發(fā)布時間：2018-06-14 00:26

  本文選題：骨髓基質(zhì)干細胞 + 內(nèi)皮祖細胞��； 參考：《青島大學(xué)》2010年碩士論文

【摘要】： 骨髓基質(zhì)干細胞片層制備與內(nèi)皮祖細胞分離培養(yǎng)的研究 目的：體外分離培養(yǎng)犬骨髓間充質(zhì)干細胞(Bone marrow stem cells BMSCs)純化、擴增后向成骨細胞誘導(dǎo)分化,對其成骨和礦化能力等生物學(xué)特性進行檢測。體外分離培養(yǎng)犬內(nèi)皮祖細胞(endothelial progenitor cells EPCs)并向內(nèi)皮細胞誘導(dǎo)分化。應(yīng)用溫度反應(yīng)性培養(yǎng)皿收獲骨髓基質(zhì)干細胞形成完整的細胞片層。制備同種異體的凍干骨(demineralized bone matrix DBM)支架。將種子細胞、支架和生長因子三要素結(jié)合,以期構(gòu)建血管化的大塊組織工程骨。 方法：密度梯度離心法分離培養(yǎng)犬骨髓基質(zhì)干細胞,向成骨細胞方向誘導(dǎo)分化后,倒置顯微鏡和掃描電鏡下進行形態(tài)學(xué)觀察；將誘導(dǎo)后的BMSCs接種至溫度反應(yīng)性培養(yǎng)皿中,37℃、5%CO2飽和濕度培養(yǎng),然后降溫至20℃制備BMSCs細胞片層；密度梯度離心法分離培養(yǎng)內(nèi)皮祖細胞,并用血管內(nèi)皮生長因子等向內(nèi)皮細胞誘導(dǎo)分化；制備凍干骨支架；制備BMSCs細胞片層/BMSCs/EPCs/DBM復(fù)合體,包裹犬的胸背動靜脈成骨。 結(jié)果： 1.原代培養(yǎng)的骨髓基質(zhì)干細胞呈長梭形,漩渦狀生長。 2.向成骨細胞誘導(dǎo)后的BMSCs呈卵圓形或多角形,有較強的成骨能力。 3.當(dāng)溫度降至20℃時,BMSCs從溫度反應(yīng)性培養(yǎng)皿上完全脫落,掃描電鏡下可見富含細胞外基質(zhì)的完整的細胞片層。 4.原代培養(yǎng)的內(nèi)皮祖細胞呈小圓形,向內(nèi)皮祖細胞誘導(dǎo)后呈紡錘形。 結(jié)論： 1.應(yīng)用密度梯度離心法和貼壁法可獲取內(nèi)皮祖細胞并向內(nèi)皮細胞誘導(dǎo)； 2.應(yīng)用溫度反應(yīng)性培養(yǎng)皿能獲取完整的BMSCs細胞片層。并用于功能性成骨； 3.將BMSCs片層、BMSCs、EPCs和DBM制備成復(fù)合體植入犬體內(nèi)有望形成血管化的組織工程骨。
[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，

本文編號：2016194