本文選題：骨髓間充質(zhì)干細(xì)胞　切入點(diǎn)：增強(qiáng)型綠色熒光蛋白　出處：《山東大學(xué)》2006年碩士論文

【摘要】：目的 體外擴(kuò)增培養(yǎng)兔骨髓間充質(zhì)干細(xì)胞(MSCs)。定向誘導(dǎo)MSCs為類Schwann細(xì)胞。逆轉(zhuǎn)錄病毒載體法使MSCs轉(zhuǎn)染增強(qiáng)型綠色熒光蛋白基因(EGFP)并穩(wěn)定表達(dá)，觀測轉(zhuǎn)染后熒光表達(dá)強(qiáng)度及時(shí)間，，轉(zhuǎn)染前后骨髓間充質(zhì)干細(xì)胞的生物學(xué)特性變化，及轉(zhuǎn)染對MSCs向類Schwann細(xì)胞誘導(dǎo)分化的影響。探討MSCs作為組織工程人工神經(jīng)種子細(xì)胞及利用EGFP作為體內(nèi)試驗(yàn)示蹤劑的可行性。 方法 密度梯度離心聯(lián)合貼壁篩選法分離擴(kuò)增培養(yǎng)兔骨髓間充質(zhì)干細(xì)胞。采用β-巰基乙醇、全反視黃酸、福斯克林、堿性成纖維細(xì)胞生長因子、重組人血小板衍生生長因子等依次誘導(dǎo)MSCs向類Schwann細(xì)胞定向分化，S100免疫細(xì)胞化學(xué)染色鑒定。 同時(shí)用pLEGFP-N1逆轉(zhuǎn)錄病毒載體質(zhì)粒轉(zhuǎn)染PT67包裝細(xì)胞，提取病毒上清液后感染骨髓間充質(zhì)干細(xì)胞，G418篩選及單克隆培養(yǎng)得到穩(wěn)定表達(dá)綠色熒光蛋白的MSCs細(xì)胞(EGFP-MSCs)。觀察測定轉(zhuǎn)染前后骨髓間充質(zhì)干細(xì)胞的生長曲線，細(xì)胞活性和貼壁率并比較。并以相同條件誘導(dǎo)EGFP-MSCs為類Schwann細(xì)胞，比較轉(zhuǎn)染前后S100陽性表達(dá)率。 結(jié)果 1．培養(yǎng)獲取了較高純度的MSCs，細(xì)胞增殖迅速，細(xì)胞活性保持較好。 2．類Schwann細(xì)胞誘導(dǎo)分化：經(jīng)β-巰基乙醇誘導(dǎo)24 h后，細(xì)胞體
[Abstract]:Objective to expand and culture rabbit bone marrow mesenchymal stem cells (MSCs) in vitro.MSCs was induced to be Schwann-like cells.MSCs was transfected and stably expressed by retrovirus vector method. The intensity and time of fluorescence expression after transfection were observed, and the biological characteristics of bone marrow mesenchymal stem cells before and after transfection were observed.And the effect of transfection on the differentiation of MSCs into Schwann-like cells.To explore the feasibility of MSCs as artificial neural seeding cells for tissue engineering and EGFP as tracer in vivo.Methods Bone marrow mesenchymal stem cells were isolated and cultured by density gradient centrifugation and adherent screening.尾 -mercaptoethanol, total retinoic acid, forskine, basic fibroblast growth factor and recombinant human platelet-derived growth factor were used to induce MSCs to differentiate into Schwann like cells by immunocytochemical staining.At the same time, the PT67 packaging cells were transfected with pLEGFP-N1 retrovirus vector plasmid. The MSCs cells expressing green fluorescent protein (GFP) were obtained by G418 screening of bone marrow mesenchymal stem cells infected with virus supernatant, and the EGFP-MSCs cells were obtained by monoclonal culture.The growth curve, cell activity and adherent rate of bone marrow mesenchymal stem cells before and after transfection were observed and compared.The positive expression rate of S100 was compared before and after transfection of EGFP-MSCs as Schwann cells induced by the same conditions.Result 1.High purity MSCs were obtained by culture, the cells proliferated rapidly and the cell activity remained better.2.Differentiation of Schwann like cells: after induced by 尾 -mercaptoethanol for 24 h, cell body
【學(xué)位授予單位】：山東大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2006
【分類號】：R329.2

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