced osteogenesis.It has broad prospects in the field of vertebroplasty and bone defect filling materials.However,there are still some problems in the preliminary application,including low compressive strength,poor injectability,poor collapse resistance and so on.Based on the biomechanical characteristics of osteoporotic vertebral body,this subject combines dicalcium silicate,tricalcium silicate and hemihydrate gypsum to design a composite calcium-silica-based bone cement,which is intended to solve the problems of mismatch between its elastic modulus and cancellous bone,poor injectability and unsatisfactorycolllapsibility.By using the characteristics of the poly（lactic co-glycolic acid）polyethylene glycol poly（lactic acid glycolic acid）（poly（lacticacid）,poly（Glycolicacid）andpoly（ethyleneglycol）,PLGA-PEG-PLGA）tri-block polymer temperature-sensitive hydrogel,PLGA-PEG-PLGA hydrogel was used as a curing agent and mixed with calcium silicon based bone cement powder.In this way,polyethylene glycol poly（lactic co glycolic acid）copolymer thermosensitive hydrogel three calcium silicate/calcium silicate two/gypsum（PLGA-PEG-PLGA/C₃S/C₂S/POP）composite cement was prepared.The bone cement made in this project has significantly improved injectability,good performance in inducing autologous osteogenesis and anti-collapsibility,and the compressive strength and elastic modulus match with human cancellous bone,so it has certain application value in bone defect repair and vertebroplasty operation.Objectives:To design the preparation method of PLGA-PEG-PLGA/C3S/C2S/POP composite system bone cement.To compare the effects of different proportion of temperature-sensitive hydrogel content on the curing time,compressive strength,degradability,injectability,anti-collapse,cytocompatibility,and induced osteogenesis of the composite bone cement.Animal tests were performed to analyze the safety,efficacy and bioactivity of the cement after defining the optimal protocol.Methods:（1）Preparation of bone cement:tricalcium silicate was prepared by sol-gel method.Preparation of dicalcium silicate powder by coprecipitation method.The calcium sulfate dihydrate was dried and dehydrated to obtain hemihydrate gypsum.PLGA-PEG-PLGA triblock polymers were extracted by ring-opening polymerization using polyethylene glycol as initiator.Various concentrations of hydrogels were used for the liquid phase of the bone cement,and the PLGA-PEG-PLGA/C₃S/C₂S/POP composite bone cement was finally obtained.（2）Determination of physical and chemical properties:the hydration products of bone cement were studied by X-ray diffraction;the cement curing time was tested using a Vicat apparatus;When judging the injectability of cement,the ratio between the mass of bone cement slurry extruded from the syringe and the total mass was mainly used;the compressive strength was tested by a mechanical testing machine;to detect the anti-collapse property of cement,the cement slurry was added to the simulated body fluid and the appearance of loosening was observed after shaking.After shaking,observe whether loosening occurs.（3）In vitro cell test:The in vitro cytotoxicity of cement was measured by CCK8 test;the osteogenic properties were determined by alkaline phosphatase activity and alizarin red staining.（4）Animal experimental study:20 healthy female New Zealand rabbits were selected as research objects,and firstly,bilateral ovaries were surgically removed for classical osteoporosis modeling.After 12 weeks,2osteoporosis models were randomly selected to test the results.The remaining 18 rabbits were divided into 3 groups A,B and C,and performed rabbit femoral condylar bone defect filling.Group A was implanted with 30%PLGA-PEG-PLGA/C₃S/C₂S/POP composite bone cement with good physical and chemical functions and in vitro tests at the early stage of implantation.Group B was PMMA bone cement control group.Group C was the blank control group.The distribution of bone cement,degradability and new bone tissue were examined by Micro-CT and gross anatomical observation at 6 and 12weeks after surgery.Results:（1）According to the phase analysis by X-ray diffractometer,the composition of24-hour hydration products of PLGA-PEG-PLGA/C₃S/C₂S/POP composite bone cement included calcium hydroxide,calcium silicate hydrate,gypsum dihydrate,and insufficiently reacted tricalcium silicate and dicalcium silicate.The addition of temperature-sensitive hydrogel did not affect the hydration results.In terms of compressive strength,the hydrogel composite bone cement was improved to some extent.The average compressive strength of 30%PLGA-PEG-PLGA/C₃S/C₂S/POP at day 14was 45.56±6.56MPa,which was significantly different from that of bone cement without hydrogel（P<0.05）.The addition of polymer significantly improved the injectability of the cement,and the injectability coefficient of 30%PLGA-PEG-PLGA/C₃S/C₂S/POP composite bone cement reached 85.93±3.54%,with significantly improved performance（p<0.05）;at the 10th week of curing,the degradation rates of 30%and 40%PLGA-PEG-PLGA groups were 46.32±4.42%and 48.32±5.16%,which was significantly faster degradation rate than that of C₃S/C₂S/POP bone cement alone（p<0.05）;the anti-collapse properties of all groups with different proportions of hydrogel added were improved.（3）According to the test data of cell proliferation viability by CCK-8colorimetric method,different proportions of hydrogel bone cement showed good performance in primary osteoblast proliferation,without cytotoxicity.According to the test data of alkaline phosphatase activity and alizarin red staining,30%PLGA-PEG-PLGA/C₃S/C₂S/POP had the best induced osteogenicity.（4）The model of rabbit osteoporosis was established by ovariectomization,and the trabecular bone of the distal femur was sparse at 12 weeks after the operation;different materials were used to fill the distal femoral bone defect in rabbits,12 weeks after the operation,imaging and anatomical results showed that the cement partially degraded and new bone appeared near the filled area.Conclusions:When PLGA-PEG-PLGA tri-block temperature-sensitive hydrogel was added to C₃S/C₂S/POP calcium-silica-based bone cement,it showed good performance in injectability,failure resistance,compressive strength matching with cancellous bone strength,as well as the induced mineralization of calcium-silica-based bone cement,and the promotion of biological activity on bone cell proliferation and differentiation.Animal experiments showed that 30%PLGA-PEG-PLGA/C₃S/C₂S/POP composite bone cement has good operability,safety and effectiveness.