Chinese Journal of Tissue Engineering Research ›› 2022, Vol. 26 ›› Issue (30): 4834-4839.doi: 10.12307/2022.763
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Li Ruiyu, Shi Xu, Chen Qi, Zuo Hua, Li Kezhen
Received:
2021-08-30
Accepted:
2021-10-15
Online:
2022-10-28
Published:
2022-03-29
Contact:
Chen Qi, MD, Associate chief physician, Affiliated Hospital of Jiangsu University, Zhenjiang 212000, Jiangsu Province, China
About author:
Li Ruiyu, Master candidate, Affiliated Hospital of Jiangsu University, Zhenjiang 212000, Jiangsu Province, China
Supported by:
CLC Number:
Li Ruiyu, Shi Xu, Chen Qi, Zuo Hua, Li Kezhen. Transforming growth factor beta 3 in mediating chondrogenic differentiation of mesenchymal stem cells from different sources[J]. Chinese Journal of Tissue Engineering Research, 2022, 26(30): 4834-4839.
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2.1 原代细胞与成软骨诱导分化过程中的镜下及整体结构改变 原代细胞接种后静置于培养箱2 d,行半换液去除部分悬浮细胞,可见细胞贴壁生长,形态不规则,10 d后镜下观察:骨髓间充质干细胞大部分呈不规则三角形,核呈椭圆形,可见双核细胞,见图1A;鼻黏膜间充质干细胞大部分呈梭形生长,核形态与骨髓间充质干细胞相似,见图1B。第3代骨髓间充质干细胞和鼻黏膜间充质干细胞在未含转化生长因子β3的成软骨诱导分化培养基中培养4 d镜下观察:骨髓间充质干细胞形成大小不一的集落,呈放射状紧密排列,多为长梭形,见图1C;鼻黏膜间充质干细胞贴壁似鱼群样分布,形态均一,生长速度较快,见图1D。第3代骨髓间充质干细胞和鼻黏膜间充质干细胞在含有转化生长因子β3的成软骨诱导分化培养基中培养4 d镜下观察:细胞交织成网状,难以区分细胞间隙,见图1E,F。"
2.2 RT-PCR检测成软骨分化过程中的相关基因表达 干预7 d后,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的COL2A1、Aggrecan mRNA表达均较各自对照组显著上调(P < 0.01);鼻黏膜间充质干细胞实验组的COL2A1、Aggrecan mRNA表达较骨髓间充质干细胞实验组显著上调(P < 0.01);骨髓间充质干细胞实验组的SOX-9 mRNA表达与其对照组无显著差异(P > 0.05);鼻黏膜间充质干细胞实验组的SOX-9 mRNA表达较其对照组显著上调(P < 0.01);鼻黏膜间充质干细胞实验组的SOX-9 mRNA表达较骨髓间充质干细胞实验组显著上调(P < 0.01);骨髓间充质干细胞对照组和鼻黏膜间充质干细胞对照组在成软骨分化过程中COL2A1、SOX-9及Aggrecan mRNA表达无明显差异(P > 0.05),见图3。"
2.3 免疫印迹检测诱导软骨分化过程中相关蛋白表达水平 干预7,14 d后,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的COL2A1、SOX-9蛋白表达均高于各自对照组,差异均有显著性意义(P < 0.01);干预7 d时,两实验组的COL2A1蛋白表达差异无显著性意义(P > 0.05),然而干预14 d时,骨髓间充质干细胞实验组COL2A1蛋白表达高于鼻黏膜间充质干细胞实验组,差异有显著性意义(P < 0.01),干预7 d时,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的COL2A1蛋白表达均增加约1.4倍,而干预14 d时,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的COL2A1蛋白表达增加约3.2,2.2倍。干预7,14 d时,鼻黏膜间充质干细胞实验组的SOX-9蛋白表达高于骨髓间充质干细胞实验组,差异有显著性意义(P < 0.01),干预7 d时,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的SOX-9蛋白表达增加约1.2,1.6倍,而干预14 d时,骨髓间充质干细胞实验组和鼻黏膜间充质干细胞实验组的SOX-9蛋白表达增加约1.1,1.3倍;骨髓间充质干细胞对照组和鼻黏膜间充质干细胞对照组在成软骨分化过程中COL2A1、SOX-9蛋白表达无明显差异(P > 0.05),见图4。"
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