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, researchers at the Policlinico San Donato-IRCCS investigated transcriptomic changes induced by endothelial cell exposure to hypoxia. 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.
Enrichment analysis of Gene Ontology Biological Processes (GOBP) of genes modulated in HUVEC upon hypoxia exposure.
Top 10 GOBP ranked by combined score associated with (a) up-modulated and (b) down-modulated signatures using Enrichr analysis tool. Terms belonging to closely related categories are highlighted in the same color. A Benjamini-Hochberg adjusted p-value < 0.05 was used as significance threshold.