Strategies beyond Decellularization for Optimal Tissue Engineering of Cardiac Xenografts

So Young Kim; Gi Beom Kim; Hong Gook Lim*; Yong Jin Kim

**So Young Kim, Gi Beom Kim, Hong Gook Lim* and Yong Jin Kim**

Volume5-Issue9
Dates: Received: 2024-09-25 | Accepted: 2024-09-30 | Published: 2024-09-30
Pages: 1253-1265

Abstract

The immunogenic carbohydrates, known as xenoantigens such as the Galα1-3Gal (α-Gal) epitope, and the non-human sialic acid N-glycolylneuraminic acid (Neu5Gc), play a major role in the immune response to xenotransplantation. Optimal decellularization protocols and strategies beyond decellularization are essential to eliminate xenoimmunogenicity. The aim of this study was to evaluate the safety and efficasy of our novel decellularization protocol and carbohydrase such as Peptide N-Glycosidase F (PNGase-F) for the removal of xenoantigens. We investigated the biomechanical properties, and efficacy for xenoantigen removal through expression of carbohydrate-binding lectins in porcine pericardium decellularized and treated with PNGase-F. Our novel decellularization protocols using 0.25% SDS + Triton X-100 through the multi-step methods with hypotonic, isotonic and hypertonic buffer solution demonstrated complete decellularization without histological changes. There were no histological changes depending on PNGase-F treatment concentration (0~2 unit/ml). There were no significant differences in tensile stress, tensile displacement, tensile strain at break, and permeability tests among porcine pericardia treated with different concentrations of PNGase-F. PNGase-F-treated porcine pericardium was stained with Jacalin, MAL I, WGA, RCA, GSL, ECA, PNA, SBA, WFA, and DSL, and showed lower fluorescence than native pericardium. PNGase-F treatment also resulted in a significant inhibition of lectin binding levels in a concentration-dependent manner. The fluorescent signal of decellularization effectively decreased lectin expression. Additional PNGase-F treatment for decellularization significantly reduced lectin expression without differences in PNGase-F concentration, demonstrating the synergistic effect of decellularization and PNGase-F. In conclusion, our novel decellularization protocols combined with PNGase-F treatment effectively removed the carbohydrates-associated xenoimmunogenicity and maintained biomechanical stability for cardiac xenografts. Our strategies beyond decellularization for optimal tissue engineering of cardiac xenografts can improve the biocompatibility of the graft.

FullText HTML FullText PDF DOI: 10.37871/jbres2012

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How to cite this article

Kim SY, Kim GB, Lim HG, Kim YJ. Strategies beyond Decellularization for Optimal Tissue Engineering of Cardiac Xenografts. J Biomed Res Environ Sci. 2024 Sept 30; 5(9): 1253-1265. doi: 10.37871/jbres2012, Article ID: JBRES2012, Available at: https://www.jelsciences.com/articles/jbres2012.pdf

Subject area(s)

Biomedical Science

Cellular Biology

References

Choi SY, Jeong HJ, Lim HG, Park SS, Kim SH, Kim YJ. Elimination of alpha-gal xenoreactive epitope: alpha-galactosidase treatment of porcine heart valves. J Heart Valve Dis. 2012 May;21(3):387-97. PMID: 22808845.
Jeong HJ, Adhya M, Park HM, Kim YG, Kim BG. Detection of Hanganutziu-Deicher antigens in O-glycans from pig heart tissues by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Xenotransplantation. 2013 Nov-Dec;20(6):407-17. doi: 10.1111/xen.12045. Epub 2013 Jul 16. PMID: 23855430.
Salama A, Evanno G, Harb J, Soulillou JP. Potential deleterious role of anti-Neu5Gc antibodies in xenotransplantation. Xenotransplantation. 2015 Mar-Apr;22(2):85-94. doi: 10.1111/xen.12142. Epub 2014 Oct 13. PMID: 25308416.
He W, Wang Q, Tian X, Pan G. Recapitulating dynamic ECM ligand presentation at biomaterial interfaces: Molecular strategies and biomedical prospects. Exploration (Beijing). 2022 Jan 24;2(1):20210093. doi: 10.1002/EXP.20210093. PMID: 37324582; PMCID: PMC10191035.
Liu W, Zhang X, Jiang X, Dai B, Zhang L, Zhu Y. Decellularized extracellular matrix materials for treatment of ischemic cardiomyopathy. Bioact Mater. 2023 Nov 29:33:460-482. doi: 10.1016/j.bioactmat.2023.10.015. PMID: 38076651; PMCID: PMC10697850.
WY, Wang WH, Xu PY, Kankala RK, Chen AZ. Decellularised extracellular matrix-based injectable hydrogels for tissue engineering applications. Biomaterials Translational. 2024;5(2):114-128. doi: 10.12336/biomatertransl.2024.02.003.
Lim HG, Kim SH, Choi SY, Kim YJ. Anticalcification effects of decellularization, solvent, and detoxification treatment for genipin and glutaraldehyde fixation of bovine pericardium. Eur J Cardiothorac Surg. 2012 Feb;41(2):383-90. doi: 10.1016/j.ejcts.2011.05.016. Epub 2011 Dec 12. PMID: 21683607.
Park S, Kim SH, Lim HG, Lim C, Kim YJ. The Anti-calcification Effect of Dithiobispropionimidate, Carbodiimide and Ultraviolet Irradiation Cross-linking Compared to Glutaraldehyde in Rabbit Implantation Models. Korean J Thorac Cardiovasc Surg. 2013 Feb;46(1):1-13. doi: 10.5090/kjtcs.2013.46.1.1. Epub 2013 Feb 6. PMID: 23424053; PMCID: PMC3573159.
Pagoulatou E, Triantaphyllidou IE, Vynios DH, Papachristou DJ, Koletsis E, Deligianni D, Mavrilas D. Biomechanical and structural changes following the decellularization of bovine pericardial tissues for use as a tissue engineering scaffold. J Mater Sci Mater Med. 2012 Jun;23(6):1387-96. doi: 10.1007/s10856-012-4620-8. Epub 2012 Mar 28. PMID: 22454140.
Jang W, Kim YJ, Choi K, Lim HG, Kim WH, Lee JR. Mid-term results of bioprosthetic pulmonary valve replacement in pulmonary regurgitation after tetralogy of Fallot repair. Eur J Cardiothorac Surg. 2012 Jul;42(1):e1-8. doi: 10.1093/ejcts/ezs219. Epub 2012 May 4. PMID: 22561653.
Kim GB, Kwon BS, Lim HG. First in human experience of a new self-expandable percutaneous pulmonary valve implantation using knitted nitinol-wire and tri-leaflet porcine pericardial valve in the native right ventricular outflow tract. Catheter Cardiovasc Interv. 2017 Apr;89(5):906-909. doi: 10.1002/ccd.26910. Epub 2017 Feb 1. PMID: 28145041.
Kim GB, Song MK, Bae EJ, Park EA, Lee W, Lim HG, Kim YJ. Successful Feasibility Human Trial of a New Self-Expandable Percutaneous Pulmonary Valve (Pulsta Valve) Implantation Using Knitted Nitinol Wire Backbone and Trileaflet α-Gal-Free Porcine Pericardial Valve in the Native Right Ventricular Outflow Tract. Circ Cardiovasc Interv. 2018 Jun;11(6):e006494. doi: 10.1161/CIRCINTERVENTIONS.118.006494. PMID: 29871940.
Lee SY, Kim GB, Kim SH, Jang SI, Choi JY, Kang IS, Kim YH. Mid-term outcomes of the Pulsta transcatheter pulmonary valve for the native right ventricular outflow tract. Catheter Cardiovasc Interv. 2021 Nov 1;98(5):E724-E732. doi: 10.1002/ccd.29865. Epub 2021 Jul 6. PMID: 34227733.
Shamis Y, Patel S, Taube A, Morsi Y, Sbarski I, Shramkov Y, Croft RJ, Crawford RJ, Ivanova EP. A new sterilization technique of bovine pericardial biomaterial using microwave radiation. Tissue Eng Part C Methods. 2009 Sep;15(3):445-54. doi: 10.1089/ten.tec.2008.0350. PMID: 19563253.
Nam J, Choi SY, Sung SC, Lim HG, Park SS, Kim SH, Kim YJ. Changes of the Structural and Biomechanical Properties of the Bovine Pericardium after the Removal of α-Gal Epitopes by Decellularization and α-Galactosidase Treatment. Korean J Thorac Cardiovasc Surg. 2012 Dec;45(6):380-9. doi: 10.5090/kjtcs.2012.45.6.380. Epub 2012 Dec 7. PMID: 23275920; PMCID: PMC3530722.
Min BJ, Kim YJ, Choi JW, Choi SY, Kim SH, Lim HG. Histologic Characteristics and Mechanical Properties of Bovine Pericardium Treated with Decellularization and α-Galactosidase: A Comparative Study. Korean J Thorac Cardiovasc Surg. 2012 Dec;45(6):368-79. doi: 10.5090/kjtcs.2012.45.6.368. Epub 2012 Dec 7. PMID: 23275919; PMCID: PMC3530721.
Seong YW, Kim YJ, Kim SH, Min BJ, Lee YO, Lim HG. Effect of trypsin on physico-dynamic and histological changes after decellularization of bovine pericardium. Korean J Thorac Cardiovasc Surg 2010;43:565-75. doi: 10.5090/kjtcs.2010.43.6.565.
Park CS, Kim YJ, Sung SC, Park JE, Sun-Young C, Woong-Han K, Kyung-Hwan K. Study on an effective decellularization technique for cardiac valve, arterial wall and pericardium xenographs: Optimization of decellularization. Korean J Thorac Cardiovasc Surg. 2008;41:550-62.
Sung SC, Kim YJ, Choi SY, Park JE, Kim KH, Kim WH. A study on an effective decellularization technique for a xenograft cardiac valve: The effect of osmotic treatment with hypotonic solution. Korean J Thorac Cardiovasc Surg. 2008; 41:679-686.
Jang WS, Kim YJ, Kim SH. Effects on tensile strength and elasticity after treatment with glutaraldehyde, solvent, decellularization and detoxification in fresh bovine pericardium. Korean J Thorac Cardiovasc Surg. 2010;43:1-10. doi: 10.5090/kjtcs.2010.43.1.1.
Lim HG, Choi SY, Yoon EJ, Kim SH, Kim YJ. In vivo efficacy of alpha-galactosidase as possible promise for prolonged durability of bioprosthetic heart valve using alpha1,3-galactosyltransferase knockout mouse. Tissue Eng Part A. 2013 Nov;19(21-22):2339-48. doi: 10.1089/ten.TEA.2013.0062. Epub 2013 Jun 18. PMID: 23672462.
Lim HG, Choi SY, Jeong S, Shin JS, Park CG, Kim YJ. In vivo efficacy for novel combined anticalcification treatment of glutaraldehyde-fixed cardiac xenograft using humanized mice. J Biomater Appl. 2015 Feb;29(7):929-40. doi: 10.1177/0885328214552710. Epub 2014 Sep 22. PMID: 25245780.
Kim GB, Lim HG, Kim YJ, Choi EY, Kwon BS, Jeong S. Novel self-expandable, stent-based transcatheter pulmonic valve: a preclinical animal study. Int J Cardiol. 2014 Apr 15;173(1):74-9. doi: 10.1016/j.ijcard.2014.02.005. Epub 2014 Feb 20. PMID: 24588954.
Lim HG, Kim GB, Jeong S, Kim YJ. Development of a next-generation tissue valve using a glutaraldehyde-fixed porcine aortic valve treated with decellularization, α-galactosidase, space filler, organic solvent and detoxification. Eur J Cardiothorac Surg. 2015 Jul;48(1):104-13. doi: 10.1093/ejcts/ezu385. Epub 2014 Oct 14. PMID: 25315752.
Kim CY, Kim KH, Moon KC, Kim WH, Sung SC, Kim YJ. Comparison of different methods of aortic valve conduit xenograft preservation in an animal experiment model; fresh cryopreservation versus acellularized cryopreservation. Korean J Thorac Cardiovasc Surg. 2010;43:11-9. doi: 10.5090/kjtcs.2010.43.1.11.
Park CS, Kim YJ, Lee JR, Lim HG, Chang JE, Jeong S, Kwon N. Anticalcification effect of a combination of decellularization, organic solvents and amino acid detoxification on glutaraldehyde-fixed xenopericardial heart valves in a large-animal long-term circulatory model. Interact Cardiovasc Thorac Surg. 2017 Sep 1;25(3):391-399. doi: 10.1093/icvts/ivx131. PMID: 28505294.
Kim DJ, Kim YJ, Kim WH, Kim SH. Xenograft Failure of Pulmonary Valved Conduit Cross-linked with Glutaraldehyde or Not Cross-linked in a Pig to Goat Implantation Model. Korean J Thorac Cardiovasc Surg. 2012 Oct;45(5):287-94. doi: 10.5090/kjtcs.2012.45.5.287. Epub 2012 Oct 9. PMID: 23130301; PMCID: PMC3487011.
Lim HG, Jeong S, Kim GB, Lee W, Son KH, Kim YJ. Next-generation transcatheter aortic valve implantation. JTCVS Open. 2020 Jun 24;3:14-24. doi: 10.1016/j.xjon.2020.06.008. PMID: 36003870; PMCID: PMC9390526.
Macher BA, Galili U. The Galalpha1,3Galbeta1,4GlcNAc-R (alpha-Gal) epitope: a carbohydrate of unique evolution and clinical relevance. Biochim Biophys Acta. 2008 Feb;1780(2):75-88. doi: 10.1016/j.bbagen.2007.11.003. Epub 2007 Nov 22. PMID: 18047841; PMCID: PMC2271034.
Varki A. Biological roles of oligosaccharides: all of the theories are correct. Glycobiology. 1993 Apr;3(2):97-130. doi: 10.1093/glycob/3.2.97. Update in: Glycobiology. 2017 Jan;27(1):3-49. doi: 10.1093/glycob/cww086. PMID: 8490246; PMCID: PMC7108619.