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Identification of potentially immunogenic proteins in porcine cholecyst extracellular matrix

Tissue engineering scaffolds of animal origin have many medical uses but the residual immunogenicity can significantly shadow their potential clinical applications as tissue grafts. Therefore, there is a continued search for non-immunogenic scaffolds of animal origin. Although the phylogenetic proximity of the donor species and the decellularization protocols are important, the native load of immunogens in the extracellular matrices of the source organ is a major determinant of residual immunogenicity. Recent observations indicate that, as a tissue graft, porcine cholecyst extracellular matrix (CEM) induces a constructive tissue remodelling reaction rather than a pro-inflammatory graft rejection reaction. The objective of the study was to identify potentially immunogenic proteins in the extractable proteins of CEM. Proteins were extracted and separated by 2D-gel electrophoresis and probed with hyper-immune serum developed against CEM in Sprague-Dawley rats. The reactive proteins were identified by Mass Spectrometry and Uniprot database search. The study detected 18 potentially immunogenic proteins in the extract but none of them was of extracellular matrix origin. There were sixteen cellular and two secretary proteins. Among the cellular immunogenic proteins, lactate dehydrogenase, vimentin, actin, ATP synthase beta chain and annexin-A2 were previously reported as xeno-antigens contributing for the graft rejection. The extract also contained Caviolin-1, a risk factor critical for renal transplant rejection. The results of the study warrant detailed study on the identification of immunogenic proteins in the context of evaluating the biomaterial properties of mammalian-derived scaffolds intended for regenerative medical applications.