关节骨软骨仿生支架浸提液对种子细胞生物学特性的影响

Effects of the extract solution of bionic articular osteochondral scaffold on the biological characteristics of seed cells

为了观测自制胶原涂层羟基磷灰石纳米晶/聚磷酸钙纤维/L-聚乳酸基含“界层结构”仿生一体化关节骨软骨复合组织工程支架材料对种子细胞增殖活性及相关分化功能的影响,采用CCK-8、流式细胞仪检测及体外分化诱导培养,分别检测新型支架材料浸提液对人骨髓间充质干细胞(hBMSCs)及人脐带间充质干细胞(hUCMSCs)的增殖活性、凋亡及成骨、成软骨分化能力的影响;通过实时荧光定量PCR和Western blot检测细胞成骨及成软骨分化诱导后ALP、Osterix、Runx2、COL1A1及COL2A1的mRNA和蛋白表达水平.结果显示,新型支架材料浸提液对hBMSCs及hUCMSCs的增殖活性及凋亡均无显著影响;体外分化诱导后均可呈现成骨及成软骨分化表型,5种基因的mRNA和蛋白表达均显著上调;支架材料浸提液处理后,其成骨及成软骨分化能力更明显,5种基因的mRNA和蛋白表达上调更显著.提示自制新型仿生一体化支架材料可保持hBMSCs及hUCMSCs种子细胞良好的增殖活性及成骨、成软骨分化功能.

To investigate the effects of collagen-coated nano-hydroxyapatite/calcium polyphosphate fibers/poly-L-lactic acid bionic integrated articular bone and cartilage composite tissue engineered scaffold material with a boundary layer structure on the growth,proliferation and related differentiation function of seed cells in vitro.We examined the effects of novel scaffold material extract solution on the proliferation,apoptosis and osteogenic or chondrogenic differentiation capacity of human bone marrow mesenchymal stem cells(hBMSCs)and human umbilical cord mesenchymal stem cells(hUCMSCs)using cell counting kit-8,flow cytometry,and differentiation induction cultures.The mRNA and protein expression levels of alkaline phosphatase(ALP),Osterix,Runx2,COL1A1 and COL2A1 in two kinds of mesenchymal stem cells were measured by real-time fluorescence quantitative PCR and Western blot after osteogenic and chondrogenic differentiation induction.The results indicated that there were no significant effects on the proliferation and apoptosis of hBMSCs and hUCMSCs after treatment with the novel scaffold material extract solution.hBMSCs and hUCMSCs showed osteogenic or chondrogenic differentiation phenotypes after the induction of osteogenic or chondrogenic differentiation in vitro.The mRNA and protein expression of ALP,Osterix,Runx2,COL1A1 and COL2A1 were significantly upregulated.The differentiation of osteogenesis and chondrogenesis were more obvious when cells were treated with the scaffold material extract solution,and the mRNA and protein expression of ALP,Osterix,Runx2,COL1A1 and COL2A1 were upregulated more significantly.The results suggested that the selfmade novel bionic integrated scaffolds could maintain the proliferative viability and the osteogenic and chondrogenic differentiation functions of both hBMSCs and hUCMSCs in vitro.