Characterization of Laminins in Healthy Human Aortic Valves and a Modified Decellularized Rat Scaffold

Figures

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  FIG. 1.. Histology of donor aortic valve cusps. (A–C) H&E. (D–F) Collagen I (green) and elastin (red), nuclei counterstained with DAPI (blue). All three cusps had an intact three-layered structure and an overall normal histology. Elastin identifies the ventricularis (D–F), as well as the interstitial vessels (D). A representative image of four (A–C) or one (D–F) tissue sections is shown. All images were captured with 20 × magnification. Scale bar represents 100 μm. DAPI, 4,6-diamidino-2-phenylindole; NCC, noncoronary cusp; RCC, right coronary cusp; LCC, left coronary cusp; H&E, Hematoxylin and Eosin.

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  FIG. 2.. Laminin chain expression in human aortic valves. The endothelial basement membrane covering the valve cusps were positive for laminin α4, α5, β1, β2, and γ1, as well as the endothelial lining of interstitial vessels and minor patches in the interstitium. Images of α1, α2, α3, β3, and γ2 show absence of immunoreactivity. A representative image of ≥3 tissue sections from each of the three donor valve cusps is shown. All images shown at 20 × magnification. Scale bar represents 100 μm.

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  FIG. 3.. Expression of laminin α4, α5, and γ1 in rat aortic valves. The endothelial basement membrane stained positive for all three laminin chains. Minimal immunoreactivity was seen in the interstitium. A representative image of ≥4 tissue sections from three animals per group is shown. All images captured with 20 × magnification. Scale bar represents 100 μm.

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  FIG. 4.. Histology of normal and decellularized rat aortic valves. H&E images and DAPI staining of cell nuclei reveal absence of intact nuclei after decellularization. Laminin γ1, collagen IV, perlecan, heparan sulfate, and fibronectin was visualized in both normal controls and decellularized valves, with a largely preserved distribution pattern after decellularization. A representative image of ≥3 tissue sections from three animals per group is shown. All images were captured at 20 × magnification. Scale bar represents 100 μm. LNγ1, laminin γ1; COL IV, collagen IV; PLN, perlecan; HS, heparan sulfate; FN, fibronectin.

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  FIG. 5.. Quantification of DNA in rat aortic valves before and after decellularization. (A) DNA content in normal controls (n = 6) and decellularized valves treated with SDS for 3 h (n = 3), 12 h (n = 3), or 3 h with additional treatment with BE (n = 3). (B, C) Dissection technique for excision of intact valve cusps in normal control (B) and decellularized aortic root (C). *p < 0.05. **p < 0.0001. SDS, sodium dodecyl sulfate; BE, benzonase endonuclease.