Long non‑coding RNA NORAD regulates angiogenesis of human umbilical vein endothelial cells via miR‑590‑3p under hypoxic conditions

Zhao,Xiaoxue; Wei,Xiufang; Wang,Xueying; Qi,Guoxian

doi:10.3892/mmr.2020.11064

Long non‑coding RNA NORAD regulates angiogenesis of human umbilical vein endothelial cells via miR‑590‑3p under hypoxic conditions

Authors:
- Xiaoxue Zhao
- Xiufang Wei
- Xueying Wang
- Guoxian Qi
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Affiliations: Department of Geriatrics, The First Hospital of China Medical University, Shenyang, Liaoning 110001, P.R. China
Published online on: April 8, 2020 https://doi.org/10.3892/mmr.2020.11064
Pages: 2560-2570
Copyright: © Zhao et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Dysregulation of angiogenesis can be caused by hypoxia, which may result in severe diseases of the heart, including coronary artery disease. Hypoxia‑inducible factor 1 (HIF‑1) modulates angiogenesis via the regulation of several angiogenic factors. However, the underlying mechanism of hypoxia‑induced angiogenesis remains unknown. In the present study, it was hypothesized that long non‑coding RNA (lncRNA) non‑coding RNA activated by DNA damage (NORAD) may serve a role in the process of angiogenesis via the regulation of microRNA(miR)‑590‑3p under hypoxic conditions. The effect of NORAD and miR‑590‑3p on cell viability and properties associated with angiogenesis, including cell migration and tube formation in human umbilical vein endothelial cells (HUVECs) under hypoxic conditions, were assessed. Potential downstream angiogenic factors of miR‑590‑3p were also determined by molecular experiments. It was identified that NORAD expression was upregulated and miR‑590‑3p expression was downregulated in hypoxia‑exposed HUVECs, and also in myocardial infarction (MI) left ventricle tissues in mice. Moreover, downregulation of NORAD expression resulted in decreased cell viability and angiogenic capacity, but further knocking down miR‑590‑3p expression reversed these alterations, resulting in increased cell migration and tube formation in HUVECs under hypoxic conditions for 24 h. It was demonstrated that NORAD overexpression also increased cell vitality and tube‑formation capacity. Furthermore, NORAD was identified to bind with miR‑590‑3p directly, and miR‑590‑3p was shown to target certain proangiogenic agents, such as vascular endothelial growth factor (VEGF)A, fibroblast growth factor (FGF)1 and FGF2 directly. Therefore, the present results suggested that lncRNA NORAD may bind with miR‑590‑3p to regulate the angiogenic ability of HUVECs via the regulation of several downstream proangiogenic factors under hypoxia. Thus, the lncRNA NORAD/miR‑590‑3p axis may be a novel regulatory pathway in the angiogenic mechanisms in HUVECs, which highlights a potentially novel perspective for treating ischemia/hypoxia‑induced angiogenic diseases.

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1	Carmeliet P: Mechanisms of angiogenesis and arteriogenesis. Nat Med. 6:389–395. 2000. View Article : Google Scholar : PubMed/NCBI
2	Risau W: Mechanisms of angiogenesis. Nature. 386:671–674. 1997. View Article : Google Scholar : PubMed/NCBI
3	Yancopoulos GD, Davis S, Gale NW, Rudge JS, Wiegand SJ and Holash J: Vascular-specific growth factors and blood vessel formation. Nature. 407:242–248. 2000. View Article : Google Scholar : PubMed/NCBI
4	Ferrara N: Vascular endothelial growth factor: Molecular and biological aspects. Curr Top Microbiol Immunol. 237:1–30. 1999.PubMed/NCBI
5	Zimna A and Kurpisz M: Hypoxia-inducible factor-1 in physiological and pathophysiological angiogenesis: Applications and therapies. Biomed Res Int. 2015:5494122015. View Article : Google Scholar : PubMed/NCBI
6	Greijer AE, van der Groep P, Kemming D, Shvarts A, Semenza GL, Meijer GA, van de Wiel MA, Belien JA, van Diest PJ and van der Wall E: Up-regulation of gene expression by hypoxia is mediated predominantly by hypoxia-inducible factor 1 (HIF-1). J Pathol. 206:291–304. 2005. View Article : Google Scholar : PubMed/NCBI
7	Semenza GL: Angiogenesis in ischemic and neoplastic disorders. Annu Rev Med. 54:17–28. 2003. View Article : Google Scholar : PubMed/NCBI
8	Mori S, Tran V, Nishikawa K, Kaneda T, Hamada Y, Kawaguchi N, Fujita M, Saegusa J, Takada YK, Matsuura N, et al: A dominant-negative FGF1 mutant (the R50E mutant) suppresses tumorigenesis and angiogenesis. PLoS One. 8:e579272013. View Article : Google Scholar : PubMed/NCBI
9	Semenza GL: Oxygen sensing, hypoxia-inducible factors, and disease pathophysiology. Annu Rev Pathol. 9:47–71. 2014. View Article : Google Scholar : PubMed/NCBI
10	Go AS, Mozaffarian D, Roger VL, Benjamin EJ, Berry JD, Borden WB, Bravata DM, Dai S, Ford ES, Fox CS, et al: Heart disease and stroke statistics-2013 update: A report from the American heart association. Circulation. 127:e6–e245. 2013. View Article : Google Scholar : PubMed/NCBI
11	Murray CJ and Lopez AD: Mortality by cause for eight regions of the world: Global burden of disease study. Lancet. 349:1269–1276. 1997. View Article : Google Scholar : PubMed/NCBI
12	Lavine KJ, Kovacs A, Weinheimer C and Mann DL: Repetitive myocardial ischemia promotes coronary growth in the adult mammalian heart. J Am Heart Assoc. 2:e0003432013. View Article : Google Scholar : PubMed/NCBI
13	Fatica A and Bozzoni I: Long non-coding RNAs: New players in cell differentiation and development. Nat Rev Genet. 15:7–21. 2014. View Article : Google Scholar : PubMed/NCBI
14	Kung JT, Colognori D and Lee JT: Long noncoding RNAs: Past, present, and future. Genetics. 193:651–669. 2013. View Article : Google Scholar : PubMed/NCBI
15	Han P, Li W, Lin CH, Yang J, Shang C, Nuernberg ST, Jin KK, Xu W, Lin CY, Lin CJ, et al: A long noncoding RNA protects the heart from pathological hypertrophy. Nature. 514:102–106. 2014. View Article : Google Scholar : PubMed/NCBI
16	Wang K, Long B, Zhou LY, Liu F, Zhou QY, Liu CY, Fan YY and Li PF: CARL lncRNA inhibits anoxia-induced mitochondrial fission and apoptosis in cardiomyocytes by impairing miR-539-dependent PHB2 downregulation. Nat Commun. 5:35962014. View Article : Google Scholar : PubMed/NCBI
17	Lee S, Kopp F, Chang TC, Sataluri A, Chen B, Sivakumar S, Yu H, Xie Y and Mendell JT: Noncoding RNA NORAD regulates genomic stability by sequestering PUMILIO proteins. Cell. 164:69–80. 2016. View Article : Google Scholar : PubMed/NCBI
18	Liu H, Li J, Koirala P, Ding X, Chen B, Wang Y, Wang Z, Wang C, Zhang X and Mo YY: Long non-coding RNAs as prognostic markers in human breast cancer. Oncotarget. 7:20584–20596. 2016.PubMed/NCBI
19	Li H, Wang X, Wen C, Huo Z, Wang W, Zhan Q, Cheng D, Chen H, Deng X, Peng C and Shen B: Long noncoding RNA NORAD, a novel competing endogenous RNA, enhances the hypoxia-induced epithelial-mesenchymal transition to promote metastasis in pancreatic cancer. Mol Cancer. 16:1692017. View Article : Google Scholar : PubMed/NCBI
20	Miao Z, Guo X and Tian L: The long noncoding RNA NORAD promotes the growth of gastric cancer cells by sponging miR-608. Gene. 687:116–124. 2019. View Article : Google Scholar : PubMed/NCBI
21	Kawasaki N, Miwa T, Hokari S, Sakurai T, Ohmori K, Miyauchi K, Miyazono K and Koinuma D: Long noncoding RNA NORAD regulates transforming growth factor-β signaling and epithelial-to-mesenchymal transition-like phenotype. Cancer Sci. 109:2211–2220. 2018. View Article : Google Scholar : PubMed/NCBI
22	Ekhteraei-Tousi S, Mohammad-Soltani B, Sadeghizadeh M, Mowla SJ, Parsi S and Soleimani M: Inhibitory effect of hsa-miR-590-5p on cardiosphere-derived stem cells differentiation through downregulation of TGFβ signaling. J Cell Biochem. 116:179–191. 2015. View Article : Google Scholar : PubMed/NCBI
23	Jafarzadeh M and Soltani BM: Hsa-miR-590-5p interaction with SMAD3 transcript supports its regulatory effect on The TGFβ signaling pathway. Cell J. 18:7–12. 2016.PubMed/NCBI
24	Nallamshetty S, Chan SY and Loscalzo J: Hypoxia: A master regulator of microRNA biogenesis and activity. Free Radic Biol Med. 64:20–30. 2013. View Article : Google Scholar : PubMed/NCBI
25	Kumar MM and Goyal R: LncRNA as a therapeutic target for angiogenesis. Curr Top Med Chem. 17:1750–1757. 2017. View Article : Google Scholar : PubMed/NCBI
26	Rehmsmeier M, Steffen P, Hochsmann M and Giegerich R: Fast and effective prediction of microRNA/target duplexes. RNA. 10:1507–1517. 2004. View Article : Google Scholar : PubMed/NCBI
27	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
28	Guide for the Care and Use of Laboratory Animals. Institute for Laboratory Animal Research. National Academies Press. (Washington, DC). 2011.
29	Yang Z, Zhao Y, Lin G, Zhou X, Jiang X and Zhao H: Noncoding RNA activated by DNA damage (NORAD): Biologic function and mechanisms in human cancers. Clin Chim Acta. 489:5–9. 2019. View Article : Google Scholar : PubMed/NCBI
30	Madanecki P, Kapoor N, Bebok Z, Ochocka R, Collawn JF and Bartoszewski R: Regulation of angiogenesis by hypoxia: The role of microRNA. Cell Mol Biol Lett. 18:47–57. 2013. View Article : Google Scholar : PubMed/NCBI
31	Folkman J and Shing Y: Angiogenesis. J Biol Chem. 267:10931–10934. 1992.PubMed/NCBI
32	Geng L, Chaudhuri A, Talmon G, Wisecarver JL and Wang J: TGF-Beta suppresses VEGFA-mediated angiogenesis in colon cancer metastasis. PLoS One. 8:e599182013. View Article : Google Scholar : PubMed/NCBI
33	Aplin AC and Nicosia RF: Hypoxia paradoxically inhibits the angiogenic response of isolated vessel explants while inducing overexpression of vascular endothelial growth factor. Angiogenesis. 19:133–146. 2016. View Article : Google Scholar : PubMed/NCBI
34	Noren Hooten N, Abdelmohsen K, Gorospe M, Ejiogu N, Zonderman AB and Evans MK: microRNA expression patterns reveal differential expression of target genes with age. PLoS One. 5:e107242010. View Article : Google Scholar : PubMed/NCBI