| Introduction:The bone fracture caused by many reasons, such as aging, disease and trauma,is common in clinic. For a long time, the metallic materials are the traditionalfracture internal fixation for their high mechanics strength. But previous studieshave shown that metallic materials did not provide the optimum therapy forinternal fixation due to their shortcomings such as stress shielding duringpost-healing, chronic inflammation caused by corrosion, fatigue and loosening ofthe implant. Bioabsorbable materials for internal fixation have become a focus ofmany researchers in the past 30 years because of their incomparable advantages tometal materials in therapy of bone fracture. However, they can only be used forinternal fixation of non weight-bearing bones for their low initial mechanicalquality and rapid early period degradation, which originate from their materialand technical characteristics. Therefore, the researches on the biodegradable andbioabsorbable materials with high initial mechanical quality used for internalfixation of bone fracture are the hot topics focused by many scientists. Poly-L-lactide (PLLA) is a kind of polymer material with wellbiocompatibility and biodegradation. However, there are still many defects inPLLA material prepared for internal fixation, such as complicated fabricatingprocesses, expensive price, and slowly intension degradation. Moreover, aciddegradation products of PLLA might result in aseptic inflammation of tissue.β-calcium metaphosphate (β-CMP), Ca(PO3)2, similar to the content of naturebone, is a kind of bioactive ceramic with high strength, good biologicalcompatibility. However,β-CMP couldn't be used for fracture fixationindependently for its characteristics of ceramics such as brittleness and stiffness,which limits its application in clinic. To meet the demand for mechanical andbiological properties of internal-fracture-fixation materials, more and moreattentions have been paid to combining PLLA withβ-CMP.In this study,β-calcium metaphosphate whiskers were prepared by controlledcrystallization in the glass andβ-CMP whiskers were used to reinforce themechanical properties of PLLA in preparation of PLLA/β-CMP composites forfracture internal fixation. The mechanical properties, bioactivity andbiocompatibility ofβ-CMP whiskers and PLLA/β-CMP composites wereevaluated.Methods:1.β-calcium metaphosphate whiskers had been prepared by controlledcrystallization in the glass using raw materials such as Ca(H2PO4)2, CaCO3 andH3PO4 (85％liquid). The factors influencing the morphology of theβ-CMPwhiskers such as component, time of crystallization treatment, washing time withNaOH solution and deionized water were discussed. DTA was used to determinethe temperature of crystallization treatment. The crystal phase and themorphology of the samples were analyzed by means of XRD and SEM. Phospharidylcholine, a kind of biosurfactant, was chosen to modifyβ-CMPwhiskers in order to improve the hydrophobic property.2. Degradation kinetics in vitro ofβ-CMP whiskers in physiological salineunder the static state soak and kinetic circuit of solution had been studied. Thekinetic model equation and activation energy of degradation ofβ-CMP whiskershad been simulated according to the degradation data. The cytocompatibility wasevaluated by the effect ofβ-CMP on osteoblasts adhesion and proliferation assayby means of MTT analysis and SEM image.3. A simple and economic method, two-step compression molding method(two-step method), was developed to prepare PLLA/β-CMP fracture fixationcomposite. After PLLA andβ-CMP were homogeneously mixed and placed in adisk mold, the mixture was pressed at a low-temperature and a high-temperature,respectively. The low-temperature and high-temperature were determined by theglass transition temperature and melting-out temperature of PLLA measured byDSC analysis. The composites were analyzed by scanning electron microscope.The bending strength (σb) and Young's modulus (E) were performed on theuniversal mechanical testing machine.4. The degradation of PLLA/β-CMP composite has been studied byimmersion experiments with SBF at 37℃.5. The biological properties of PLLA/β-CMP composite were evaluated bycytology and animal experiments. After seeded and cultured with osteoblasts,osteoblasts proliferation and adhere on PLLA/β-CMP composite were analyzedby MTT and SEM, respectively. The animal safety assessments of PLLA/β-CMPcomposite were carried out, including acute toxicity tests on mice, pyrogen testson rabbits, hemolysis tests and muscle implant tests. Results:1.β-calcium metaphosphate whiskers with high aspect ratios (＞30) anddiameters of 0.2～2μm had been prepared by controlled crystallization from theglass phase. Results showed that high quality ofβ-calcium metaphosphatewhiskers could be obtained by the best CaO: P2O5 ratio of 0.85, crystallizationtreating for 36 hours and washing with 0.20 mol·L-1NaOH solution and deionizedwater for 48 hours, respectively. The testing results of contact-angle indicated thatthe hydrophobic property of whiskers modified with phospharidylcholine wasbetter than unmodified.2. The results of degradation in vitro ofβ-CMP whiskers in physiologicalsaline showed that the degradation rate ofβ-CMP under dynamic solution wasmuch higher than under the static immersion, due to its higher driving force ofdegradation.According to the degradation data, it was simulated that the degradation ofβ-CMP accorded with Avrami kinetic model equation: -ln(1-x)=ktn, and itsactivation energy (Ea) was 26.28kJ·mol-1.The results of MTT analysis and SEM images showed thatβ-CMP couldpromote cells growth, osteoblasts adhered and spread well on the material. Such asuperior characteristic indicated that theβ-CMP particles could contact directlywith cells/tissues to stimulate the cell proliferation. In addition, the numbers ofosteoblasts adhered on the material proliferated with the increase of culture time,indicating thatβ-CMP had good in vitro biocompatibility ofβ-CMP.3. PLLA/β-CMP fracture fixation composites with high mechanicalproperties were prepared by a two-step method. The bending strength andYoung's modulus of composites increased with the increase ofβ-CMP weightcontent and reached a maximum of 126.33 MPa and 4.36 GPa at aβ-CMP contentof 25wt%, which are close to those of the cortical bone and can meet the request of internal fixation for human bone fractures. At the process of compressionmolding, the temperatures of low-temperature and high-temperature molding were80℃(a little higher than Tg of PLEA) and 190℃(a little higher than Tm ofPLEA).4. The results of degradation experiments indicated that the degradation rateof composite increased with the increase ofβ-CMP weight content. The bendingstrength of PLLA/β-CMP composite with 25wt%β-CMP content was maintainedat 110.15MPa (approximately 92％of the initial values) after degradation for 12weeks, which can meet the request of internal fixation for human bone fractures.5. The results of cytology experiments showed that PLLA/β-CMPcomposite with high weight content ofβ-CMP could promote cells growth andosteoblasts adhered and spread well on the materials. Furthermore, the osteoblastsproliferation increased significantly with the increase ofβ-CMP weight content incomposite. These results indicate that PLLA/β-CMP composite has a good invitro biocompatibility and can meet the request of internal fixation.6. The results of acute toxicity tests on mice and pyrogen tests on rabbitsshowed that PLLA/β-CMP composite had not any acute toxic effects and couldmeet the accepted criteria for the pyrogen test. The hemolysis rate of compositewas 1.46％, i.e. less than 5％. There was no obvious inflammation after thePLLA/β-CMP composite was implanted in muscle of rabbit for 90d, indicated thatthe composite had good in vitro histocompatibility. |
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