Modulating <i>ACVRL1</i> Expression in HMEC1 Cells as a Simplified <i>In Vitro</i> Model for Hereditary Hemorrhagic Telangiectasia (HHT) Type 2 Studies.

Abstract

<h4>Background/aim</h4>Hereditary hemorrhagic telangiectasia (HHT) is a rare disease with an incidence of 1:5,000. HHT is inherited in an autosomal dominant manner and is associated with vascular malformations. It particularly affects the genes <i>ENG</i> (HHT1) and <i>ACVRL1</i> (HHT2). Clinically, patients typically exhibit pronounced recurrent epistaxis. The aim of this study was to evaluate if <i>ACVRL1</i> knockdown in HMEC-1 endothelial cells could induce a HHT2-like phenotype which could be deployed as a model for HHT2.<h4>Materials and methods</h4>The human immortalized endothelial cell line HMEC-1 was used for the experiments. RNAi knockdown was performed using a pool of four siRNAs targeting the <i>ACVRL1</i> gene. The gene knockdown was verified using RT-qPCR and Western blot analysis. The effects of <i>ACVRL1</i> knockdown on angiogenesis were compared to a non-target (NT) small RNA control using a tube formation assay. The expression of 84 endothelia-associated genes was analyzed with RT-qPCR.<h4>Results</h4>Tube formation ability was significantly affected by <i>ACVRL1</i> knockdown. In particular, the parameters total tube length, total segment length, total master segment length, total branching length, total mesh area and branching interval were significantly increased whereas total isolated branch length, number of master junctions, number of branches and the number of isolated segments decreased after <i>ACVRL1</i> knockdown. Significant changes in the expression of angiogenesis related genes were detected by qPCR analysis and discussed.<h4>Conclusion</h4>Knockdown of <i>ACVRL1</i> in HMEC-1 endothelial cells leads to pathological angiogenesis with enhanced tube formation capacity, which is similar to the angiogenesis <i>in vivo</i> in patients with HHT2. The presented HMEC-1 cell-based system therefore has the potential to be deployed as an <i>in vitro</i> model for HHT2 studies.