Enrichment and spatial convergence of transthyretin, OxPL, plasminogen and Lp(a) in calcific aortic valve stenosis

Background Lipoprotein(a) [Lp(a)] and plasminogen (PLG) carry oxidized phospholipids (OxPL) that promote inflammatory and osteogenic signaling in calcific aortic valve stenosis (CAVS) and may link lipid oxidation, fibrinolysis, and transthyretin (TTR) amyloidogenesis. How these pathways are spatially organized within human valve tissue across histologic stages of CAVS remains incompletely defined. Objectives To determine whether lipid, oxidative, protease, and TTR-related processes are spatially enriched and co-localized within human aortic valve leaflets across histologic grades of CAVS and to evaluate associations with circulating oxidative biomarkers. Methods Aortic valve leaflets from 45 patients undergoing surgical replacement for severe CAVS were pathologically graded (Grade 1: mild thickening; Grade 2: fibrocalcific remodeling; Grade 3: nodular calcification; Grade 4: confluent calcification) and analyzed by immunohistochemistry for TTR, PLG, Lp(a), OxPL, autotaxin, and malondialdehyde (MDA)-lysine epitopes, with digital quantification of percent immunostaining area and semi-quantitative overlapping area scores. In an independent cohort of 30 patients with paired plasma and valve samples, circulating Lp(a), OxPL-apoB, OxPL-apo(a), OxPL-PLG, PLG, and MDA IgG/IgM autoantibodies were measured and correlated with tissue measures. Results All molecular targets were detectable within valve leaflets and demonstrated progressive expansion from early (Grades 1-2) to advanced (Grades 3-4) lesions. Percent immunostaining area increased approximately fourfold for Lp(a) and threefold for PLG and OxPL, while TTR exhibited the largest absolute area in advanced lesions (P<0.001 for all). Overlapping area scores increased stepwise across grades for all molecular pairs, with the most pronounced changes observed for OxPL/TTR and OxPL/PLG (all p < 0.001). OxPL area of immunostaining strongly correlated with TTR area (r=0.871, p < 0.001) and PLG area (r=0.785, p < 0.001). Conclusions Human aortic valve leaflets in CAVS demonstrate progressive enrichment and spatial convergence of TTR, PLG, Lp(a), and OxPL with increasing histologic severity, suggesting a pathophysiologically linked process contributing to advancing fibrocalcific leaflet pathology.