The semi-selective barrier provided by the endothelium ensures vascular integrity and tissue homeostasis. Under physiological conditions, interendothelial junction-complexes, including adherence and tight junctions, orchestrate vascular permeability in tissue-specific manner. In contrast, disintegration of these complexes is a hallmark of several detrimental pathologies including inflammation, oedema, and tissue ischemia. A huge endeavour has been made to understand the molecular mechanisms and pathways controlling endothelial barrier function; however, little is still known regarding the role of long non-coding RNAs in this context.

Here we report on the endothelial-enriched lncRNA NTRAS as a gatekeeper of endothelial barrier function. In vivo, pharmacological Ntras silencing suffices to induce vascular leakage, demonstrated by increased extravasation of FTSC-dextran (123 ± 10%) and TMR-dextran (172 ± 27%) in murine heart tissue. Strikingly, Ntras-deficient mice showed a significantly reduced survival (-25% during 10 days follow up; p=0.01). Besides, ~60% of Ntras-silenced mice exhibited a broad range of cardiac pathologies, ranging from mild inflammation to severe defects such as myocarditis and scar formation (p<0.05). Mechanistically, we demonstrate NTRAS to interact with hnRNPL to fine-tune alternative splicing of tight junction protein 1 (TJP1) pre-mRNA, eventually regulating endothelial permeability. Specifically, by sequestration of the splicing regulator hnRNPL, NTRAS promotes the expression of the so-called TJP1a+ isoform which we found to contribute to endothelial resistance. In contrast, NTRAS silencing and targeted splicing modulation increase TJP1a- expression, and thereby impaired barrier function. Deleting a bona fide motif within Ntras, serving as a binding site for hnRNPL, impaired cardiac vascular integrity and manifested as induced tracer penetration (FTSC-dextran 147 ± 15%; TMR-dextran 163 ± 21%; p<0.05). At the same time, these mice showed induced cardiac inflammation characterized by an increased CD45⁺ cells infiltration (299 ± 34%; p<0.001) into heart tissue.

Collectively, we unveil NTRAS as a constituent of a splicing-regulatory network detrimental in sustaining vascular integrity through preserving the equilibrium between TJP1α+ and α- isoforms. These results further expand our knowledge regarding the physiological roles of lncRNAs in vascular biology, especially in the context of regulating barrier function.