摘要
本研究探讨人脂肪间充质干细胞来源的外泌体(ADSC-Exos)对人表皮干细胞(EpSCs)增殖的影响及机制.首先使用Ⅰ型胶原酶分离人脂肪组织ADSCs,中性蛋白酶Ⅱ和胰酶分离人皮肤组织EpSCs;采用ExoQuick-TC试剂分离ADSCs培养上清中的外泌体.然后通过MTT法、细胞免疫荧光检测Ki67,BrdU掺入实验检测ADSC-Exos对EpSCs增殖的影响,流式细胞术检测ADSC-Exos对EpSCs细胞周期的影响.通过H&E染色、免疫组化染色检测细胞增殖标志分子及表皮干细胞标志分子的表达,以观察ADSC-Exos对体外培养皮肤组织的结构及EpSCs增殖的影响.结果显示:ADSC-Exos能以浓度依赖性和时间依赖性的方式促进EpSCs增殖,增加S期细胞数,减少G1期细胞数;ADSC-Exos也能促进体外培养皮肤组织中的EpSCs增殖.机制研究发现:ADSC-Exos对EpSCs的促增殖作用能部分被β-catenin抑制剂XAV-939或c-Myc抑制剂10058-F4所抑制.ADSC-Exos能促进EpSCs表达β-catenin、c-Myc及cyclin E1、A2、D1,XAV-939能抑制ADSC-Exos诱导的β-catenin、c-Myc以及cyclinE1、A2、D1表达,10058-F4能抑制ADSC-Exos诱导的c-Myc和cyclin D1、A2、E1表达.综上,ADSC-Exos能显著促进EpSCs增殖,其作用部分是由上调β-catenin、c-Myc和cyclinE1、A2、D1表达所介导.
This study aimed to investigate the effect of exosomes from adipose-derived mesenchymal stem cells(ADSC-Exos)on the proliferation of human epidermal stem cells(EpSCs)and explore the underlying mechanisms.ADSCs were isolated from human adipose tissue by typeⅠcollagenase digestion.EpSCs were isolated from human skin tissue by dispaseⅡand trypsin digestion.The exosomes were isolated from the supernatant of ADSCs using the ExoQuick-TC reagent.EpSCs proliferation was examined by MTT assay,immunofluorescence staining of Ki67 and BrdU incorporation assay.Cell cycle phase distribution was analyzed by flow cytometry.Cultured human skin tissues were examined for the structure and expression of the markers of cell proliferation and epidermal stem cells by H&E staining and immunohistochemistry,respectively.The results showed that ADSC-Exos promoted EpSCs proliferation in concentration-and time-dependent manners.ADSCExos increased the number of cells in S phase,and decreased the number of cells in G1 phase.ADSC-Exos also significantly promoted the proliferation of EpSCs in cultured skin tissue.Mechanistic study showed that the proliferative activity of ADSC-Exos was partially inhibited byβ-catenin inhibitor XAV-939 or c-Myc inhibitor10058-F4.ADSC-Exos upregulated the expressions ofβ-catenin,c-Myc,cyclins E1,A2 and D1.XAV-939 suppressed ADSC-Exos-induced expressions ofβ-catenin,c-Myc,cyclins E1,A2 and D1.10058-F4 inhibited the upregulation of c-Myc and cyclins E1,A2 and D1 by ADSC-Exos.Collectively,these results indicate that ADSCExos promote EpSCs proliferation partly through upregulating the expression ofβ-catenin,c-Myc and cyclins.
作者
刘金伟
曹高标
程贺云
杜伟伟
张广亮
金光哲
张苹
王兆东
刘喆
巨积辉
乐颖影
傅奕
侯瑞兴
LIU Jin-Wei;CAO Gao-Biao;CHENG He-Yun;DU Wei-Wei;ZHANG Guang-Liang;JIN Guang-Zhe;ZHANG Ping;WANG Zhao-Dong;LIU Zhe;JU Ji-Hui;LE Ying-Ying;FU Yi;HOU Rui-Xing(Department of Hand Surgery,Suzhou Ruihua Orthopedics Hospital,Suzhou 215104,China;Department of Hand Surgery,Ruihua Affiliated Hospital of Soochow University,Suzhou 215104,China;Institute of Hand Surgery,Ruihua Affiliated Hospital of Soochow University,Suzhou 215104,China;Department of Human Anatomy,Histology and Embryology,School of Biology and Basic Medical Sciences,Soochow University,Suzhou 215104,China;Shanghai Institute of Nutrition and Health,Chinese Academy of Sciences,Shanghai 200031,China)
出处
《生物化学与生物物理进展》
SCIE
CAS
CSCD
北大核心
2021年第5期590-600,共11页
Progress In Biochemistry and Biophysics
基金
国家自然科学基金(31900969)
姑苏卫生人才培养项目(GSWS2019088)资助。
关键词
外泌体
脂肪间充质干细胞
表皮干细胞
细胞增殖
exosome
adipose-derived mesenchymal stem cell
epidermal stem cells
cell proliferation
