Downregulation of the Long Non-Coding RNA KLRK1-AS1 Disturbs Endothelial Barrier Integrity and Promotes Angiogenic Sprouting

Endothelial integrity is essential for cardiovascular health, and circulating endothelial progenitor cells, particularly endothelial colony-forming cells (ECFCs), are key contributors to vascular repair and maintenance. Long non-coding RNAs (lncRNAs) have emerged as novel epigenetic regulators of endothelial physiology and pathology. Building on our previous work identifying the lncRNA <i>KLRK1-AS1</i> as a positive modulator of ECFC wound healing, we aimed to elucidate its role in endothelial biology. Cord blood-derived ECFCs were subjected to siRNA-mediated silencing of <i>KLRK1-AS1</i>, followed by blinded evaluations of monolayer morphology, barrier stability using ECIS impedance measurements, assessments of proliferation, and spheroid-based angiogenic activity. SiRNA-mediated silencing of <i>KLRK1-AS1</i> induced detectable alterations in ECFC monolayer morphology (<i>p</i> = 0.047), while proliferation remained unaffected. Notably, <i>KLRK1-AS1</i> knockdown significantly compromised endothelial barrier integrity, resulting in a 44% reduction in impedance after 48 h (<i>p</i> < 0.001), suggesting weakened intercellular contacts. In contrast, loss of <i>KLRK1-AS1</i> enhanced angiogenic behaviour, demonstrated by an increased number of sprouts (+62%, <i>p</i> = 0.031). Together, these findings indicate that <i>KLRK1-AS1</i> supports a quiescent, stable endothelial phenotype, with intact barrier function, while its depletion shifts ECFCs toward a more angiogenic, activated state. Our results identify <i>KLRK1-AS1</i> as a previously unrecognised regulator of endothelial function.