本文選題：間充質(zhì)干細胞 + Buffy��； 參考：《延邊大學》2017年碩士論文

【摘要】：目的:比較分析兔骨髓間充質(zhì)干細胞(Mesenchymal Stem Cells,MSC)與含有單核細胞、生長因子和細胞因子等的Buffy coat在體外軟骨分化,為Buffy coat治療軟骨損傷提供實驗依據(jù)。方法:動物選取新西蘭系大白兔12只(16周齡,體重3±0.5kg),隨機分為:MSC組(陰性對照組,n=3),采集骨髓的間充質(zhì)干細胞(MSC),不添加轉(zhuǎn)化生長因子-β(Transforing growth factor-β,TGF-β)的誘導下培養(yǎng)。Buffy coat組(實驗組,n=6),采用Buffy coat不經(jīng)過體外的傳代培養(yǎng),在軟骨誘導液下培養(yǎng)。MSC+TGF組(陽性對照組,n=3),采集骨髓的間充質(zhì)干細胞,添加TGF-β的誘導下培養(yǎng)。通過大體觀察、組織學觀察、糖胺多糖(Glycosaminoglycan,GAG)檢測,和體外軟骨BERN的評分比較三組在體外分化的能力。結(jié)果:1)大體觀察:MSC組培養(yǎng)的軟骨部分已經(jīng)溶解,邊緣模糊;MSC+TGF組與Buffy coat組基本沒有溶解現(xiàn)象,邊緣圓潤、清晰。2)番紅O染色鏡下觀察:MSC組軟骨細胞生長緩慢,分布不均勻,數(shù)量極少,有極少量GAG的分泌現(xiàn)象出現(xiàn),分泌不規(guī)則;有少數(shù)細胞可以找到軟骨細胞特有的Cartilage lacuna結(jié)構(gòu)。Buffy coat組與MSC+TGF組軟骨細胞生長速度較快,細胞分布較均勻,細胞數(shù)量較多,表面較平整、周圍較圓潤,有較多GAG分泌現(xiàn)象出現(xiàn),基本分布在軟骨細胞周圍;有較多細胞可以找到Cartilage lacuna結(jié)構(gòu)。3)糖胺多糖(GAG)含量測定:MSC 組:7.78± 1.79μg,Buffy coat 組:22.83±3.11μg,MSC+TGF 組:24.05±3.35μg。三組間存有顯著差異,MSC 組與 MSC+TGF 組、MSC 與 Buffy coat 組,組間存在顯著差異(P0.05);Buffy coat組與MSC+TGF組,組間無顯著差異(P0.05)。4)體外軟骨 BERN 評分:MSC 組:2.83± 1.16,Buffy coat 組:6.16±1.16,MSC+TGF組:6.68±0.81。三組間存在顯著差異,MSC組與MSC+TGF組、MSC與Buffy coat組,組間存在顯著差異(P0.05),Buffy coat組與MSC+TGF組,組間無顯著差異(P0.05)。結(jié)論:Buffy coat具有良好的體外軟骨分化能力,可以解決目前體外分化軟骨必須添加生長因子這個難題,為Buffy coat在臨床應用提供了實驗依據(jù)。
[Abstract]:Aim: to compare and analyze the cartilage differentiation of rabbit bone marrow mesenchymal Stem cells (MSCs) and Buffy coat containing monocytes, growth factors and cytokines in vitro, so as to provide experimental evidence for the treatment of cartilage injury by Buffy coat. Methods: twelve New Zealand white rabbits aged 16 weeks were selected. Body weight was 3 鹵0.5 kg 路kg ~ (-1), and was randomly divided into two groups: 1 / growth group (negative control group). MSCs were collected from bone marrow and cultured without TGF- 尾-尾 -Transforing growth factor- 尾 (TGF- 尾). Buffy coat group (experimental group) was cultured without Buffy coat in vitro. Mesenchymal stem cells (MSCs) were collected from bone marrow and cultured with TGF- 尾. The ability of differentiation of the three groups in vitro was compared by gross observation, histological observation, glycosaminoglycan glycosaminoglycan gag test and cartilage BERN score in vitro. Results (1) the chondrocytes cultured in the TGF group were dissolved, but there was no dissolution in the TGF group and the Buffy coat group. The edges were smooth and clear. Under the microscope, the chondrocytes in the Buffy coat group grew slowly and distributed unevenly. A few cells could find the specific Cartilage lacuna structure of chondrocytes. Buffy coat group and MSC TGF group had a faster growth rate, more evenly distributed cells and more cells. The surface was smooth, the surrounding was smooth, and there were more GAG secretion phenomena, which were mainly distributed around the chondrocytes, and the Cartilage lacuna structure. 3) glycosaminoglycan (Gagg) content could be found in the Cartilage lacuna group. The content of Cartilage was 7. 78 鹵1. 79 渭 g / m Buffy coat group and 20. 83 鹵3. 11 渭 g / m GAG TGF group. There was significant difference among the three groups. There was significant difference between the two groups (P0.05 Buffy coat group and MSC TGF group). There was no significant difference among the three groups (P0.05. 4) the in vitro cartilage BERN score was 2.83 鹵1.16 Buffy coat group, 6.16 鹵1.16 MSC coat group, 6.68 鹵0.81MSC TGF group. There was significant difference among the three groups. There was no significant difference between the two groups (P0.05Buffy coat group and MSC TGF group). Conclusion coat has a good ability of cartilage differentiation in vitro, which can solve the problem that growth factor must be added to the differentiation cartilage in vitro, and provide experimental basis for the clinical application of Buffy coat.
【學位授予單位】：延邊大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R687;R318.08

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