De Novo Valve Tissue Morphology Following Bioscaffold Mitral Valve Replacement in a Juvenile Non-Human Primate Model

Figures

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  Figure 1. Porcine Small Intestinal Submucosa (PSIS) hand-made mitral valve schematic and implantation points. The PSIS sheets are made into mitral valves (A) consisting of three main areas of (B) interest, the leaflets, annulus and the “legs”. Once implanted (C), the leaflets are the components that open and close as the heart is being pumped. The annulus is a thicker area of the valve that allows the opening for blood to flow through and divides the left atrium and left ventricle. The distal limbs or “legs” of the PSIS mitral valves are attached to the papillary muscles and heart wall when implanted (C), functioning similar to the chordae tendineae. The blue lines on the hand-made PSIS mitral valve signify the sutures (A,C). Panels (A,C) are reused by permission of Oxford University Press [29].

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  Figure 2. Porcine Small Intestinal Submucosa (PSIS) hand-made mitral valve explants. PSIS mitral valves (A) before implantation and (B) 3 months, (C) 11 months, and (D) 20 months post-implantation. There was (B–D) new tissue formation clearly observed on the explanted PSIS valves compared to the (A) original PSIS bioscaffold before implantation.

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  Figure 3. Native juvenile baboon mitral valve explant at 14-months stained with H&E. Histological image of native mitral valve leaflets (A,B) at 40× demonstrating the normal presence of cells on the leaflet. The label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 4. PSIS mitral valve explant at 3-month post-implantation in juvenile baboon stained with H&E. Mitral valve leaflet at lower magnification of 10× (A; H&E) with underlying myocardium. Higher magnification (B-40×) showing presence of interstitial cells and endothelium. In (A), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 5. PSIS mitral valve explant at 11-month post implantation in juvenile baboon stained with H&E. Mitral leaflet and annulus at 1.25× (A–C) with calcification (A-arrow) and chronic inflammatory response including multinucleated giant cells (C-10×) but with cellular infiltration of neo-leaflet (B-40×) as well. In (A), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 6. PSIS mitral valve explant at 20-month post implantation in juvenile baboon stained with H&E. Mitral valve leaflet and annulus at 1.25× (A–C) with neo-endocardial tissue growth (asterisk; B-20×) and calcification and chronic inflammation at the annulus (C-10×). In (A), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 7. Native juvenile baboon mitral valve explant and PSIS mitral valve explants staining for α-SMA. Explants of the native mitral valve leaflets with the presence of α-SMA (smooth muscle cells) in brown (A,B; 20×) was observed throughout the leaflets but was predominant on the ventricularis (v) side. Presence of α-SMA (smooth muscle cells) in brown was also seen at the fragmented edge of the mitral valve leaflet (C-40×) explanted at 3-month post-implantation. The 11-month (D-1.25×, E-20×) and 20-month (F-10×) explants had the preferential presence of α-SMA (smooth muscle cells) stained in brown on the ventricular surface but with a distribution that occurred till its centerline location, similar to the native baboon leaflet. In (A–D,F), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 8. Native juvenile baboon mitral valve explant and PSIS mitral valve explants staining for CD31. The native leaflets had a presence of CD31 (endothelial cells) in brown that lined at the outer surface of the leaflet (A-40×, B-20×). For the 11-month explant, the presence of CD31 positive staining was present at the central region of leaflet (C-40×). Finally, the 20-month explant showed the presence of CD31 positive staining distribution at the surface of the leaflet (D-20×) similar to the native leaflet.

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  Figure 9. Movat’s histological stain to depict the ECM components of native juvenile baboon and PSIS mitral valve explants. The native mitral valve leaflet (A-10×) and the 3-month PSIS explant (B-2×) exhibited matrix consisting of collagen, elastin, and proteoglycans (yellow/green, black, and blue, respectively). Notable, there was evidence of elastin deposition along the atrialis layer (A-arrow). Both the 11- (C-1.25×) and 20-month (D-1.25×) PSIS explants exhibited similar matrix components (collagen, elastin and proteoglycans), but with the added presence of fibrin (dark pink). In (A–D), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.

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  Figure 10. Movat’s histological stain showing the ECM components of a raw PSIS bioscaffold (Cormatrix, Roswell, GA, USA). The decellularized bioscaffold was predominantly composed of collagen (yellowish) with small vessels (black arrows) and scattered spindle cells (blue arrows; 20×). Spindle cells are fibroblasts that are long in shape and are a natural part of the body’s response to injury. These spindle cells are present before implantation.

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  Figure 11. Movat’s histological stain showing the ECM components of an acellular PSIS bioscaffold (Cormatrix, Roswell, GA, USA). This version two, acellular bioscaffold, was predominantly composed of collagen (yellowish) with small vessels (black arrows) and scattered spindle cells (blue arrows; 20×). Spindle cells are fibroblasts that are long in shape and are a natural part of the body’s response to injury. These spindle cells are present before implantation. This version of PSIS has fewer cells than the decellularized version 1 seen in Figure 10, but the cells are still not completely removed.

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  Figure 12. Spatial intensity maps to determine the ECM percent content. The movat’s histological stain of the native mitral valve (A-10×) is shown to demonstrate the spatial intensity maps computed for each image collected. Here, specifically, the matrix proteins of collagen (B), elastin (C), and proteoglycans (D) (yellow/green, black, and blue, respectively), were found to be 54%, 15%, and 31%, respectively, of each ECM protein. The collective data are shown in Table 1, Table 2 and Table 3 for the leaflets, annulus, and neochordae locations, respectively. In (A), the label, “A” depicts the atrialis side while the label “V” indicates the ventricularis side of the valve.