N-乙酰半胱氨酸丝素微球复合磷酸钙骨水泥的制备及表征

摘要

BACKGROUND:Calcium phosphate bone cement has been applied to clinical surgery because of its good biocompatibility and osteoconduction. However poor mechanical properties and lack of osteoinductivity limit its wide application. OBJECTIVE:To develop calcium phosphate cement incorporated with N-acetylcysteine (NAC) loaded silk fibroin microspheres (SFM), which is a kind of new injectable bone graft material with slow-release function, and evaluate its physical and chemical properties and cel compatibility. METHODS: Empty SFMs were prepared with emulsion solvent evaporation to absorb NAC solution of different concentrations by NAC-SFM and the concentration of NAC at the maximum drug loading ratio was determined. Then, NAC-SFM was loaded into calcium phosphate bone cement to test the drug release propertiesin vitro. MC3T3-E1 osteoblasts were cultured on the surface of NAC-SFM calcium phosphate bone cement and cel attachment and growth were observed by scanning electron microscope. Additionaly, MC3T3-E1 cels were cultured with three kinds of bone cement extracts (calcium phosphate cement, SFM-calcium phosphate cement, NAC-SFM-calcium phosphate cement, as wel as cultured in theα-minimum essential medium containing a volume fraction of 10% fetal bovine serum and 1% penicilin-streptomycin double antibody as the control. MTS assay was used to evaluate cel proliferation. RESULTS AND CONCLUSION:Microspheres in the composite bone cement presented with smooth surface, same size, diffused distribution and no obvious destroy. Thus, the SFM could remain stable in the reaction process of the composite bone cement. The double slow release system which contained silk fibroin microspheres and calcium phosphate bone cement showed a significant decrease in the cumulative release percentage of NAC within the first 24 hours compared with the control group (P < 0.05). In the next 28 days, the release speed of NAC was significantly lower in the NAC-SFM-calcium phosphate cement group than the calcium phosphate cement group (P< 0.05). In addition, different extracts had no significant cytotoxicity to the growth of MC3TC-E1 cels. Thus, the NAC-SFM-calcium phosphate cement has good cytocompatibility, which provide a new insight into the development of bone repair biomaterials.%背景：磷酸钙骨水泥因其良好的生物相容性及骨传导性已被用于临床，但因其力学性能较差以及骨诱导性的欠缺限制了其广泛应用。　　目的：制备复合N-乙酰半胱氨酸丝素微球的注射性磷酸钙骨水泥，构建一种具有药物缓释功能的新型注射性植骨材料，并评价其理化性能及细胞相容性。　　方法：采用乳化溶剂蒸发法制备空丝素微球，用丝素微球吸附不同浓度N-乙酰半胱氨酸溶液，确定最大理论载药率时N-乙酰半胱氨酸浓度，再将载有N-乙酰半胱氨酸的丝素微球载入磷酸钙骨水泥中，检测负载N-乙酰半胱氨酸丝素微球的磷酸钙骨水泥的体外释药性能；体外将小鼠MC3T3-E1成骨细胞与N-乙酰半胱氨酸丝素微球磷酸钙骨水泥共培养，扫描电镜观察细胞在材料表面的贴附及生长。将单纯磷酸钙、丝素微球磷酸钙、N-乙酰半胱氨酸丝素微球磷酸钙骨水泥共3种骨水泥浸提液，以及含有体积分数10%胎牛血清和1%青霉素-链霉素的α-MEM培养基，与MC3T3-E1细胞共培养，对照组仅用上述培养基培养，采用MTS法评估各组细胞增殖情况。　　结果与结论：①复合骨水泥中的微球表面光滑平整，大小基本均一，散在分布，球体未见明显破坏，可见在制备复合骨水泥的过程中微球能保持稳定，不被破坏；②采用丝素微球、磷酸钙骨水泥作为药物双重缓释体系在前24 h累计释药百分比较对照组明显减少(P <0.05)，后续28 d中N-乙酰半胱氨酸丝素微球磷酸钙骨水泥组释药速度也较单纯磷酸钙骨水泥显著降低(P <0.05)；③实验中各组材料浸提液对MC3T3-E1细胞的生长均无明显影响，无明显细胞毒性；④结果说明：N-乙酰半胱氨酸丝素微球复合磷酸钙骨水泥材料具备良好的细胞相容性，有望为骨修复生物材料的研究提供参考。