大鼠骨髓间充质干细胞与髁突软骨细胞共培养诱导软骨化分化的实验研究

摘要/Abstract

摘要： 目的研究大鼠骨髓间充质干细胞与髁突软骨细胞共培养对骨髓间充质干细胞（bone marrow stromal cells,BMSCs）向软骨化分化的影响。方法分离、培养大鼠BMSCs与髁突软骨细胞,取第2代细胞以 2∶1 比例混匀为共培养组,细胞终浓度为1.2×10⁴/mL;以相同浓度的髁突软骨细胞作为实验组,以相同浓度的BMSCs作为对照组。倒置显微镜下观察细胞形态;以CCK-8法检测细胞增殖;阿利新蓝染色、ALP染色及定量分析检测细胞成软骨分化;Western免疫印迹检测Ⅱ型胶原、Ⅹ型胶原的表达。采用SPSS13.0 软件包对数据进行统计学分析。结果共培养组与实验组细胞呈多角形、铺路石样排列,为典型软骨细胞形态,而对照组细胞呈长梭形。细胞生长曲线显示,共培养组较其他2组细胞增殖能力强;共培养组与实验组阿利新兰染色、ALP染色均为阳性;Western 免疫印迹显示,共培养组细胞的Ⅱ型胶原、Ⅹ型胶原的表达显著升高。结论在共培养中,BMSCs有利于髁突软骨细胞表型的维持以及促进软骨细胞的增殖;同时,髁突软骨细胞具有诱导BMSCs向软骨分化的作用。

关键词: 髁突, 软骨细胞, 骨髓间充质干细胞, 共培养, 种子细胞

Abstract: PURPOSE: To investigate the influence of co-culture of rat bone marrow mesenchymal stem cells (BMSCs) and condylar chondrocytes on chondrogenic differentiation of BMSCs. METHODS: BMSCs and condylar chondrocytes were isolated and cultured from SD rat. Passage 2 BMSCs and chondrocytes (1.2×10⁴/mL) were co-cultured at a ratio of 2 to 1 as co-culture group, condylar chondrocytes with same concentration and passage as experimental group, BMSCs with same concentration and passage as control group . Inverted microscope was used to observe cell morphology; CCK-8 was used to evaluate the proliferation of the cells; alcian blue staining, alkaline phosphatase(ALP) staining and quantitative analysis were used to evaluate cell differentiation; Western blot was used to detect the expression of COL2A1,COL10A1. One-way ANOVA with SPSS13.0 software package was performed for statistical analysis. RESULTS: The cells of co-culture group and experimental group were polygon-shaped and displayed cobble-stone morphology which were typical morphology of chondrocytes, while cells in the control group were long spindle-shaped; cell growth curve indicated cells in the co-culture group had greater proliferation ability compared with the other two groups; alcian blue and ALP staining of cells in the co-culture group and experimental group were positive ;the expression of COL2A1 and COL10A1 were significantly increased. CONCLUSIONS: BMSCs can maintain the phenotype of condylar chondrocytes and promote proliferation of chondrocytes. At the same time, condylar chondrocytes can induce BMSCs to chondrogenic differentiation.

Key words: Condyle, Chondrocytes, Bone marrow mesenchymal stem cells, Co-culture, Seed cell

中图分类号:

Q254

盛夏涵, 曲禄瑶, 蔡协艺. 大鼠骨髓间充质干细胞与髁突软骨细胞共培养诱导软骨化分化的实验研究[J]. 中国口腔颌面外科杂志, 2016, 14(6): 490-494.

SHENG Xia-han, QU Lu-yao, CAI Xie-yi. Co-culture of rat bone marrow mesenchymal stem cells and condylar chondrocytes induces chondrogenic differentiation[J]. China J Oral Maxillofac Surg, 2016, 14(6): 490-494.

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