LncRNA H19 knockdown in human amniotic mesenchymal stem cells suppresses angiogenesis by associating with EZH2 and activating VASH1
LncRNA H19 knockdown in human amniotic mesenchymal stem cells suppresses angiogenesis by associating with EZH2 and activating VASH1
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人羊膜间充质干细胞中 LncRNA H19 敲低通过与 EZH2 结合并激活 VASH1 抑制血管生成
DOI：
10.1089/scd.2019.0014
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Ming Shen
中科院分区：
文献类型：
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作者：
Zhiyao Yuan;Yifeng Bian;Xiaojie Ma;Zichun Tang;Ning Chen;Ming Shen
人羊膜间充质干细胞（human amniotic mesenchymal stem cells，HAMSCs）是一种很有前途的种子细胞，在促进血管新生方面具有很大的优势。然而，HAMSC促进血管生成的机制仍不清楚。长链非编码RNA H19参与许多生物学过程，例如增强血管生成和癌细胞的增殖、侵袭和迁移。本研究利用慢病毒载体构建稳定低表达H19（HAMSC-shH19）和随机对照（HAMSC-shNC）的HAMSC，并与人脐静脉内皮细胞（HUVECs）进行三维共培养，研究H19基因敲减对HAMSC血管生成的影响。我们的研究结果表明，H19基因敲低显着抑制HAMSCs的血管生成功能在早期阶段在体外和体内。CCK-8和transwell实验结果表明，HAMSC分泌的条件培养基下调H19后可抑制HUVECs的增殖和迁移。HAMSC-shH19表达和分泌的血管生成因子与对照组相比减少，而血管生成抑制因子升高。此外，我们进行了染色质免疫沉淀和RNA结合蛋白免疫沉淀测定，发现H19可以与Zeste同源物2的组蛋白甲基转移酶增强子（EZH2）相互作用，并且H19敲低抑制了EZH2将甲基募集到血管生成抑制剂基因vasohibin-1（VASH1）的启动子区域的能力，从而增加了VASH1的表达和HAMSC的分泌，抑制血管生成。综上所述，我们的研究确定H19是HAMSCs中促进血管生成的重要调节因子，这将有助于构建理想的基因修饰种子细胞，以促进再生医学中的血管生成。
Human amniotic mesenchymal stem cells (HAMSCs) are promising seed cells with great advantages in promoting angiogenesis. However, the mechanisms underlying angiogenesis facilitated by HAMSCs are still unclear. Long noncoding RNA H19 is involved in many biological processes, such as enhancing angiogenesis and proliferation, invasion, and migration of cancer cells. In this study, we constructed HAMSCs of stable low-expression H19 (HAMSC-shH19) and the scramble control (HAMSC-shNC) using lentiviral vectors, and in a three-dimensional coculture with human umbilical vein endothelial cells (HUVECs) to investigate the effect of H19 knockdown in HAMSCs on angiogenesis. Our results demonstrated that H19 knockdown significantly inhibited the angiogenic function of HAMSCs at an early stage in vitro and in vivo. The results of CCK8 and transwell assays demonstrated that the conditioned medium secreted by HAMSCs reduced proliferation and migration of HUVECs after downregulating H19. The angiogenesis factors expressed and secreted by HAMSC-shH19 were decreased compared with those secreted by the control, while angiogenesis inhibitors were elevated. Furthermore, we conducted chromatin immunoprecipitation and RNA-binding protein immunoprecipitation assays and found that H19 could interact with the histone methyltransferase Enhancer of Zeste homolog 2 (EZH2) and that H19 knockdown inhibited the ability of EZH2 to recruit methyl groups to the promoter region of the angiogenesis inhibitor gene vasohibin-1 (VASH1), thus increasing VASH1 expression and secretion of HAMSCs, suppressing angiogenesis. In summary, our study identified H19 as an important regulator in HAMSCs for promoting angiogenesis, which would help to construct ideal gene-modified seed cells to enhance angiogenesis in regenerative medicine.