Implication of Long noncoding RNAs in the endothelial cell response to hypoxia revealed by RNA-sequencing

C. Voellenkle, J. M. Garcia-Manteiga, S. Pedrotti, A. Perfetti, I. De Toma, D. Da Silva, B. Maimone, S. Greco, P. Fasanaro, P. Creo, G. Zaccagnini, C. Gaetano, F. Martelli

Research output: Contribution to journalArticle

Abstract

Long noncoding RNAs (lncRNAs) are non-protein coding RNAs regulating gene expression. Although for some lncRNAs a relevant role in hypoxic endothelium has been shown, the regulation and function of lncRNAs is still largely unknown in the vascular physio-pathology. Taking advantage of next-generation sequencing techniques, transcriptomic changes induced by endothelial cell exposure to hypoxia were investigated. Paired-end sequencing of polyadenylated RNA derived from human umbilical vein endothelial cells (HUVECs) exposed to 1% O 2 or normoxia was performed. Bioinformatics analysis identified ≈2000 differentially expressed genes, including 122 lncRNAs. Extensive validation was performed by both microarray and qPCR. Among the validated lncRNAs, H19, MIR210HG, MEG9, MALAT1 and MIR22HG were also induced in a mouse model of hindlimb ischemia. To test the functional relevance of lncRNAs in endothelial cells, knockdown of H19 expression was performed. H19 inhibition decreased HUVEC growth, inducing their accumulation in G1 phase of the cell cycle; accordingly, p21 (CDKN1A) expression was increased. Additionally, H19 knockdown also diminished HUVEC ability to form capillary like structures when plated on matrigel. In conclusion, a high-confidence signature of lncRNAs modulated by hypoxia in HUVEC was identified and a significant impact of H19 lncRNA was shown.

Original languageEnglish
Article number24141
JournalScientific Reports
Volume6
DOIs
Publication statusPublished - Apr 11 2016

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Long Noncoding RNA
RNA Sequence Analysis
Endothelial Cells
Human Umbilical Vein Endothelial Cells
Untranslated RNA
Hypoxia
G1 Phase
Hindlimb
Computational Biology
Endothelium
Blood Vessels
Cell Cycle
Ischemia
Pathology
Gene Expression
Messenger RNA

ASJC Scopus subject areas

  • General

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Implication of Long noncoding RNAs in the endothelial cell response to hypoxia revealed by RNA-sequencing. / Voellenkle, C.; Garcia-Manteiga, J. M.; Pedrotti, S.; Perfetti, A.; De Toma, I.; Da Silva, D.; Maimone, B.; Greco, S.; Fasanaro, P.; Creo, P.; Zaccagnini, G.; Gaetano, C.; Martelli, F.

In: Scientific Reports, Vol. 6, 24141, 11.04.2016.

Research output: Contribution to journalArticle

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abstract = "Long noncoding RNAs (lncRNAs) are non-protein coding RNAs regulating gene expression. Although for some lncRNAs a relevant role in hypoxic endothelium has been shown, the regulation and function of lncRNAs is still largely unknown in the vascular physio-pathology. Taking advantage of next-generation sequencing techniques, transcriptomic changes induced by endothelial cell exposure to hypoxia were investigated. Paired-end sequencing of polyadenylated RNA derived from human umbilical vein endothelial cells (HUVECs) exposed to 1{\%} O 2 or normoxia was performed. Bioinformatics analysis identified ≈2000 differentially expressed genes, including 122 lncRNAs. Extensive validation was performed by both microarray and qPCR. Among the validated lncRNAs, H19, MIR210HG, MEG9, MALAT1 and MIR22HG were also induced in a mouse model of hindlimb ischemia. To test the functional relevance of lncRNAs in endothelial cells, knockdown of H19 expression was performed. H19 inhibition decreased HUVEC growth, inducing their accumulation in G1 phase of the cell cycle; accordingly, p21 (CDKN1A) expression was increased. Additionally, H19 knockdown also diminished HUVEC ability to form capillary like structures when plated on matrigel. In conclusion, a high-confidence signature of lncRNAs modulated by hypoxia in HUVEC was identified and a significant impact of H19 lncRNA was shown.",
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AU - Pedrotti, S.

AU - Perfetti, A.

AU - De Toma, I.

AU - Da Silva, D.

AU - Maimone, B.

AU - Greco, S.

AU - Fasanaro, P.

AU - Creo, P.

AU - Zaccagnini, G.

AU - Gaetano, C.

AU - Martelli, F.

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