【摘要】：目的：聯(lián)合應用堿性成纖維細胞生長因子（basic fibroblast growth factor，bFGF）與胰島素樣生長因子IGF-1作為誘導劑探討向肌腱細胞方向轉化的誘導技術，以探索一種新的肌腱組織工程種子細胞的獲取方法。 方法： 1.分離、培養(yǎng)大鼠BMSCs并進行分子生物學鑒定：密度梯度離心法體外分離BMSCs，并對培養(yǎng)的BMSCs進行干細胞表型鑒定與成骨、成脂肪及成軟骨多向誘導確定所獲取的BMSCs是具有多向分化能力的BMSCs。 2.觀察bFGF和IGF-1聯(lián)合應用對BMSCs向肌腱細胞分化的作用：將BMSCs隨機分為4組，即實驗組A(IGF-1+bFGF)、實驗組B(IGF-1)、實驗組C(bFGF)及空白對照組D于37℃、5%CO2孵箱中培養(yǎng)。用相差顯微鏡觀察細胞形態(tài)的變化；RT-PCR方法檢測各組細胞內I型膠原、III型膠原以及scleraxis mRNA在3天、7天、14天和21天時的表達水平；免疫熒光染色檢測各組細胞內I型膠原、III型膠原14天的表達水平。 結果： 1.分離培養(yǎng)的BMSCs在形態(tài)學上表現(xiàn)為長梭形、呈漩渦狀生長。流式細胞儀檢測結果顯示細胞表面CD29和CD90表達陽性，分別為99.24%、99.06%；CD45表達陰性，為98.71%。BMSCs經(jīng)誘導后茜素紅、油紅O、甲苯胺藍及II型膠原免疫細胞化學染色均為陽性。 2.誘導后的BMSCs A組、B組、C組、與空白對照組相比其I、III、SCX在基因及蛋白水平上具有顯著差異，從3天開始不斷遞增，14天后轉而下降，以7天至14天的變化最明顯與空白對照組差異最大。而在A、B、C三組中，以B組I、III、SCX基因的表達最明顯，免疫熒光染色檢測中B組細胞內I型膠原、Ⅲ型膠原的表達水平最高。 結論： 大鼠BMSCs體外誘導各組所獲得的細胞與陰性對照組細胞在形態(tài)與功能上存在顯著差別，，其形態(tài)特征及胞內基質表達近似肌腱細胞；尤其是IGF-1組。bFGF與IGF-1兩種細胞因子聯(lián)用沒有明顯明顯協(xié)同效應。
[Abstract]:Objective: to explore the induction technique of transformation to tendon cells by using basic fibroblast growth factor (basic fibroblast growth factor,bFGF and insulin-like growth factor IGF-1 as inducers. In order to explore a new method for obtaining seed cells from tendon tissue engineering. Method: 1. Rat BMSCs was isolated, cultured and identified by molecular biology: BMSCs, was isolated in vitro by density gradient centrifugation and stem cell phenotype and osteogenesis of cultured BMSCs were identified. Multidirectional induction of adipogenesis and chondrogenesis determines that the obtained BMSCs is a BMSCs. with multidirectional differentiation ability. 2. To observe the effect of bFGF and IGF-1 on the differentiation of BMSCs into tendon cells: BMSCs was randomly divided into four groups: experimental group A (IGF-1 bFGF), experimental group B (IGF-1), experimental group C (bFGF) and blank control group D at 37 鈩

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