400014 重庆,重庆医科大学附属儿童医院儿研所干细胞研究室,国家儿童健康与疾病临床医学研究中心,儿童发育疾病研究教育部重点实验室,儿科学重庆市重点实验室
Received:22 November 2022,
Revised:2023-11-06,
Published:15 December 2023
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Nannan ZHANG, Chaoqun HU, Yun HE, et al. Role of hypoxia in promoting migration and cell fusion of urine-derived stem cells[J]. Journal of Army Medical University, 2023, 45(23): 2467-2475.
Nannan ZHANG, Chaoqun HU, Yun HE, et al. Role of hypoxia in promoting migration and cell fusion of urine-derived stem cells[J]. Journal of Army Medical University, 2023, 45(23): 2467-2475. DOI: 10.16016/j.2097-0927.202211171.
目的
2
体外实验中探讨低氧对尿源性干细胞(Urine derived stem cells
USCs)迁移及融合能力的影响及该过程中细胞骨架结构和极性的改变。
方法
2
常氧(20%)或低氧(3%)条件下培养USCs 48h,应用划痕实验、Transwell实验检测USCs迁移能力;共聚焦显微镜及流式细胞术观察并测定USCs与肝细胞(HP14-19和HepG2)的融合能力;透射电子显微镜及鬼笔环肽荧光探针观察细胞骨架结构;免疫荧光、流式细胞术及Real-time PCR实验检测细胞极性相关蛋白RhoA、Cdc42及Rac1的表达。
结果
2
与常氧组USCs相比,低氧组USCs的迁移能力及与HP14-19和HepG2的细胞融合能力明显提升。透射电镜及鬼笔环肽荧光染色可见低氧处理后USCs细胞骨架重塑,微丝微管排列更加有序且规律。免疫荧光、流式细胞术及Real-time PCR结果提示,低氧组USCs极性相关蛋白RhoA、Cdc42及Rac1的表达量较常氧组明显增加。
结论
2
低氧处理USCs可促进细胞的迁移及融合能力,其机制可能与细胞骨架的重塑和细胞极性上调有关。
Objective
2
To explore the effect of hypoxia on cell migration and fusion of urine-derived stem cells (USCs)
in vitro
and observe the changes in cytoskeleton and cell polarity during the process.
Methods
2
After USCs were cultured under normoxia (20%) or hypoxia (3%) for 48 h
cell migration were detected by cell scratch test and Transwell assay
USCs' fusion with hepatic HP14-19 and HepG2 cells were observed and determined using confocal microscopy and flow cytometry
and structure of cytoskeleton was observed with transmission electron microscopy (TEM) and phalloidin fluorescence probe. The expression of cell polarity related proteins
RhoA
Cdc42 and Rac1 was measured with immunofluorescence staining
flow cytometry and real-time PCR.
Results
2
The USCs from the hypoxia group showed stronger migration capacity and cell fusion with hepatocytes when compared with the cells from the normoxia group. The results of TEM and phalloidin fluorescence staining revealed that hypoxia induced remodeling of cytoskeleton
and regularly and well-arranged microfilament and microtubule. The results of immunofluorescence staining
flow cytometry and real-time PCR indicated that the expression levels of cell polarity related proteins RhoA
Cdc42 and Rac1 were much higher in the hypoxia group than in the normoxia group.
Conclusion
2
Hypoxia preconditioning promotes cell migration and fusion in USCs
which may be associated with cytoskeleton remodeling and up-regulation of cell polarity.
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