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. Author manuscript; available in PMC: 2021 Aug 1.
Published in final edited form as: Arterioscler Thromb Vasc Biol. 2020 May 28;40(8):1838–1853. doi: 10.1161/ATVBAHA.118.314087

Figure 6. The important role of S100A9 and RAGE in human diabetic calcified carotid arteries.

Figure 6.

(A) 25 human atheroma obtained from carotid endarterectomy. Staining for S100A9, RAGE, and ALP in human atheroma (n=25). One of low S100A9 % positive area (left panel) and high S100A9 % positive area (right panel) are shown. Scale bars: 100 μm. Each graph shows the correlation between percentage of S100A9 positive area and ALP- and RAGE-positive area. The average of 5–10 high power fields was used for the analysis. (B) Serial section of human atheroma double-immune stained with anti-S100A9 (red) to anti-RAGE or anti-ALP (green) antibodies. Yellow area in Merge indicate S100A9 positive cells for either RAGE or ALP. Scale bars: 20 μm. The data represent 5 donors that showed similar results. (C) Transmission electron microscopy–based immunogold staining of S100A9 (red arrows) in EVs from human carotid plaques. One of 3 plaques is shown. Scale bar: 300 nm. (D) Carotid plaque specimens were obtained from diabetic (n = 4) and non-diabetic patients (n = 4) undergoing carotid endarterectomy. Volcano plot for the proteomics data. Red and blue markers indicate the significantly enriched (p<0.05) proteins in diabetic patient group and non-diabetic patient group, respectively, with a fold change (FC) cutoff of 2.0. (E) The schematic of the potential mechanism for macrophage-mediated EV microcalcification in diabetes. S100A9-RAGE axis may regulate pro-inflammatory and pro-osteogenic macrophage activation via Nrf2 and NF-κB pathways.