Fig 2: (A) Representative photographs of immunofluorescence staining showing S100A8 (marked with green) and DAPI (marked with blue) in healthy group and aSAH group, and quantitative analysis of positive cells. n = 3/group; p = 0.0033. (B) Representative photographs of immunofluorescence staining showing S100A9 (marked with red) and DAPI (marked with blue) in healthy group and aSAH group, and quantitative analysis of positive cells. n = 3/group; p = 0.0003. (C) Representative photographs of immunofluorescence staining showing SRPK1 (marked with yellow) and DAPI (marked with blue) in healthy group and aSAH group, and quantitative analysis of positive cells. n = 3/group; p = 0.0027. (D) Representative photographs of immunofluorescence staining showing ZNF281 (marked with yellow) and DAPI (marked with blue) in healthy group and aSAH group, and quantitative analysis of positive cells. n = 3/group; p = 0.0077. Healthy control: Healthy Middle meningeal arteries; Aneurysm: Ruptured aneurysm tissue. Scale bar: 75 µm (**p < 0.01, ***p < 0.001).

Fig 3: (A) Representative Western blot bands of S100A8, S100A9, SRPK1, and ZNF281 expression in blood samples in different groups. (B) Quantitative analysis of S100A8, S100A9, SRPK1, and ZNF281 expression in blood samples in different groups. n = 3/group. (C) Representative Western blot bands of S100A8, S100A9, SRPK1, and ZNF281 expression in arterial walls and aneurysms walls from different groups. (D) Quantitative analysis of S100A8, S100A9, SRPK1, and ZNF281 expression in arterial samples in different groups. n = 3/group. Healthy control: Healthy middle meningeal arteries; Aneurysm: Ruptured aneurysm tissue (*p < 0.05, **p < 0.01).

Fig 4: GSEA plot showing that a high expression of S100A8 and S100A9 is positively correlated with these significant signaling pathways. (A) In the group with high S100A8 expression in calcific aortic valve disease (CAVD), the enriched pathways included calcium signaling, cell adhesion molecules, chemokine signaling, cytokine–cytokine receptor interaction, FC epsilon RI signaling, Hedgehog signaling, hypertrophic cardiomyopathy, vascular smooth muscle contraction, and VEGF signaling. (B) In the group with a high expression of S100A9 in CAVD, the enriched pathways included arrhythmogenic right ventricular cardiomyopathy, cardiac muscle contraction, cell adhesion molecules, dilated cardiomyopathy, ERBB signaling, FC epsilon RI signaling, Hedgehog signaling, hypertrophic cardiomyopathy, leukocyte transendothelial migration, MAPK signaling, and vascular smooth muscle contraction.

Fig 5: Diagnostic performance of S100A8/S100A9 expression levels for calcific aortic valve disease (CAVD) patients in the datasets. (A–C) The expression levels of S100A8/S100A9 in CAVD were based on the GSE databases (GSE153555, GSE51472, and GSE83453). (D–F) Diagnostic value of S100A8 and S100A9 in GSE databases (GSE153555, GSE51472, and GSE83453). Receiver operating characteristic (ROC) curve of normal and CAVD valve tissues. ** P < 0.01, *** P < 0.001 when two groups were compared as indicated or compared with the corresponding control.