摘要
在骨组织工程领域中,生长因子的调控对于促进骨组织的再生和血管化具有非常重要的意义,但外源性生长因子的不当使用所带来的组织过度增生及血管畸形等不良反应亦需给予高度的重视。经成骨定向诱导的骨髓间充质干细胞(BMSCs)的胞外基质(ECM)富含其自身表达的成骨相关生长因子,适应于BMSCs的成骨分化、基质矿化和血管生成。本研究针对BMSCs胞外基质在骨组织工程的应用,进行免疫原性去除、活性成分保留及ECM在组织工程支架有效涂装等关键问题的研究。以地塞米松/β-甘油磷酸钠/维生素C对BMSCs进行成骨化诱导并获取其在成骨分化过程中分泌的ECM,通过反复冻融法和DNA酶酶解法去除ECM中残留的细胞和遗传物质。ELISA测试结果表明,获取的脱细胞胞外基质(dECM)中COL-I为230.89 ng/mg、BMP-2为10.047 ng/mg、VEGF为10.857 ng/mg、OC为16.866 ng/mg。采用溶剂浇铸/颗粒滤沥法制备孔径32~48 μm,孔隙率为80%的聚乳酸–羟基乙酸共聚物(PLGA)及聚己内酯/聚乙醇酸(PGA/PCL)三维多孔支架,将dECM乙酸分散液通过真空浸渍–乙酸蒸发工艺对三维多孔支架进行涂装。碱性磷酸酶(ALP)活性测定和茜素红矿化结节染色等结果表明,dECM涂装的三维多孔支架具有明显的成骨促进效应。
In the field of bone tissue engineering, the regulation of growth factor (GFs) is of great significance in promoting bone tissue regeneration and vascularization, but excessive tissue hyperplasia and vascular malformation caused by improper use of exogenous growth factors should be paid more attention. The extracellular matrix (ECM) of osteogenic-induced bone marrow mesenchymal stem cells (BMSCs) is rich in self-expressed osteogenic growth factors and is suitable for osteogenic dif-ferentiation, stromal mineralization and angiogenesis of BMSCs. Focusing on the ECM application in bone tissue engineering, some key issues, such as immunogenicity removal, active component retention, and effective coating on scaffolds were studied in this study. BMSCs were osteogenically induction with DXMS/β-sodium glycerophosphate (β-GP)/vitamin C, and the ECM secreted by BMSCs during osteogenic differentiation was collected. Then the residual cells and genetic components were removed by the repeated freezing-thawing and the DNase hydrolysis. ELISA test results showed that the decellularized extracellular matrix (dECM) still retained the moderate amount of active components with the COL-I of 230.89 ng/mg, the BMP-2 of 10.047 ng/mg, the VEGF of 10.857 ng/mg, and the OC of 16.866 ng/mg. Polylactic acid-glycolic acid copolymer (PLGA) and polycaprolactone/polyglycolic acid (PGA/PCL) three-dimensional porous scaffolds with pore diameter of 32 - 48 μm and porosity of 80% were prepared by solvent casting/particle filtration. DECM acetic acid dispersion was coated on the three-dimensional porous scaffold by vacuum im-pregnation-acetic acid evaporation process. The results of alkaline phosphatase (ALP) activity de-termination and alizarin red mineralized nodule staining showed that the three-dimensional po-rous scaffolds with dECM coating exhibited the obvious osteogenic promotion effect.
出处
《材料科学》
CAS
2020年第7期570-580,共11页
Material Sciences
关键词
骨髓间充质干细胞
胞外基质
脱细胞化
DNA酶解
生长因子
Bone Marrow Mesenchymal Stem Cells
Extracellular Matrix
Decellularization
DNA Enzymolysis
Growth Factors
