BrdU标记对大鼠骨髓间充质干细胞增殖与成骨能力的影响

发布时间：2018-01-04 09:33

本文关键词：BrdU标记对大鼠骨髓间充质干细胞增殖与成骨能力的影响　出处：《福建医科大学》2010年硕士论文　论文类型：学位论文

【摘要】： 目的:观察5-溴-2-脱氧尿嘧啶核苷(5-bromo-2-deoxyuridine, BrdU)标记前后对SD大鼠骨髓间充质干细胞(Mesenchymal stem cells, MSCs)增殖与成骨能力的影响。方法:直接贴壁法分离培养大鼠骨髓MSCs, 0.2%Brdu标记后检测其增殖能力、克隆形成能力;经成骨诱导液诱导培养3周后,利用茜素红染色,荧光免疫组化和Real Time PCR观测标记前后MSCs的成骨能力的变化情况。结果:BrdU标记后MSCs克隆形成能力明显减弱,在克隆个数和面积上均小于未标记组(P0.05);在增殖能力方面, BrdU标记后第4 d开始进入对数增长期时增殖较慢,其中第6 d,8 d,12 d和14 d两组之间有显著性差别,且BrdU标记组增殖高峰较低(P0.05);在成骨能力方面,经成骨诱导后标记组与未标记组的MSCs都表达骨钙素、I型胶原,并且都有明显的钙结节形成,Real Time PCR检测显示标记前后MSCs在骨钙素基因的表达方面无显著性差异(P0.05)。结论:BrdU标记抑制MSCs的克隆增殖形成能力,但不会降低其成骨分化能力,可以广泛应用于MSCs作为种子细胞的干细胞研究的示踪剂。
[Abstract]:Objective: to observe 5-bromo-2-deoxyuridine (5-bromo-2-deoxyuridine). Effects of BrdU labeling on the proliferation and osteogenesis of bone marrow mesenchymal stem cells (MSCs) in SD rats. Methods: MSCs were isolated from rat bone marrow by direct adherent method. The proliferative ability and clone formation ability were detected after labeling with 0.2 Brdu. The osteogenic ability of MSCs was observed by alizarin red staining, fluorescence immunohistochemistry and Real Time PCR. Results the clone forming ability of MSCs was obviously weakened after being labeled with 1: BrdU, and the number and area of clone were lower than that of the unlabeled group (P0.05). In terms of proliferative ability, the proliferation was slower at the beginning of logarithmic growth at the 4th day after BrdU labeling, and there was a significant difference between the two groups on the 6th day, 8 days and 14 days. The peak of proliferation in BrdU labeled group was lower than that of P0.05; In terms of osteogenic ability, osteocalcin type I collagen was expressed in MSCs of both the labeled group and the unlabeled group after osteogenic induction, and obvious calcium nodules were formed. Real Time PCR showed that there was no significant difference in the expression of osteocalcin gene in MSCs before and after labeling. Conclusion the MSCs labeled with 1: BrdU inhibits the ability of clone proliferation but does not decrease its osteogenic differentiation. It can be widely used as a tracer for stem cell research of seed cells by using MSCs as a tracer.
【学位授予单位】：福建医科大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R329

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