尿酸影响人骨髓间充质干细胞成骨分化过程中BMP-2表达

发布时间：2018-02-11 11:28

本文关键词： 人骨髓间充质干细胞尿酸成骨分化骨形态形成蛋白-2　出处：《青岛大学》2015年硕士论文　论文类型：学位论文

【摘要】：目的:探讨人类骨髓间充质干细胞(h BMSCs)向成骨细胞分化过程中,不同浓度尿酸对骨形态形成蛋白-2(BMP-2)表达的影响。方法:无菌取健康成人的骨髓样本,利用全骨髓贴壁法分离、培养h BMSCs,选取生长状态良好的第3代h BMSCs为研究对象。将其分为5个组,分别为空白对照组(加入完全培养基)、成骨诱导组(加入成骨诱导液及含0、0.2、0.4、0.8 mmol/L尿酸的完全培养基)。倒置显微镜下观察细胞形态的变化,各类培养基干预培养14天后,进行茜素红染色及碱性条件下抽磷酸酶(ALP)活性检测来评估h BMSCs成骨分化情况。RT-PCR技术检测各组BMP-2 m RNA的表达情况。结果:第3代的h BMSCs大多为形态单一的长梭形,并逐渐形成旋涡状生长;干预诱导后的细胞逐渐变为不规则的立方形,局部细胞重叠生长形成团块状结节,以含尿酸浓度为0.8mmol/L的成骨诱导培养基组最为显著。连续尿酸干预14天后,空白对照组未观察到被茜素红染色的橘红色结节,而各成骨诱导组细胞均有橘红色结节形成。ALP活性随尿酸浓度的增加和尿酸干预时间的延长而增强(P0.05)。RT-PCR检测结果显示,空白对照组无BMP-2 m RNA的表达。成骨诱导组随培养基中尿酸浓度的增加,BMP-2 m RNA表达水平逐渐升高,并呈尿酸浓度依赖性(P0.05)。结论:尿酸可以促进h BMSCs的成骨分化过程,并可通过上调BMP-2 m RNA的表达促进成骨,呈一定的浓度依赖性。
[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.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R580

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