Table 1.
Overview of patient group characteristics.
Fig 1.
Determination of decellularization extent in NA as well as aneurysmal/dissection tissue of different origin.
In (A) upper row, the native tissues of the corresponding groups are shown. In blue the cell nuclei are depicted and in red the plasma membrane is shown by WGA staining. In the row below, representative images of corresponding decellularized tissues are depicted. Scale bar represents 50μm. (B) Quantification of cell nuclei and (C) WGA positive staining are shown using box plots with mean and standard deviation. NA n = 9; BAV-aTAA n = 9; TAV-aTAA n = 9; AAD type A n = 8. * p<0.05, ** p<0.01, *** p<0.001.
Fig 2.
Quantification of ECM components of NA and aneurysmal/dissection tissues.
(A) Representative images of the auto-fluorescence properties of elastic fibers and other ECM components to quantify the ECM of native and decellularized tissue are shown (elastic fibers with stronger auto-fluorescence in GFP-B channel compared to residual ECM). Scale bar represents 50μm. In (B) and (C) the quantification of elastic fiber area as well as ECM area without elastic fibers are shown in box plots with mean and standard deviation. Box plots in (D) show the quantification of elastin expression in native as well as decellularized NA and aneurysmal/dissection tissue. (E) shows representative images of antibody specific elastin staining (shown in red) in native and decellularized NA and aneurysmal/dissection tissue. Cell nuclei depicted in blue are stained with Hoechst. NA n = 9; BAV-aTAA n = 9 and for elastin n = 8; TAV-aTAA n = 9; AAD type A n = 8. * p<0.05, ** p<0.01, *** p<0.001.
Fig 3.
Elastika van Gieson and Masson Trichrome Goldner stained aortic NA and aneurysmal/dissection tissue.
In (A) representative images of Elastika van Gieson stained NA and aneurysmal aortic tissue, either native (upper row) or decellularized (with 2.5% NaOH; lower row), are shown as well as magnified areas of the representative images. Yellow asterisks represent muscle cells and yellow arrowheads elastic fibers. In (B) representative images as well as magnified areas of Masson Trichrome Goldner stained aortic tissue are shown. The upper row shows native tissue of NA specimen, aneurysmal tissue of BAV- or TAV-origin as well as dissection specimen. Black arrowheads indicate cellular material. In the row below the corresponding decellularized tissues are depicted. Scale bar represents 50μm and in the magnified areas 20μm.
Fig 4.
Determination of aortic collagen content before and after decellularization.
In (A) representative images of Picrosirius red stained aortic NA and aneurysmal/dissection tissue after image acquisition using a fluorescence microscope are depicted (collagen in red, elastic fibers and cellular material in green). The upper row shows the native tissue und the lower the tissue after decellularization. Scale bar represents 50μm. (B) shows quantification of medial collagen content in native and decellularized NA tissue as well as aneurysmal/dissection tissue. NA n = 9; BAV-aTAA n = 9; TAV-TAA n = 9; AAD type A n = 8. * p<0.05, ** p<0.01.
Fig 5.
SEM based evaluation of elastic fiber sheets and other ECM components of NA and aneurysmal/dissection tissues.
In (A) representative SEM pictures of native (upper row) and decellularized tissue (lower row) of controls (NA) and BAV-aTAAs are shown. In (B) SEM pictures of native and decellularized TAV-aTAAs tissue as well as tissue from a patient with an aortic dissection (AAD type A) are shown. White arrows are indicating elastic fibers, white asterisks are indicating cellular material, and white arrowheads are indicating interlaminar elastin fibers.
Fig 6.
Comparison of SEM pictures with histological stainings and higher magnification pictures to show elastic fiber sheets after decellularization.
In (A) the focal medial degeneration (white arrows) of TAV-aTAAs is depicted by SEM images as well as in Elastika van Gieson stained tissue. Scale bar in the Elastika van Gieson staining represents 250μm. (B) Representative SEM images of NA and aneurysmal tissue after decellularization (white arrows are indicating elastic fiber disks). Int = intima; Med = media; Adv = adventitia.
Fig 7.
Representative images indicating the dissection lamellae and the ECM structure of aortic dissection specimens aside from the dissection lamellae.
In (A) overview pictures of the dissection lamellae (white/black arrows) before and after decellularization are shown using SEM-based image acquisition and the corresponding tissue specimen after Elastika van Gieson staining. (B) Representative SEM images of the ECM structure in native and decellularized dissection tissue aside from the dissection lamellae and the corresponding pictures of Elastika van Gieson staining. White scale bar represents 200μm and black scale bar 250μm. Scale bar in the small pictures (magnification x40) represents 50μm. Int = intima; Med = media; Adv = adventitia.