title: Implication of Long noncoding RNAs in the endothelial cell response to hypoxia revealed by RNA-sequencing.
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% O2 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.
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: C
authors.initials: C
authors.lastname: Voellenkle
authors.affiliation: Center for Translational Genomics and Bioinformatics, San Raffaele Scientific Institute, 20132 Milan, Italy.
authors.forename: J M
authors.initials: JM
authors.lastname: Garcia-Manteiga
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Pedrotti
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Perfetti
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: I
authors.initials: I
authors.lastname: De Toma
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: D
authors.initials: D
authors.lastname: Da Silva
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: B
authors.initials: B
authors.lastname: Maimone
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: S
authors.initials: S
authors.lastname: Greco
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Fasanaro
authors.affiliation: Laboratory of Stem Cells for Tissue Engineering, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Creo
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: G
authors.initials: G
authors.lastname: Zaccagnini
authors.affiliation: Division of Cardiovascular Epigenetics, Goethe University, 60590 Frankfurt am Main, Germany.
authors.forename: C
authors.initials: C
authors.lastname: Gaetano
authors.affiliation: Laboratory of Molecular Cardiology, Policlinico San Donato-IRCCS, 20097 Milan, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Martelli
mesh_terms: D000818:Animals; D015687:Cell Hypoxia; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism; D004195:Disease Models, Animal; D059585:G1 Phase Cell Cycle Checkpoints; D059014:High-Throughput Nucleotide Sequencing; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D007150:Immunohistochemistry; D007511:Ischemia / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D020320:Oligoribonucleotides, Antisense; D034622:RNA Interference; D062085:RNA, Long Noncoding / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017423:Sequence Analysis, RNA; D059467:Transcriptome
chemical_list: D050759:Cyclin-Dependent Kinase Inhibitor p21; C094335:H19 long non-coding RNA; D020320:Oligoribonucleotides, Antisense; D062085:RNA, Long Noncoding