敲降ZEB1基因表达对人脂肪间充质干细胞增殖、周期与凋亡的影响

基础医学与临床 ›› 2017, Vol. 37 ›› Issue (3): 335-340.

敲降ZEB1基因表达对人脂肪间充质干细胞增殖、周期与凋亡的影响

刘妍¹,李红凌²,赵春华¹

1. 中国医学科学院基础医学研究所
2. 中国医学科学院北京协和医学院基础医学研究所

收稿日期:2016-11-14 修回日期:2016-12-20 出版日期:2017-03-05 发布日期:2017-02-23
通讯作者: 赵春华 E-mail:zhaochunhua@vip.163.com
基金资助:
国家自然科学基金面上项目

Effects of ZEB1 gene knockdown on proliferation,cell cycle and apoptosis of human adipose-derived mesenchymal stem cells

Received:2016-11-14 Revised:2016-12-20 Online:2017-03-05 Published:2017-02-23

摘要/Abstract

摘要： 目的探讨敲降ZEB1基因表达对人脂肪间充质干细胞（hADSCs）增殖、周期和凋亡的影响。方法胶原酶消化法提取人脂肪组织中原代间充质干细胞，对其进行免疫学表型、成骨和成脂分化能力鉴定后用于实验。脂质体转染法将siRNA转入hADSCs，实时定量PCR和Western blot检测ZEB1、细胞增殖、周期和凋亡相关基因mRNA和蛋白的表达。MTS法检测细胞增殖。流式细胞术检测细胞周期和细胞凋亡率。结果与si-NC转染组相比，si-ZEB1转染组可使ZEB1 mRNA和蛋白表达水平降低（p<0.01），细胞增殖能力明显下降（p<0.05），增殖相关基因CCND1、MKI67、MYC和PCNA表达显著下调（p<0.05或p<0.01）；G1期细胞比例从50.71%增加到58.94%（p<0.01），S期和G2期细胞比例分别减少了6.16%和2.07%；细胞凋亡率上升了10.2%，促凋亡相关基因TP53和BAX表达上调，抗凋亡基因MCL1和BCL2表达下调（p<0.05或p<0.01）。结论 ZEB1具有促进hADSCs增殖和周期进展，抑制细胞凋亡的作用。

关键词: 关键词：人脂肪间充质干细胞, ZEB1, 增殖, 周期, 凋亡

Abstract: Objective To investigate the effects of ZEB1 (Zinc finger E-box binding homeobox 1) gene knockdown on proliferation, cell cycle and apoptosis of human adipose-derived mesenchymal stem cells(hADSCs). MethodsPrimary mesenchymal stem cells were obtained from human adipose tissue by collagenase digestion and identified by immunological phenotype and differentiation potential of osteogenic and adipogenic lineages. The siRNA was transfected into hADSCs by lipofectamine 2000.The expression of ZEB1 and key proliferation, cell cycle and apoptosis-related genes were detected by real-time PCR and Western blot at mRNA and protein level respectively. The cell proliferation capacity was assessed by MTS. The apoptosis and cell cycle of hADSCs were evaluated by flow cytometry. ResultsThe ZEB1 expression at mRNA and protein level was significantly decreased in si-ZEB1 transfection cells compared to transfection with si-NC. Inhibition of ZEB1 decreased the cell proliferation ability, and significantly inhibited the expression of cell proliferation related genes,including CCND1,MKI67, MYC and PCNA. After transfection withsi-ZEB1, the percentage of cells in G1 phase increased from 50.17% to 58.94%, and S phase and G2 phase decreased by 6.16% and 2.07% separately.Meanwhile, the apoptosis rate increased by 10.2%, the expression of apoptosis related genes TP53 and BAX was up-regulated, and the expression of anti-apoptotic genes MCL1 and BCL2was down-regulated in si-ZEB1 transfection cells compared to transfection with si-NC. ConclusionsZEB1 may play an important role in promoting hADSCs proliferation, cell cycle progression and inhibit their apoptosis.

Key words: Key words：human adipose-derived mesenchymal stem cells, ZEB1, proliferation, cell cycle, apoptosis

中图分类号:

Q291

刘妍李红凌赵春华. 敲降ZEB1基因表达对人脂肪间充质干细胞增殖、周期与凋亡的影响[J]. 基础医学与临床, 2017, 37(3): 335-340.

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