Angiopoietin-1 haploinsufficiency affects the endothelial barrier and causes hereditary angioedema.

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Different mutations of the angiopoietin-1 gene (ANGPT1) have been associated with the occurrence of hereditary angioedema (HAE). The purpose of the study is to clarify whether the ANGPT1 A119S variant plays its role via haploinsufficiency or a dominant negative effect. The ability of ANGPT1 A119S variant to affect the endothelial barrier function was assessed by immunocytochemistry. Inter-endothelial gap formation molecules primarily responsible for cell-cell adhesions of HUVECs, vascular endothelial (VE)-cadherin and β-catenin, and reorganization of the F-actin cytoskeletal were evaluated. In invitro conditions mimicking the heterozygous state, the p.A119S variant significantly reduced the capability to bind its natural receptor (80.7% of normal), less than the homozygous condition (59.1%). After stimulation of VEGF or bradykinin, the addiction to equimolar amounts of wtANGPT1 and ANGPT1 p.A119S clearly reduced the expression of VE-cadherin on the endothelial cell surface (31% and 24% respectively). Likewise, cell surface expression of β-catenin was reduced and severe gap formation between adjacent HUVECs developed. In cultured cells, β-catenin expression was mostly observed along the cell surface. Treatment with equimolar amounts of wtANGPT1 and ANGPT1 p.A119S failed to restore the reorganization of the F-actin cytoskeletal elements. ANGPT1 p.A119S variant in homozygous condition further diminished VE-cadherin and β-catenin expression and failed to reduce stress fibre formation significantly affecting the endothelial barrier functionality. Present data show that in a heterozygous state the p.A119S substitution results in a pathogenic loss of function of the protein due to a mechanism of haploinsufficiency. The ANGPT1 reduced ability to counteract the increment of endothelial permeability produced by inducers, such as VEGF and bradykinin, stimulate vascular leakage and reorganization of the F-actin cytoskeletal elements. As a result, a partial impairment of the ANGPT1 functionality, like when dominant mutations occur, represents a pathophysiological cause of HAE.