Abstract Body (Do not enter title and authors here): Introduction: Fibrocalcific remodeling is an end-stage feature of bioprosthetic (BP) structural valve degeneration and calcific aortic valve (AV) disease. However, the processes governing BP calcification are understudied. Here we conduct a histopathological assessment of BP degeneration in the aortic position and build a proteomic comparison map of BP degeneration versus AV disease in humans.
Methods: Macroscopic segmentation was performed on entire explanted degenerated bovine pericardial BP leaflets (n=48) and diseased AV valves (n=19) and validated with histology to classify their state (BP: non-degenerated/thrombotic/neotissue/calcified, AV: non-diseased/fibrotic/calcified). Segment-specific bulk mass spectrometry-based proteomics was performed on BP/AV tissues. Laser capture microdissection enabled spatially resolved proteomics of BP calcification within discrete bioprosthetic/thrombotic/neotissue matrices versus explanted non-calcified regions.
Results: Principal component analysis revealed BP and AV proteomes (2,005 and 2,012 proteins) clustered according to their degenerated and diseased segments; BP thrombotic and BP calcified sample clusters overlapped (Fig.A). Correlations of segment proteome-wide abundances revealed the highest intra-tissue similarity between non-degenerated BP and calcified BP (r_p=0.87). The highest inter-tissue similarity was found between BP neotissue and calcified AV (r_p=0.69) (Fig.B). Histopathological observations supported these proteomic findings, confirming the presence of calcification within discrete BP matrices: bioprosthetic (22/59), thrombus (22/59), and neotissue (15/59) (Fig.C). Laser capture microdissection revealed that 252 proteins were enriched in either bioprosthetic, thrombotic, or neotissue matrix calcification compared to explanted non-calcified regions. Only 3% of differentially enriched proteins overlapped, suggesting different mechanisms (Fig.D).
Conclusions: This is the first comparative proteomic study of segmented degenerated BP and diseased AV tissue. We identified 3 subtypes of BP calcification within bioprosthetic, thrombotic, or neotissue matrices. Spatial proteomics revealed different proteins associated with the calcification of these matrices.