成骨生长肽促进大鼠骨髓间充质干细胞成脂、成骨分化

发布时间：2018-10-05 14:21

【摘要】：目的观察成骨生长肽(OGP)在成脂及成骨性培养基中对大鼠骨髓间充质干细胞(MSC)增殖分化的影响。方法 Percoll法分离大鼠骨髓间充质干细胞,通过频繁更换培养液及反复传代的方法纯化细胞。用传至2代细胞作MTT实验,以此确定OGP对MSC起作用的最大活性浓度;传3代MSC接种于6孔培养板上,共接种13板,分为成脂、成骨诱导两组,每板细胞都分为实验组和对照组,对照组不含成骨生长肽,实验组含有10~(-11)mol/L的成骨生长肽。脂肪细胞鉴定:成脂诱导7、14、21天各取1板细胞做实验,用TG试剂盒在全自动生化分析仪上测定细胞中甘油三酯的总量;成脂诱导21天后取预置盖玻片的1板细胞行脂肪细胞油红O染色。成骨细胞鉴定:取成骨诱导8、12、16天细胞各2板,分别用ALP试剂盒、钙试剂盒测定细胞内碱性磷酸酶活性及每孔钙含量;成骨诱导30天后取预置盖玻片的1板细胞,做Von Kossa染色,计算每张玻片钙结节数。结果传3代大鼠骨髓间充质干细胞成脂诱导7天、14天、21天后分化为脂肪细胞的阳性率以及甘油三酯含量,实验组均高于对照组;成骨诱导8天、12天、16天后细胞内碱性磷酸酶活性测定以及钙含量测定,实验组均高于对照组,30天后矿化结节的形成,实验组高于对照组。结论随着培养条件不同,成骨生长肽可促进大鼠骨髓间充质干细胞向脂肪细胞或成骨细胞分化。
[Abstract]:Objective to observe the effect of osteogenic growth peptide (OGP) on the proliferation and differentiation of rat bone marrow mesenchymal stem cells (MSC) in adipogenic and osteoblast medium. Methods Rat bone marrow mesenchymal stem cells were isolated by Percoll method. The maximum active concentration of OGP on MSC was determined by the MTT assay of passage 2 cells, 13 plates were inoculated on 6 well culture plate, 13 plates were inoculated on 6 well culture plate, each plate cells were divided into experimental group and control group, and each of them was divided into experimental group and control group. The control group did not contain osteogenic growth peptide, while the experimental group contained 10 ~ (- 11) mol/L osteogenic growth peptide. Adipocyte identification: 1 plate cell was taken for 21 days after adipogenic induction, and the total triglyceride content was measured by TG kit in automatic biochemical analyzer. After 21 days of adipogenic induction, 1 plate cells prepositioned with cover glass were stained with fat cell oil red O. Osteoblast identification: two plates of osteoblasts were taken from each of the cells on day 8, 12 and 16, respectively. The activity of alkaline phosphatase and the content of calcium in each hole were measured by ALP kit and calcium kit respectively. After 30 days of osteogenesis induction, the 1 plate cells prepositioned with glass slide were stained with Von Kossa. Calculate the number of calcium nodules per slide. Results the positive rate and triglyceride content of adipocytes differentiated into adipocytes from rat bone marrow mesenchymal stem cells induced by adipogenic induction for 7 days and 14 days and 21 days later were higher in the experimental group than in the control group. The activity of alkaline phosphatase and the content of calcium in the cells of the experimental group were higher than those of the control group after 30 days of osteogenic induction, and the formation of mineralized nodules in the experimental group was higher than that in the control group. Conclusion osteoblast growth peptide can promote the differentiation of rat bone marrow mesenchymal stem cells into adipocytes or osteoblasts with different culture conditions.
【学位授予单位】：兰州大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R329.2

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