Rapid anticalcification treatment for glutaraldehyde-fixed autologous tissue in cardiovascular surgery

Fig. 1. Annotation and representative histological features of the thoracic aorta. a Entire tissue area (black line), granulated tissue (blue line), and extent of calcification (red line) were annotated for each sample of thoracic aorta; area was measured in mm.2 and evaluated semiquantitatively. b Representative histological features of the explanted thoracic aorta. Granulation and calcification covered the tissue surface (stain, H&E; magnification, × 5)

Fig. 2. Tissue calcification and granulation. Explanted thoracic aorta was covered with granulated tissue composed of fibroblasts, capillary vessels, and inflammatory cells, such as lymphocytes and macrophages (yellow arrows). Collagen fibers of the adventitia were degenerated and considered calcified (red arrows) (stain, H&E; magnification, × 30)

Fig. 3. Changes in the tunica media. Microscopy findings showing almost preserved elastic arterial media. In areas with noticeable calcification, the outermost elastic fibers of the tunica media have disappeared (red arrows) (stain, VB; magnification, × 30)

Fig. 4. Semiquantitative analysis of calcification. Group 1 (control): untreated tissues; group 2: tissues fixed with glutaraldehyde (GA) for 3 min; group 3: tissues fixed with GA for 3 min and post-treated with ethanol for 10 min; group 4: tissues fixed with GA for 3 min and post-treated with ethanol for 20 min; group 5: tissues fixed with GA for 3 min and post-treated with ethanol for 30 min. Semiquantitative analysis of calcification revealed the calcified areas in the explants of groups 1, 2, 3, 4, and 5 to be 0.02 ± 0.04, 0.22 ± 0.15, 0.10 ± 0.07, 0.07 ± 0.03, and 0.08 ± 0.03 mm2, respectively. Calcification of explants from groups 1 (P = 0.0014), 4 (P = 0.0014), and 5 (P = 0.0031) was significantly lower than that of explants from group 2. Group 3 (P = 0.0503) showed slightly decreased calcification although the decrease was not statistically significant. Calcified areas are displayed as a bow-and-whiskers plot with median (thick line), 25th to 75th percentiles (box), 10th to 90th percentiles (whiskers), and outliers (circles). P-values were calculated using the Kruskal–Wallis test