NETosis is associated with the severity of aortic stenosis: Links with inflammation

Abstract

Background: An involvement of neutrophil extracellular traps (NETs) in the aortic stenosis (AS) pathogenesis is unknown.

Methods: We enrolled 50 patients, median age 66.5 years with isolated severe AS, after aortic valve replacement and 20 healthy sex/age-matched controls. Autopsy-derived aortic valves from 5 healthy donors served as controls. Valvular expression of citrullinated histone H3 (citH3), myeloperoxidase (MPO), and neutrophil elastase (NE) and macrophages (CD68) were evaluated by immunostaining. Plasma citH3 and interleukin 6 (IL-6) were also determined.

Results: All stenotic and healthy valves expressed citH3 in the leaflets' endothelial and sub-endothelial layers at the aortic side. Amount of valvular citH3-positive cells was higher in AS patients compared with controls (53.5 [33-62]% vs. 5.7 [4.1-9.0]%, p < 0.0001) and correlated with aortic valve area (AVA; r = -0.69, p < 0.0001) and mean transvalvular gradient (r = 0.6, p < 0.0001). Double-staining revealed that in AS valves 27.2 ± 9.8% of cells were citH3/MPO- and 25.3 ± 8.9% citH3/NE-positive. Within stenotic valves, 6.4 ± 2.5% of cells showed citH3/CD68 double-positivity and were identified as macrophages. AS patients compared to controls had 83% higher plasma citH3 (p < 0.0001). In AS, plasma citH3 correlated with IL-6 (r = 0.39, p = 0.0054) levels and AVA (r = -0.45, p = 0.0009).

Conclusions: The presence of NETs in stenotic valves and association with AS severity suggest novel mechanisms involved in the disease progression.

Keywords: Aortic stenosis; Citrullinated histones; Myeloperoxidase; Neutrophil elastase; Neutrophil extracellular traps.