纤维型肌动蛋白对间充质干细胞衰老的调节作用及其机制

doi:10.13481/j.1671-587x.20180305

吉林大学学报(医学版) ›› 2018, Vol. 44 ›› Issue (03): 483-486.doi: 10.13481/j.1671-587x.20180305

纤维型肌动蛋白对间充质干细胞衰老的调节作用及其机制

俞冬升¹, 蔡大敏², 陈婕妤², 吕方怡², 花扣珍², 银国利², 王俊娟²

1. 浙江省人民医院杭州医学院附属人民医院骨科, 浙江杭州 310014;
2. 杭州医学院解剖学与组织胚胎学教研室, 浙江杭州 310014

收稿日期:2017-12-13 出版日期:2018-05-28 发布日期:2018-05-31
通讯作者: 王俊娟,助教(Tel:0571-87692675,E-mail:wang.junjuan@foxmail.com) E-mail:wang.junjuan@foxmail.com
作者简介:俞冬升(1988-),男,浙江省杭州市人,主治医师,医学博士,主要从事再生医学方面的研究。
基金资助:
浙江省科技厅自然科学基金资助课题（LY18H090011）；浙江省教育厅一般科研项目资助课题（Y201738521，Y201738494）

Regulation effect of F-actin on senescence of mesenchymal stem cells and its mechanism

YU Dongsheng¹, CAI Damin², CHEN Jieyu², LYU Fangyi², HUA Kouzhen², YIN Guolin², WANG Junjuan²

1. Department of Orthopaedics, Affiliated Hostpial, Hangzhou Medical College, People's Hospital, Zhejiang Province, Hangzhou 310014, China;
2. Department of Anatomy and Histology and Embryology, Hangzhou Medical College, Hangzhou 310014, China

Received:2017-12-13 Online:2018-05-28 Published:2018-05-31

摘要/Abstract

摘要： 目的：探讨纤维型肌动蛋白（F-actin）调节人骨髓来源间充质干细胞（hBMSCs）衰老的作用，初步阐明hBMSCs衰老的分子生物学机制。方法：将分离得到的hBMSCs进行体外培养并分为对照组（P2代hBMSCs）、F-actin抑制剂组（2.5 μmol·L^-1 F-actin抑制剂Latrunculin B处理P2代hBMSCs 1 h）和P11代hBMSCs组（P2代hBMSCs连续传代得到P11代hBMSCs）。成骨、成脂和成软骨诱导液诱导各组hBMSCs，采用茜素红染色、SO染色和油红O染色确定诱导效果。免疫荧光染色观察各组hBMSCs中Ki67阳性细胞数、F-actin的形态和聚合情况、YAP在细胞内的亚细胞定位。SA-β-Gal染色检测各组hBMSCs中SA-β-Gal染色阳性细胞数。结果：茜素红染色、SO染色和油红O染色，hBMSCs具有成骨、成脂和成软骨分化的能力。免疫荧光染色和SA-β-Gal染色，对照组hBMSCs中的微丝纤维束较多较粗，F-actin长度较长，YAP主要集中在细胞核（YAP在细胞核内为活化态）；与对照组比较，P11代hBMSCs组中Ki67阳性细胞数较少，SA-β-Gal阳性细胞较多，F-actin更短更细，YAP主要集中在胞浆，且YAP出核的细胞SA-β-Gal染色呈阳性；F-actin抑制剂组hBMSCs中YAP出核失活，SA-β-Gal染色阳性的hBMSCs衰老细胞更多。结论：抑制YAP入核可以促进hBMSCs衰老，F-actin可以通过调节YAP活性影响hBMSCs衰老。

关键词: YAP蛋白, 纤维型肌动蛋白, 间充质干细胞, 衰老

Abstract: Objective: To investigate the regulation effects of fibrous-actin (F-actin) on senescent of the human bone marrow mesenchymal stem cells (hBMSCs), and to elucidate the molecular mechanisms of senescence the of hBMSCs. Methods: The hBMSCs were cultured in vitro and divided into control group (P2 hBMSCs), F-actin inhibitor group (P2 hBMSCs were treated with 2.5 μmol·L^-1 F-actin inhibitor Latrunculin B for 1 h) and P11 hBMSCs group (P11 hBMSCs passaged from P2 hBMSCs continuously). The hBMSCs in various groups were induced by osteogenic, adipogenic and chondrogenic induction mediums and the induction effects were identified by Alizarin red staining, SO staining and oil red O staining. The number of Ki67 positive cells, polymerization and morphology of F-actin and YAP subcellular localization were detected by immunofluorescence staining. SA-β-Gal staining was uesd to detect the SA-β-Gal staining positive cells in the hBMSCs in various groups. Results: The Alizarin red staining, SO staining and oil red O staining results showed that the hBMSCs in various groups had the osteogenic, adipogenic and chondrogenic abilities.The immunofluorescence staining and SA-β-Gal staining results showed that the microfilament bundles in the hBMSCs in control group were more thick, the F-actin length was longer, and the YAP mainly concentrated in the nucleus (YAP was activated in the nucleus). Compared with control group, the number of Ki67 positive cells in P11 hBMSCs group was less and the number of SA-β-Gal positive cells was more;the Factin was shorter and thinner,the YAP mainly concentrated in the cytoplasm,and the high cytoplasmic YAP cells was positive for SA-β-Gal staining;in F-actin inhibitor group,the high cytoplasmic YAP inactivation was found in the hBMSCs and the SA-β-Gal staining positive hBMSCs had more senescent cells. Conclusion: To inhibit the entering of YAP into the nucleus can promote the senescence of hBMSCs and F-actin can affect the senescence of hBMSCs by regulating the YAP activity.

Key words: senescence, mesenchymal stem cells, fibrous-actin YAP protein

中图分类号:

R329.2

俞冬升, 蔡大敏, 陈婕妤, 吕方怡, 花扣珍, 银国利, 王俊娟. 纤维型肌动蛋白对间充质干细胞衰老的调节作用及其机制[J]. 吉林大学学报(医学版), 2018, 44(03): 483-486.

YU Dongsheng, CAI Damin, CHEN Jieyu, LYU Fangyi, HUA Kouzhen, YIN Guolin, WANG Junjuan. Regulation effect of F-actin on senescence of mesenchymal stem cells and its mechanism[J]. Journal of Jilin University Medicine Edition, 2018, 44(03): 483-486.

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纤维型肌动蛋白对间充质干细胞衰老的调节作用及其机制

Regulation effect of F-actin on senescence of mesenchymal stem cells and its mechanism

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