骨髓基质干细胞片层制备与内皮祖细胞分离培养的研究

发布时间：2018-06-14 00:26

  本文选题：骨髓基质干细胞 + 内皮祖细胞　； 参考：《青岛大学》2010年硕士论文

【摘要】： 骨髓基质干细胞片层制备与内皮祖细胞分离培养的研究 目的：体外分离培养犬骨髓间充质干细胞(Bone marrow stem cells BMSCs)纯化、扩增后向成骨细胞诱导分化,对其成骨和矿化能力等生物学特性进行检测。体外分离培养犬内皮祖细胞(endothelial progenitor cells EPCs)并向内皮细胞诱导分化。应用温度反应性培养皿收获骨髓基质干细胞形成完整的细胞片层。制备同种异体的冻干骨(demineralized bone matrix DBM)支架。将种子细胞、支架和生长因子三要素结合,以期构建血管化的大块组织工程骨。 方法：密度梯度离心法分离培养犬骨髓基质干细胞,向成骨细胞方向诱导分化后,倒置显微镜和扫描电镜下进行形态学观察；将诱导后的BMSCs接种至温度反应性培养皿中,37℃、5%CO2饱和湿度培养,然后降温至20℃制备BMSCs细胞片层；密度梯度离心法分离培养内皮祖细胞,并用血管内皮生长因子等向内皮细胞诱导分化；制备冻干骨支架；制备BMSCs细胞片层/BMSCs/EPCs/DBM复合体,包裹犬的胸背动静脉成骨。 结果： 1.原代培养的骨髓基质干细胞呈长梭形,漩涡状生长。 2.向成骨细胞诱导后的BMSCs呈卵圆形或多角形,有较强的成骨能力。 3.当温度降至20℃时,BMSCs从温度反应性培养皿上完全脱落,扫描电镜下可见富含细胞外基质的完整的细胞片层。 4.原代培养的内皮祖细胞呈小圆形,向内皮祖细胞诱导后呈纺锤形。 结论： 1.应用密度梯度离心法和贴壁法可获取内皮祖细胞并向内皮细胞诱导； 2.应用温度反应性培养皿能获取完整的BMSCs细胞片层。并用于功能性成骨； 3.将BMSCs片层、BMSCs、EPCs和DBM制备成复合体植入犬体内有望形成血管化的组织工程骨。
[Abstract]:Preparation of Bone Marrow Mesenchymal Stem cells and isolation and Culture of Endothelial progenitor cells objective: to purify bone marrow mesenchymal stem cells (BMSCs) from canine bone marrow in vitro and to differentiate into osteoblasts. The biological characteristics such as osteogenesis and mineralization ability were examined. Canine endothelial progenitor cells (ECs) were isolated and cultured in vitro and differentiated into endothelial cells. Bone marrow mesenchymal stem cells (BMSCs) were harvested in a temperature-reactive culture dish to form a complete layer of cells. The allogeneic freeze-dried bone demineralized bone matrix DBM scaffolds were prepared. Seed cells, scaffolds and growth factors were combined to construct vascularized large tissue engineered bone. Methods: canine bone marrow stromal cells were isolated and cultured by density gradient centrifugation. After differentiation into osteoblasts, morphology was observed under inverted microscope and scanning electron microscope. The induced BMSCs were inoculated into the temperature-reactive culture dish and incubated with CO2 saturation humidity at 37 鈩，

本文编号：2016194