组织工程与重建外科杂志

组织工程与重建外科杂志 ›› 2017, Vol. 13 ›› Issue (6): 318-321.doi: 10.3969/j.issn.1673-0364.2017.06.004

• 论著 • 上一篇    下一篇

bFGF对hBMSCs增殖及成软骨能力影响的实验研究

赵丹丹，陶然，刘豫，曹谊林，周广东   

  1. 潍坊医学院整形外科研究所；上海交通大学医学院附属第九人民医院整复外科，上海市组织工程研究重点实验室；上海市 组织工程国家工程研究中心；上海交通大学医学院附属第九人民医院整复外科，上海市组织工程研究重点实验室；上海市 组织工程国家工程研究中心
  • 收稿日期:2017-10-11 发布日期:2020-07-23

Experimental Study of the Impacts of bFGF on Proliferation and Chondrogenesis of Human Bone Marrow Stromal Cells

ZHAO D andan,TAO Ran,LIU Yu,CAO Yilin,ZHOU Guangdong   

  • Received:2017-10-11 Published:2020-07-23

摘要： 目的 探索碱性成纤维细胞生长因子(b FGF)对人骨髓间充质干细胞(h BMSCs)增殖和成软骨能力的影响,并确定适用于h BMSCs体外软骨构建的细胞代次。方法 获取h BMSCs,分别用DMEM和DMEM+b FGF培养基进行培养。两组细胞均传代至第4代,比较两组各代次h BMSCs的形态变化、细胞得率;取第3代细胞,用pellet法体外成软骨诱导培养3周,观察两组pellet大体观并进行Ⅱ型胶原染色。在上述实验基础上,取h BMSCs用DMEM+b FGF培养基进行培养,1∶3传代培养至第8代,观察各代次细胞的形态变化;对各代次pellet体外成软骨诱导培养3周后,行大体观察并进行Ⅱ型胶原染色。结果 DMEM+b FGF培养体系下的细胞形态、细胞得率及成软骨能力等方面均优于DMEM组。DMEM+b FGF培养体系下,按照1∶3传代的细胞传至第6代仍能维持较好的细胞形态;第1~4代细胞均表达软骨特异性细胞外基质Ⅱ型胶原,第5及后续代次的细胞成软骨能力较差。结论 b FGF可明显促进h BMSCs增殖,并可更好地维持h BMSCs的成软骨能力,在该培养体系下的第1~4代细胞适用于体外软骨构建。

关键词: 碱性成纤维细胞生长因子, 人骨髓间充质干细胞, 细胞代次, 软骨再生

Abstract: Objective To explore the effect of basic fibroblast growth factor (bFGF) on the proliferation and chondrogenesis of human bone marrow mesenchymal stem cells (hBMSCs), and to determine the cell generation schedule suitable for in vitro cartilage construction of hBMSCs. Methods hBMSCs were obtained and cultured respectively in two culture systems:DMEM and DMEM+bFGF. Two groups of cells were repeatedly passaged to P4 generation, the morphological changes and cell yield rates of hBMSCs were compared between the two groups. At the same time the two groups of P3 cells were induced for chondrogenic differentiation using pellet culture for 3 weeks in vitro. Gross observation, immunohistochemical staining of collagen Ⅱ were used to evaluate the results of each group. Based on the study mentioned above, the DMEM+bFGF culture group was subcultured to P8 to observe the morphological changes of each subculture. After chondrogenic differentiation using pellet culture for 3 weeks in vitro, gross observation and immunohistochemical staining of collagenⅡwere evaluated as before. Results The cell morphology, cell yield and cartilage differentiation in DMEM+bFGF group was far greater than that of DMEM group. Under the culture system of DMEM+bFGF, cells passaged at 1:3 could still maintain a good cell morphology in P6 generation. Immunohistochemistry showed that cartilage-specific extracellular matrix typeⅡcollagen was expressed better in P1-4 generation. Conclusion bFGF can significantly promote the proliferation andchondrogenesis of hBMSCs. P1-4 generation cells in DMEM+bFGF culture system is suitable for in vitro cartilage construction.

Key words: Basic fibroblast growth factor, Bone marrow stromal stem cells, Cell generation, Cartilage regeneration

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