肾上腺髓质素微球复合PLGA/纳米羟基磷灰石支架材料的制备及生物活性评价

Preparation and Bioactivities of PLGA/Nano-hydroxyapatite Scaffold Containing Chitosan Microspheres for Controlled Delivery of Mutifuncational Peptide-adrenomedullin

利用离子乳化交联法制备了负载肾上腺髓质素的壳聚糖微球,应用热致相分离法制备了乳酸和乙醇酸共聚物/纳米羟基磷灰石(PLGA/nHA)支架材料并在其中包覆载药微球.通过扫描电子显微镜、体外释放行为、材料溶血行为、碱性磷酸酶(ALP)活性的测定、支架材料表面细胞荧光染色和MTT[3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐]比色法等手段综合评价载药支架材料的性能及生物活性.结果表明,微球直径均匀,载药支架孔径大小合适并相互穿通.支架材料的溶血率小于5%,符合医用材料的溶血实验要求.载药支架及支架材料本身对成骨细胞及血管内皮细胞的增殖以及成骨细胞的分化均有一定的促进作用.

By tripolyphosphate（TPP） as cross-linker,chitosan microspheres（CMs） loaded with adrenomedullin（ADM） were prepared by an emulsion-ionic cross-linking method.Then,porous poly（lactide-co-glycolide）（PLGA）/nano-hydroxyapatite（nHA）/CMs scaffold was developed by thermally induced phase separation.Scanning electron microscope was used to analyze the characterization of the microspheres and the scaffold and the drug release from them was tested in vitro.Also,the hemolytic effect of the scaffold was tested to evaluate the toxicity of the biomaterial.MG63 cell line and HUVEC cell line were cultured for testing the bioactivity of the scaffold loading ADM.After the cells adhered on the scaffold,the cell proliferation and differentiation were analyzed by methyl thiazolyl tetrazolium（MTT） assay,fluorescent staining and alkaline phosphatase（ALP） assay.The results show that the diameter of the microspheres is well-distributed and the PLGA/nHA/CMs scaffold shows a typical microstructure of polymeric foam.The pores of the scaffold ranged from 30 μm to 220 μm and they were interconnected.The hemolytic rate of the biomaterial was less than 5%,which was in line with national standard of medical materials.This study confirmed that PLGA/nHA/CMs/ADM and PLGA/nHA/CMs scaffold could accelerate the proliferation of osteoblast-like cells and vascular endothelial cells and also could stimulate the differentiation of osteoblast-like cells.