OPG转基因小鼠模型的建立及其骨微结构研究
发布时间：2018-09-05 17:36
【摘要】： 目的建立OPG(osteoprotegerin,OPG)转基因小鼠模型,观察其骨微结构、骨密度和骨矿含量的变化,以探讨OPG在体内调节骨代谢的作用及其分子机理。 方法构建带人类OPG基因启动子的真核表达载体pCI-hOPGp-mOPG,显微注射入C57BL"6J小鼠受精卵原核,导入假孕母鼠输卵管后培养子代小鼠。通过PCR法鉴定小鼠基因型,获得OPG转基因小鼠模型(OPG transgenic mouse ,OPG trans )。取SPF级10周龄F2代OPG trans鼠和野生型小鼠(C57BL"6J)各4只,Western Blot测定小鼠OPG蛋白表达;μCT观察小鼠胫骨近端松质骨和皮质骨的骨微结构;同时应用DXA测定小鼠股骨总体骨密度和骨矿含量值。 结果1.构建携带分子标签Flag的表达载体pCI-hOPGp-mOPG质粒经酶切与测序,鉴定序列正确。在69只原代小鼠中,有7只转基因阳性Founder小鼠。目前F4代34只小鼠中,有4系共5只转基因阳性小鼠。 2.与野生型小鼠比较,OPG转基因小鼠胫骨松质骨微结构参数体积骨密度、组织骨密度、骨小梁厚度和骨小梁数目均明显增加,而骨小梁间隔则明显减小;皮质骨微结构参数皮质骨面积、皮质骨厚度、皮质骨密度、皮质骨矿含量明显增加,而内径周长则明显减小(P0.05),外径周长间无显著差异。股骨骨密度与骨矿含量值均明显上升(P0.05)。 结论1.成功建立携带分子标签Flag的OPG转基因小鼠模型。 2.OPG转基因小鼠皮质骨和松质骨的骨量均增加。
[Abstract]:Objective to establish OPG (osteoprotegerin,OPG) transgenic mice model and observe the changes of bone microstructure, bone mineral density and bone mineral content in order to investigate the role of OPG in regulating bone metabolism and its molecular mechanism. Methods Eukaryotic expression vector pCI-hOPGp-mOPG, with human OPG gene promoter was constructed and microinjected into C57BL "6J mouse fertilized egg prokaryote. The mouse genotypes were identified by PCR method, and the OPG transgenic mouse model (OPG transgenic mouse OPG trans). Was obtained. The expression of OPG protein was detected by Western Blot in 4 OPG trans mice and 4 wild type mice (C57BL "6J) in F _ 2 generation of SPF grade 10 weeks old, the bone microstructure of proximal cancellous bone and cortical bone of mouse tibia was observed by 渭 CT, and the total bone mineral density and bone mineral content of femur were measured by DXA. Result 1. The expression vector pCI-hOPGp-mOPG plasmid carrying molecular label Flag was constructed by restriction endonuclease digestion and sequencing, and the sequence was confirmed to be correct. Of 69 primary mice, 7 were transgenic positive Founder mice. At present, there are 5 transgenic positive mice in 4 lines out of 34 F 4 generation mice. 2. 2. Compared with wild-type mice, the bone mineral density (BMD), tissue bone density (BMD), trabecular thickness and the number of trabeculae were significantly increased in OPG transgenic mice, while the trabecular septum decreased significantly. Cortical bone microstructure parameters: cortical bone area, cortical bone thickness, cortical bone mineral density, cortical bone mineral content increased significantly, while the inner diameter circumference decreased significantly (P0.05), but there was no significant difference between outer diameter and circumference. Bone mineral density (BMD) and bone mineral content (BMD) of femur increased significantly (P0.05). Conclusion 1. The model of OPG transgenic mice carrying molecular label Flag was successfully established. The bone mass of cortical bone and cancellous bone of 2.OPG transgenic mice were increased.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R580;R-332
本文编号：2224975
[Abstract]:Objective to establish OPG (osteoprotegerin,OPG) transgenic mice model and observe the changes of bone microstructure, bone mineral density and bone mineral content in order to investigate the role of OPG in regulating bone metabolism and its molecular mechanism. Methods Eukaryotic expression vector pCI-hOPGp-mOPG, with human OPG gene promoter was constructed and microinjected into C57BL "6J mouse fertilized egg prokaryote. The mouse genotypes were identified by PCR method, and the OPG transgenic mouse model (OPG transgenic mouse OPG trans). Was obtained. The expression of OPG protein was detected by Western Blot in 4 OPG trans mice and 4 wild type mice (C57BL "6J) in F _ 2 generation of SPF grade 10 weeks old, the bone microstructure of proximal cancellous bone and cortical bone of mouse tibia was observed by 渭 CT, and the total bone mineral density and bone mineral content of femur were measured by DXA. Result 1. The expression vector pCI-hOPGp-mOPG plasmid carrying molecular label Flag was constructed by restriction endonuclease digestion and sequencing, and the sequence was confirmed to be correct. Of 69 primary mice, 7 were transgenic positive Founder mice. At present, there are 5 transgenic positive mice in 4 lines out of 34 F 4 generation mice. 2. 2. Compared with wild-type mice, the bone mineral density (BMD), tissue bone density (BMD), trabecular thickness and the number of trabeculae were significantly increased in OPG transgenic mice, while the trabecular septum decreased significantly. Cortical bone microstructure parameters: cortical bone area, cortical bone thickness, cortical bone mineral density, cortical bone mineral content increased significantly, while the inner diameter circumference decreased significantly (P0.05), but there was no significant difference between outer diameter and circumference. Bone mineral density (BMD) and bone mineral content (BMD) of femur increased significantly (P0.05). Conclusion 1. The model of OPG transgenic mice carrying molecular label Flag was successfully established. The bone mass of cortical bone and cancellous bone of 2.OPG transgenic mice were increased.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R580;R-332
【参考文献】
相关期刊论文 前2条
1 许勇,项佑贵,严兰珍,王龙,徐国江,费俭,傅继粱,王铸钢;hOPG基因启动子驱动报告基因LacZ的转基因小鼠模型的建立[J];中国生物工程杂志;2005年11期
2 方玲娜;戴如春;廖二元;;异黄酮与骨质疏松[J];国际病理科学与临床杂志;2006年01期
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