不同种类聚合物对猪小肠黏膜下层支架仿生矿化的影响

Effects of different polymers on biomimetic mineralization of small intestine submucosal scaffolds

目的:探究不同种类的聚合物对脱细胞猪小肠黏膜下层(small intestinal submucosa,SIS)支架体外仿生矿化的影响,并基于理化性能和生物相容性指标评价各组SIS矿化支架。方法:将冷冻干燥法制备而成的SIS支架分别浸泡在模拟体液(simulated body fluid,SBF)、含聚丙烯酸(polyacrylic acid,PAA)的矿化液和同时含PAA和聚天冬氨酸(polyaspartic acid,PASP)的矿化液中持续2周,隔天换液,依次得到SBF@SIS、PAA@SIS、PAA/PASP@SIS支架,以未矿化的SIS支架为对照组,评价上述支架的理化性能及生物相容性。结果:环境扫描电子显微镜(environment scanning electron microscopy,ESEM)下各组支架均呈适宜孔径的三维多孔结构,各组矿化支架均可见晶体附着,其中以PAA/PASP@SIS支架晶体沉积更为规则,同时可见胶原纤维增粗。能谱分析显示,3组矿化支架均可见钙、磷元素的特征峰,以PAA/PASP@SIS支架峰值最高;傅里叶变换红外光谱分析证实,3组矿化支架均实现了羟基磷灰石与SIS结合;各组支架均具有良好的亲水性;3组矿化支架压缩强度均高于对照组,以PAA/PASP@SIS支架压缩强度最优;各组支架均能有效吸附蛋白,以PAA/PASP@SIS组吸附能力最佳。CCK-8细胞增殖实验(周期为1、3、5、7、9 d)中,PAA/PASP@SIS组展现出最佳的促细胞增殖能力。结论:经同时含PAA和PASP的矿化液制备的PAA/PASP@SIS支架与其他矿化支架相比具有更优的理化性能和生物相容性,具有骨组织工程应用的潜能。

Objective:To explore the effects of different polymers on in vitro biomimetic mineralization of small intestinal submucosa(SIS)scaffolds,and to evaluate the physicochemical properties and biocompatibility of the SIS scaffolds.Methods:The SIS scaffolds prepared by freeze-drying method were immersed in simulated body fluid(SBF),mineralized liquid containing polyacrylic acid(PAA)and mine-ralized liquid containing PAA and polyaspartic acid(PASP).After two weeks in the mineralized solution,the liquid was changed every other day.SBF@SIS,PAA@SIS,PAA/PASP@SIS scaffolds were obtained.The SIS scaffolds were used as control group to evaluate their physicochemical properties and biocompatibility.We observed the bulk morphology of the scaffolds in each group,analyzed the microscopic morphology by environment scanning electron microscopy and determined the porosity and pore size.We also analyzed the surface elements by energy dispersive X-ray spectroscopy(EDX),analyzed the structure of functional groups by Flourier transformed infrared spectroscopy(FTIR),detected the water absorption rate by using specific gravity method,and evaluated the compression strength by universal mechanical testing machine.The pro-cell proliferation effect of each group of scaffolds were evaluated by CCK-8 cell proliferation method.Results:Under scanning electron microscopy,the scaffolds of each group showed a three-dimensional porous structure with suitable pore size and porosity,and crystal was observed in all the mineralized scaffolds of each group,in which the crystal deposition of PAA/PASP@SIS scaffolds was more regular.At the same time,the collagen fibers could be seen to thicken.EDX analysis showed that the characteristic peaks of Ca and P were found in the three groups of mineralized scaffolds,and the highest peaks were found in the PAA/PASP@SIS scaffolds.FTIR analysis proved that all the three groups of mineralized scaffolds were able to combine hydroxyapatite with SIS.All the scaffolds had good hydrophilicity.The compressive strength of the mineralized scaffold in the three groups was higher than that in the control group,and the best compressive strength was found in PAA/PASP@SIS scaffold.The scaffolds of all the groups could effectively adsorb proteins,and PAA/PASP@SIS group had the best adsorption capacity.In the CCK-8 cell proliferation experiment,the PAA/PASP@SIS scaffold showed the best ability to promote cell proliferation with the largest number of living cells observed.Conclusion:Compared with other mineralized scaffolds,PAA/PASP@SIS scaffolds prepared by mineralized solution containing both PAA and PASP have better physicochemical properties and biocompatibility and have potential applications in bone tissue engineering.