Elevated expression of miR302-367 in Endothelial cells inhibits developmental angiogenesis via CDC42/CCND1 mediated signaling pathways

Jingjiang Pi, Jie Liu, Tao Zhuang, Lin Zhang, Huimin Sun, Xiaoli Chen, Qian Zhao, Yashu Kuang, Sheng Peng, Xiaohui Zhou, Zuoren Yu, Ting Tao, Brian Tomlinson, Paul Chan, Ying Tian, Huimin Fan, Zhongmin Liu, Xiangjian Zheng, Edward Morrisey, Yuzhen Zhang

Research output: Contribution to journalArticle

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Abstract

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.

Original languageEnglish
Pages (from-to)1511-1526
Number of pages16
JournalTheranostics
Volume8
Issue number6
DOIs
Publication statusPublished - Jan 1 2018

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Endothelial Cells
Cell Movement
Cell Proliferation
MicroRNAs
Actins
Lim Kinases
Physiologic Neovascularization
Pathologic Neovascularization
Actin Depolymerizing Factors
High-Throughput Nucleotide Sequencing
Rhombencephalon
Chromatin Immunoprecipitation
3' Untranslated Regions
Fibrin
Luciferases
Wound Healing
Genes
Embryonic Development
Signal Transduction
Cell Cycle

Keywords

  • Actin remodeling
  • Cdc42
  • Cell cycle
  • Developmental angiogenesis
  • Endothelial cells
  • miR302-367

ASJC Scopus subject areas

  • Medicine (miscellaneous)
  • Pharmacology, Toxicology and Pharmaceutics (miscellaneous)

Cite this

Elevated expression of miR302-367 in Endothelial cells inhibits developmental angiogenesis via CDC42/CCND1 mediated signaling pathways. / Pi, Jingjiang; Liu, Jie; Zhuang, Tao; Zhang, Lin; Sun, Huimin; Chen, Xiaoli; Zhao, Qian; Kuang, Yashu; Peng, Sheng; Zhou, Xiaohui; Yu, Zuoren; Tao, Ting; Tomlinson, Brian; Chan, Paul; Tian, Ying; Fan, Huimin; Liu, Zhongmin; Zheng, Xiangjian; Morrisey, Edward; Zhang, Yuzhen.

In: Theranostics, Vol. 8, No. 6, 01.01.2018, p. 1511-1526.

Research output: Contribution to journalArticle

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 2018, 'Elevated expression of miR302-367 in Endothelial cells inhibits developmental angiogenesis via CDC42/CCND1 mediated signaling pathways', Theranostics, vol. 8, no. 6, pp. 1511-1526. https://doi.org/10.7150/thno.21986
Pi, Jingjiang ; Liu, Jie ; Zhuang, Tao ; Zhang, Lin ; Sun, Huimin ; Chen, Xiaoli ; Zhao, Qian ; Kuang, Yashu ; Peng, Sheng ; Zhou, Xiaohui ; Yu, Zuoren ; Tao, Ting ; Tomlinson, Brian ; Chan, Paul ; Tian, Ying ; Fan, Huimin ; Liu, Zhongmin ; Zheng, Xiangjian ; Morrisey, Edward ; Zhang, Yuzhen. / Elevated expression of miR302-367 in Endothelial cells inhibits developmental angiogenesis via CDC42/CCND1 mediated signaling pathways. In: Theranostics. 2018 ; Vol. 8, No. 6. pp. 1511-1526.
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abstract = "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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T1 - Elevated expression of miR302-367 in Endothelial cells inhibits developmental angiogenesis via CDC42/CCND1 mediated signaling pathways

AU - Pi, Jingjiang

AU - Liu, Jie

AU - Zhuang, Tao

AU - Zhang, Lin

AU - Sun, Huimin

AU - Chen, Xiaoli

AU - Zhao, Qian

AU - Kuang, Yashu

AU - Peng, Sheng

AU - Zhou, Xiaohui

AU - Yu, Zuoren

AU - Tao, Ting

AU - Tomlinson, Brian

AU - Chan, Paul

AU - Tian, Ying

AU - Fan, Huimin

AU - Liu, Zhongmin

AU - Zheng, Xiangjian

AU - Morrisey, Edward

AU - Zhang, Yuzhen

PY - 2018/1/1

Y1 - 2018/1/1

N2 - 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.

AB - 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.

KW - Actin remodeling

KW - Cdc42

KW - Cell cycle

KW - Developmental angiogenesis

KW - Endothelial cells

KW - miR302-367

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