Elevated Expression of miR302-367 in Endothelial Cells Inhibits Developmental Angiogenesis via CDC42/CCND1 Mediated Signaling Pathways.
Elevated Expression of miR302-367 in Endothelial Cells Inhibits Developmental Angiogenesis via CDC42/CCND1 Mediated Signaling Pathways.
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内皮细胞中 miR302-367 表达升高通过 CDC42/CCND1 介导的信号通路抑制发育性血管生成
DOI：
10.7150/thno.21986
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发表时间：
2018
期刊：
影响因子：
12.4
通讯作者：
Zhang Y
中科院分区：
文献类型：
--
作者：
Pi J;Liu J;Zhuang T;Zhang L;Sun H;Chen X;Zhao Q;Kuang Y;Peng S;Zhou X;Yu Z;Tao T;Tomlinson B;Chan P;Tian Y;Fan H;Liu Z;Zheng X;Morrisey E;Zhang Y
基本原理：血管生成对胚胎发育至关重要，microRNA可以对这一过程进行微调，但其潜在机制仍不完全清楚。使用内皮细胞（EC）特异性miR 302 -367系作为功能获得模型，使用抗miRs作为功能丧失模型，以研究miR 302 - 367对内皮细胞（EC）的作用。367关于发育性血管生成的研究，其中胚胎后脑血管系统作为体内模型，纤维蛋白凝胶珠和管形成测定作为体外模型。Boyden小室法和划痕法检测细胞迁移能力，MTT法、Ki 67免疫组化染色和PI细胞周期分析检测细胞增殖能力。RNA高通量测序鉴定了通过染色质免疫沉淀和3 '-UTR荧光素酶报告基因测定确认的miR-靶基因，并且最终靶位点阻断剂确定了对microRNA表达后观察到的表型有显著贡献的途径。EC miR 302 -367表达升高会减少发育中的血管生成，而抑制miR 302 -367则会增强这种作用，这可能是由于对EC迁移和增殖的内在抑制作用。我们将Cdc 42鉴定为直接靶基因，并且升高的EC miR 302 -367降低了总的和活性Cdc 42，并且通过WASP和Klf 2/Grb 2/Pak 1/LIM-激酶/Cofilin途径进一步抑制了F-肌动蛋白的形成。MiR 302 -367介导的Klf 2对Grb 2的调节，用于微调Pak 1的活化，有助于抑制F-actin的形成，然后减弱EC迁移。结论：miR 302 - 367通过调控内皮细胞Cdc 42和Ccnd 1信号通路促进EC的迁移和增殖，从而进一步加深了我们对发育性血管生成的认识，同时也为将来对病理性血管生成的干预提供了理论基础。
Rationale: Angiogenesis is critical for embryonic development and microRNAs fine-tune this process, but the underlying mechanisms remain incompletely understood.Methods: Endothelial cell (EC) specific miR302-367 line was used as gain-of-function and anti-miRs as loss-of-function models to investigate the effects of miR302-367 on developmental angiogenesis with embryonic hindbrain vasculature as an in vivo model and fibrin gel beads and tube formation assay as in vitro models. Cell migration was evaluated by Boyden chamber and scratch wound healing assay and cell proliferation by cell count, MTT assay, Ki67 immunostaining and PI cell cycle analysis. RNA high-throughput sequencing identified miR-target genes confirmed by chromatin immunoprecipitation and 3'-UTR luciferase reporter assay, and finally target site blocker determined the pathway contributing significantly to the phenotype observed upon microRNA expression.Results: Elevated EC miR302-367 expression reduced developmental angiogenesis, whereas it was enhanced by inhibition of miR302-367, possibly due to the intrinsic inhibitory effects on EC migration and proliferation. We identified Cdc42 as a direct target gene and elevated EC miR302-367 decreased total and active Cdc42, and further inhibited F-actin formation via the WASP and Klf2/Grb2/Pak1/LIM-kinase/Cofilin pathways. MiR302-367-mediated-Klf2 regulation of Grb2 for fine-tuning Pak1 activation contributing to the inhibited F-actin formation, and then the attenuation of EC migration. Moreover, miR302-367 directly down-regulated EC Ccnd1 and impaired cell proliferation via the Rb/E2F pathway.Conclusion: miR302-367 regulation of endothelial Cdc42 and Ccnd1 signal pathways for EC migration and proliferation advances our understanding of developmental angiogenesis, and meanwhile provides a rationale for future interventions of pathological angiogenesis that shares many common features of physiological angiogenesis.
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DOI：
10.1387/ijdb.103213sp
发表时间：
2011
期刊：
The International journal of developmental biology
影响因子：
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作者：
Patel-Hett S;D'Amore PA
通讯作者：
D'Amore PA
影响因子：
3.9
作者：
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de Gunzburg, J
影响因子：
20.3
作者：
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Ruhrberg, Christiana
分子机制和血管生成的临床应用。
影响因子：
64.8
作者：
Carmeliet, Peter;Jain, Rakesh K.
通讯作者：
Jain, Rakesh K.
DOI：
10.1083/jcb.141.5.1147
发表时间：
1998-06-01
期刊：
The Journal of cell biology
影响因子：
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Allen WE;Zicha D;Ridley AJ;Jones GE
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