摘要
目的建立稳定的比格犬骨髓间充质干细胞(BMSCs)诱导成骨分化模型;鉴定参与比格犬BMSCs细胞成骨分化过调控的蛋白质,探讨其可能的调控机制。方法分离培养比格犬BMSCs细胞,rhBMP-2诱导细胞成骨分化7 d,基于蛋白质双向凝胶电泳的蛋白质组学分析成骨分化组与对照组差异表达的蛋白质,对感兴趣的蛋白质LASP1、ferritin轻链和重链表达情况用Q-PCR、Western blot进行验证。结果成功诱导比格犬BMSCs成骨分化;蛋白质组学分析获得rhBMP-2诱导上调的蛋白质9个,下调的蛋白质11个。荧光定量PCR技术和Western blot对LASP1和ferritin表达情况进行了验证,发现rhBMP-2诱导7 d后,LASP1显著下调而ferritin显著上调,与蛋白质组学结果一致。结论 LASP1在调节细胞骨架活性方面发挥重要功能,而ferritin是细胞铁离子稳态维持的重要分子,提示这两种蛋白质在rhBMP-2诱导的比格犬BMSCs成骨分化过程中发挥重要作用。
Objective To identify the proteins involved in osteogenic differentiation of beagle dog bone marrow mesenchymal stem cells(BMSCs) and explore its possible regulation mechanism of osteogenic differentiation of BMSCs.Methods Cultured beagle dog BMSCs were induced by recombinant human bone morphogenetic protein-2(rhBMP-2) for 7 days.The differentially expressed proteins between cells with osteogenic differentiation and control cells were identified by proteomics analysis based on two-dimensional gel electrophoresis.Q-PCR and Western blotting were used to verify the interested protein of LASP1,ferritin light chain and heavy chain.Results Osteogenic differentiation was induced successfully in the BMSCs.Twenty differentially expressed proteins were identified by proteomic analysis,including 9 down-regulated and 11 upregulated ones.Q-PCR and Western blotting demonstrated a significant reduction of LASP1 expression and significant upregulation of ferritin in the BMSCs after a 7-day induction with rhBMP-2.Conclusion LASP1,which plays an important role in the regulation of the activity of the cytoskeleton,and ferritin,an important molecule in cellular iron homeostasis,can be critical in the osteogenic differentiation of beagle dog BMSCs induced by rhBMP-2.
出处
《南方医科大学学报》
CAS
CSCD
北大核心
2013年第8期1207-1212,共6页
Journal of Southern Medical University
基金
广东省自然科学基金(10451051501005738)
广东省教育厅育苗工程(LYM10050)
广东省科技计划项目(2011B031300020)
