Wnt信号通路
间充质干细胞
钙粘蛋白
软骨发生
免疫印迹
化学
细胞生物学
干细胞
连环素
硫氧化物9
染色
骨形态发生蛋白2
分子生物学
再生(生物学)
骨髓
生物
信号转导
病理
免疫学
医学
基因表达
细胞
体外
生物化学
基因
作者
Feng Qu,Xuezhen Shen,Haipeng Li,Jingbin Zhou,Bang-tuo Yuan,Chunbao Li,Wei Qi,Yujie Liu,Mingzhu Zhang
出处
期刊:Cellular and Molecular Biology
日期:2022-02-27
卷期号:67 (6): 249-259
被引量:3
标识
DOI:10.14715/cmb/2021.67.6.33
摘要
This study aimed to compare and analyze the effect of N-cadherin on chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs) and to explore the related mechanism, so as to provide a novel theoretical basis for the clinical work of articular cartilage injury regeneration and repair. For this purpose, the experimental animals were clean grade SD rats (aged 5-6 weeks, weighing 180-250g). Alcian blue staining was carried out to observe the induced chondrogenesis following N-cadherin inhibition. The specific role of N-cadherin in the Wnt signaling pathway and chondrogenic differentiation of BMSCs was detected by Western blot; while the effect of N-cadherin on the molecular level changes of β-catenin in the cytoplasm was evaluated by fluorescence quantitative real-time PCR (qRT-PCR). In addition, immunoprecipitation (IP) was used for the verification of the interaction between N-cadherin and β-catenin. Results showed that under the light microscope, 90% of the BMSCs at the third generation, 90% of the cells were fused. Alcian blue staining showed that the green staining area in the BMP2 induction group was large and dense, while that in the N-cadherin inhibition group and blank control group was small and sparse. Western blot revealed that N-cadherin and SOX9 were significantly developed in the BMP2 induction group, but Wnt3a was not significantly developed. While in the N-cadherin inhibition group, the development of Wnt3a was obvious, yet without evident development of N-cadherin and SOX9. The qRT-PCR indicated that the relative mRNA expression of Wnt3a was significantly increased in the N-cadherin inhibition group (P<0.05). However, no obvious difference was observed in the mRNA expression of β-catenin between the BMP2 induction group and the N-cadherin inhibition group (P>0.05). Western blot indicated that in the BMP2 induction group; there existed the development of β-catenin, significant development of phos-GSK-3β and total GSK-3β, but no obvious development of Wnt3a. In the N-cadherin inhibition group, there was significantly enhanced development of Wnt3a and β-catenin than that before, blurred development of phos-GSK-3β than that before, and also obvious development of total GSK-3β with little change from before. N-cadherin promoted the expression of β-catenin mostly in the cell membrane, but only a few in the cytoplasm and nucleus. Additionally, verification by IP showed that N-cadherin and β-catenin were developed on N-cadherin and β-catenin bands, suggesting an interaction between N-cadherin and β-catenin. According to these results, N-cadherin can ultimately promote chondrogenic differentiation of BMSCs by inhibiting the Wnt signaling pathway.
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