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. 2025 Aug 12;16:1655946. doi: 10.3389/fphys.2025.1655946

FIGURE 5.

Histological analysis and graphs illustrate collagen and elastin content in arterial samples from four conditions: Healthy, LR, LR+GM, and WB. Panels A to E show stained sections under different techniques. Panels F to N present bar graphs comparing medial and adventitial collagen, elastin content, medial thickness, and birefringence ratio across conditions. Significant differences are marked with asterisks indicating p-values.

Histological analysis of extracellular matrix changes in healthy and T/HS aortic tissue. Representative images of histological staining using picrosirius/hematoxylin under (A) brightfield & (B) polarized light, (C) Verhoeff’s staining method, (D) Miller’s staining method in healthy donor aorta and T/HS aortic tissues resuscitated with LR, LR+GM, and WB are shown. (E) Representative images of elastic fibers (magenta) via two-photon excitation fluorescence and collagen fibers (green) via second-harmonic generation of Healthy, LR, LR+GM, and WB vascular tissue were generated. The percentages of collagen (F, G) and elastin (H, I) in the aortic media and adventitia were quantified, along with the thickness of the medial (J), adventitial (K), and interlamellar (L) layers. Percentage of straight (non-undulated) elastic fibers in the medial layer was also measured (M), along with the ratio of red to green birefringence collagen fibers in the polarized area (N). Images were taken at ×40 magnification with lumen at the bottom of frame (scale bar = 25 μm). Data are presented as mean ± SEM. By one-way ANOVA, * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 for the marked groups.