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Mechanical Strain-Mediated Tenogenic Differentiation of Mesenchymal Stromal Cells Is Regulated through Epithelial Sodium Channels.
Stem Cells International ( IF 4.3 ) Pub Date : 2020-08-18 , DOI: 10.1155/2020/5385960
Hui Yin Nam 1 , Malliga Raman Murali 1 , Raja Elina Ahmad 2 , Belinda Pingguan-Murphy 3 , Hanumantha Rao Balaji Raghavendran 1 , Tunku Kamarul 1
Affiliation  

It has been suggested that mechanical strain may elicit cell differentiation in adult somatic cells through activation of epithelial sodium channels (ENaC). However, such phenomenon has not been previously demonstrated in mesenchymal stromal cells (MSCs). The present study was thus conducted to investigate the role of ENaC in human bone marrow-derived MSCs (hMSCs) tenogenic differentiation during uniaxial tensile loading. Passaged-2 hMSCs were seeded onto silicone chambers coated with collagen I and subjected to stretching at 1 Hz frequency and 8% strain for 6, 24, 48, and 72 hours. Analyses at these time points included cell morphology and alignment observation, immunocytochemistry and immunofluorescence staining (collagen I, collagen III, fibronectin, and N-cadherin), and gene expression (ENaC subunits, and tenogenic markers). Unstrained cells at similar time points served as the control group. To demonstrate the involvement of ENaC in the differentiation process, an ENaC blocker (benzamil) was used and the results were compared to the noninhibited hMSCs. ENaC subunits’ (α, β, γ, and δ) expression was observed in hMSCs, although only α subunit was significantly increased during stretching. An increase in tenogenic genes’ (collagen1, collagen3, decorin, tenascin-c, scleraxis, and tenomodulin) and proteins’ (collagen I, collagen III, fibronectin, and N-cadherin) expression suggests that hMSCs underwent tenogenic differentiation when subjected to uniaxial loading. Inhibition of ENaC function resulted in decreased expression of these markers, thereby suggesting that ENaC plays a vital role in tenogenic differentiation of hMSCs during mechanical loading.

中文翻译:

机械应变介导的间质基质细胞的腱生分化是通过上皮钠通道来调节的。

已经提出,机械应变可以通过激活上皮钠通道(ENaC)引起成年体细胞的细胞分化。但是,这种现象以前尚未在间充质基质细胞(MSCs)中得到证实。因此,进行本研究以研究ENaC在单轴拉伸载荷过程中在人骨髓来源的MSC(hMSCs)肌腱分化中的作用。将传代的2hMSCs接种到涂有胶原蛋白I的硅树脂室上,并以1Hz的频率和8%的应变拉伸6、24、48和72小时。在这些时间点的分析包括细胞形态和比对观察,免疫细胞化学和免疫荧光染色(胶原蛋白I,胶原蛋白III,纤连蛋白和N-钙粘蛋白)和基因表达(ENaC亚基和肌腱标记)。在相似时间点的未应变细胞用作对照组。为了证明ENaC参与分化过程,使用了ENaC阻滞剂(苯扎米尔),并将其结果与未抑制的hMSC进行了比较。ENaC亚基尽管在拉伸过程中仅α亚基显着增加,但在hMSC中观察到了αβγδ)的表达。肌腱基因的增加(胶原1胶原3核心蛋白肌腱蛋白-c硬化腱调节蛋白)和蛋白质(胶原蛋白I,胶原蛋白III,纤连蛋白和N-钙黏着蛋白)的表达表明,hMSC在承受单轴载荷时经历了肌腱分化。ENaC功能的抑制导致这些标志物的表达降低,从而表明ENaC在机械加载过程中在hMSCs的肌腱分化中起着至关重要的作用。
更新日期:2020-08-19
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