(F1104) ENHANCED RECELLULARIZATION OF DECELLULARIZED XENOANTIGEN FREE SCAFFOLDS WITH HUMAN MESENCHYMAL STEM CELLS AND HUMAN UMBILICAL ENDOTHELIAL CELLS

Abstract: Removal for major immunogenic xenoantigens of the Galα1-3Gal (α-Gal) epitope and the non-human sialic acid N-glycolylneuraminic acid (Neu5Gc) are essential to eliminate xenoimmunogenicity and optimize recellularization for cardiac xenografts. The aim of this study was to evaluate the safety and efficacy of α-galactosidase for removal of α-Gal xenoantigen and PNGase-F for removal of non-α-Gal xenoantigen combined with optimal decellularization, and the potential of in vitro recellularization was assessed with co-culturing human adipose-derived stem cells and human umbilical vein endothelial cells for xenoantigen free cardiac xenografts. We investigated the mechanical properties, and efficacy for xenoantigen removal with expression of carbohydrate-binding lectins in porcine pericardium decellularized and treated with α-Gal and PNGase-F. In the H&E stain, there were no histological changes depending on α-Gal and PNGase-F treatment. There was no difference in tensile stress, tensile displacement, tensile strain at break, and permeability test following enzymatic treatments. Both enzyme-treated xenografts were stained with all lectins, and showed synergistic effects for low fluorescence qualitatively and quantitatively. The enzymatic treatments for decellularization significantly reduced lectin expression, demonstrating the synergistic effect of both enzymes and decellularization. During recellularization, both enzyme treated xenografts with decellularized were entirely repopulated in 28 days and detected DAPI-positive cells. In addition, they were observed expression of fibronectin as an important factor in remodeling of ECM. In vitro recellularization for decellularized and both enzymes-treated xenografts was assessed with vimentin, calponin, fibronectin and CD31staining. Stronger signals were detected in decellularized xenografts, and decellularized xenografts treated with both enzymes showed significantly faster mesenchymal cell infiltration into the tissue, leading to accelerated recellularization. We have successfully produced xenoantigen-free scaffolds by demonstrating the safety and the synergistic effect of α-Gal and PNGase-F treatments, and proved effective recellularization for the xenoantigen-free scaffolds.

Funding Source: This study was supported by SNUH Lee Kun‐hee Child Cancer & Rare Disease Project, Republic of Korea (Grant Number: 23C0220100).

Clinical Trial ID number: No