Department of Experimental Medicine, Research Institute for Complex Issues of Cardiovascular Diseases, 6 Sosnovy Boulevard, Kemerovo 650002, Russia.
Int J Mol Sci. 2023 Sep 2;24(17):13602. doi: 10.3390/ijms241713602.
Hitherto, calcified aortic valves (AVs) and failing bioprosthetic heart valves (BHVs) have been investigated by similar approaches, mostly limited to various immunostaining techniques. Having employed multiple immunostaining combinations, we demonstrated that AVs retain a well-defined cellular hierarchy even at severe stenosis, whilst BHVs were notable for the stochastic degradation of the extracellular matrix (ECM) and aggressive infiltration by ECM-digesting macrophages. Leukocytes (CD45) comprised ≤10% cells in the AVs but were the predominant cell lineage in BHVs (≥80% cells). Albeit cells with uncertain immunophenotype were rarely encountered in the AVs (≤5% cells), they were commonly found in BHVs (≥80% cells). Whilst cell conversions in the AVs were limited to the endothelial-to-mesenchymal transition (represented by CD31α-SMA cells) and the formation of endothelial-like (CD31CD68) cells at the AV surface, BHVs harboured numerous macrophages with a transitional phenotype, mostly CD45CD31, CD45α-SMA, and CD68α-SMA. In contrast to immunostaining, which was unable to predict cell function in the BHVs, our whole-specimen, nondestructive electron microscopy approach (EM-BSEM) was able to distinguish between quiescent and matrix-degrading macrophages, foam cells, and multinucleated giant cells to conduct the ultrastructural analysis of organelles and the ECM, and to preserve tissue integrity. Hence, we suggest EM-BSEM as a technique of choice for studying the cellular landscape of BHVs.
迄今为止，钙化的主动脉瓣（AV）和功能失调的生物瓣（BVH）一直采用类似的方法进行研究，这些方法大多局限于各种免疫染色技术。通过使用多种免疫染色组合，我们证明即使在严重狭窄的情况下，AV 仍然保留着明确的细胞层次结构，而 BVH 的特点是细胞外基质（ECM）的随机降解和 ECM 消化巨噬细胞的侵袭。白细胞（CD45）在 AV 中占细胞的≤10%，但在 BVH 中是主要的细胞谱系（≥80%的细胞）。虽然在 AV 中很少遇到免疫表型不确定的细胞（≤5%的细胞），但在 BVH 中很常见（≥80%的细胞）。虽然 AV 中的细胞转化仅限于内皮到间充质的转化（由 CD31α-SMA 细胞表示）和 AV 表面形成的内皮样（CD31CD68）细胞，但 BVH 中存在许多具有过渡表型的巨噬细胞，主要是 CD45CD31、CD45α-SMA 和 CD68α-SMA。与免疫染色不同，免疫染色无法预测 BVH 中的细胞功能，我们的整体标本、非破坏性电子显微镜方法（EM-BSEM）能够区分静止和基质降解的巨噬细胞、泡沫细胞和多核巨细胞，进行细胞器和 ECM 的超微结构分析，并保持组织完整性。因此，我们建议 EM-BSEM 作为研究 BVH 细胞景观的首选技术。
