本文關鍵詞： 人骨髓間充質干細胞 尿酸 成骨分化 骨形態(tài)形成蛋白-2　出處：《青島大學》2015年碩士論文　論文類型：學位論文

【摘要】：目的:探討人類骨髓間充質干細胞(h BMSCs)向成骨細胞分化過程中,不同濃度尿酸對骨形態(tài)形成蛋白-2(BMP-2)表達的影響。方法:無菌取健康成人的骨髓樣本,利用全骨髓貼壁法分離、培養(yǎng)h BMSCs,選取生長狀態(tài)良好的第3代h BMSCs為研究對象。將其分為5個組,分別為空白對照組(加入完全培養(yǎng)基)、成骨誘導組(加入成骨誘導液及含0、0.2、0.4、0.8 mmol/L尿酸的完全培養(yǎng)基)。倒置顯微鏡下觀察細胞形態(tài)的變化,各類培養(yǎng)基干預培養(yǎng)14天后,進行茜素紅染色及堿性條件下抽磷酸酶(ALP)活性檢測來評估h BMSCs成骨分化情況。RT-PCR技術檢測各組BMP-2 m RNA的表達情況。結果:第3代的h BMSCs大多為形態(tài)單一的長梭形,并逐漸形成旋渦狀生長;干預誘導后的細胞逐漸變?yōu)椴灰?guī)則的立方形,局部細胞重疊生長形成團塊狀結節(jié),以含尿酸濃度為0.8mmol/L的成骨誘導培養(yǎng)基組最為顯著。連續(xù)尿酸干預14天后,空白對照組未觀察到被茜素紅染色的橘紅色結節(jié),而各成骨誘導組細胞均有橘紅色結節(jié)形成。ALP活性隨尿酸濃度的增加和尿酸干預時間的延長而增強(P0.05)。RT-PCR檢測結果顯示,空白對照組無BMP-2 m RNA的表達。成骨誘導組隨培養(yǎng)基中尿酸濃度的增加,BMP-2 m RNA表達水平逐漸升高,并呈尿酸濃度依賴性(P0.05)。結論:尿酸可以促進h BMSCs的成骨分化過程,并可通過上調BMP-2 m RNA的表達促進成骨,呈一定的濃度依賴性。
[Abstract]:Objective: to investigate the effect of different concentrations of uric acid on the expression of bone morphogenetic protein-2BMP-2 during the differentiation of human bone marrow mesenchymal stem cells (BMSCs) into osteoblasts. HBMSCs were cultured, and the third generation of h BMSCs was selected as the research object, which was divided into 5 groups. The cells were divided into blank control group (adding complete culture medium, osteogenic induction group and complete culture medium containing 0. 2 0. 4 0. 8 mmol/L uric acid) respectively. The changes of cell morphology were observed under inverted microscope. After 14 days of culture, all kinds of culture medium were interfered with each other for 14 days. Alizarin red staining and alkaline phosphatase (ALP) activity were used to evaluate the osteogenic differentiation of h BMSCs. RT-PCR was used to detect the expression of BMP-2 m RNA in all groups. The cells induced by the intervention gradually become irregular square, and the local cells overlap to form lumpy nodules. The osteogenic induction medium containing 0.8 mmol / L uric acid concentration was the most significant. After 14 days of continuous uric acid intervention, no orange nodules stained by alizarin red were observed in the blank control group. However, the activity of orange-red nodule formation and ALP increased with the increase of uric acid concentration and the time of uric acid intervention in all osteoblast induction groups. The results of RT-PCR showed that the activity of ALP was increased with the increase of uric acid concentration and the prolongation of uric acid intervention time. There was no expression of BMP-2 m RNA in the blank control group. In the osteogenic induction group, the expression of BMP-2 m RNA increased gradually with the increase of uric acid concentration in culture medium, and the expression of BMP-2 m RNA increased in a concentration dependent manner. Conclusion: uric acid can promote the osteogenic differentiation of h BMSCs. Osteogenesis could be promoted by up-regulating the expression of BMP-2 m RNA in a concentration dependent manner.
【學位授予單位】：青島大學
【學位級別】：碩士
【學位授予年份】：2015
【分類號】：R580

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