The Efficiency of Decellularization of Bovine Pericardium by Different Concentrations of Sodium Dodecyl Sulfate

Anatoliy Sokol, Dmytro Grekov, Glib Yemets, Alexander Galkin, Nataliia Shchotkina, Arkadii Dovghaliuk, Nadiia Rudenko, Iliia Yemets


Background. In modern cardiovasclar surgery, it is a promising method to use xenotissues, which in their properties are close to human tissues, in order to restore the integrity of the heart chambers, its walls or valves. Decellularization of extracellular matrix (EMC) is applied in the process of creation of such bioprotheses. In EMC, the elastin and collagen components are preserved, and antigenic molecules are eliminated resulting in reduction the risk of rejection. The study is devoted to assessment of the histological, molecular-genetic and cytotoxic properties of decellularized bovine pericardium, processed with various concentrations of trypsin enzyme.

Objective. The aim of the work is evaluation of efficiency of bovine pericardial decellularization, based on the use of trypsin enzyme with 1% Sodium Dodecyl Sulfate (SDS) and 0.1% SDS.

Methods. Bovine pericardium was used as a biomaterial for decellularization. Decellularization protocol 1 envisages processing of samples with 0.25% Trypsin solution at 24 °C with constant shaking (200 rpm) along with processing with 1% ionic SDS detergent. The samples, prepared according to protocol 2, were processed with a low concentration of 0.1% SDS. Histological and morphological properties along with detection of nucleic acids concentration in the samples were studied. Matrix samples were cultured in a human fibroblasts cell culture suspension in order to determine cytotoxicity.

Results. Histological examination has not revealed any presence of cells in tissues, decellularized in accordance with both protocols. More than 99% of the nucleic acids was removed from decellularized bovine matrix. During our study, we have not observed cytotoxic effect in vitro for protocol 2 matrix samples, decellularized with only 0.1% SDS. Focal destruction of fibroblasts was observed in conditions of long-term cultivation in protocol 1 samples (Trypsin + 1% SDS). Cells formed abnormal morphological aggregates. Samples of this group have also demonstated sructural changes in collagen and elastin fibers.

Conclusions. Studies have shown that pericardial matrix tissue, decellularized with low-concentration of 0.1% SDS, has the same biological properties as the native pericardium. Decellularization of bovine pericardium, using trypsin enzyme with 1% SDS, has a cytotoxic effect on human cells.


Pericardium; Decellularization; Sodium dodecyl sulfate; Tissue engineering

Full Text:



Mallis P, Michalopoulos E, Dimitriou C, Kostomitsopoulos N, Stavropoulos-Giokas C. Histological and biomechanical characterization of decellularized porcine pericardium. Biomed Mater Eng. 2017;28:477-88. DOI: 10.3233/BME-171689

Lima EO, Ferrasi AC, Kaasi A. Decellularization of human pericardium with potential application in regenerative medicine. Arq Bras Cardiol. 2019;113(1):18-9. DOI: 10.5935/abc.20190130

Gonçalves A, Griffiths L, Anthony R, Orton C. Decellularization of bovine pericardium for tissue-engineering by targeted removal of xenoantigens. J Heart Valve Dis. 2005 Mar;14(2):212-7.

Naso F, Gandaglia A. Different approaches to heart valve decellularization: A comprehensive overview of the past 30 years. Xenotransplantation. 2018;25(1):1-10. DOI: 10.1111/xen.12354

Pawan KC, Yi H, Ge Z. Cardiac tissue-derived extracellular matrix scaffolds for myocardial repair: advantages and challenges. Regen Biomater. 2019;6(4):185-99. DOI: 10.1093/rb/rbz017

White LJ, Taylor AJ, Faulk DM, Keane TJ, Saldin LT, Reing JE, Swinehart IT, et al. The impact of detergents on the tissue decellularization process: a ToF-SIMS study. Acta Biomater. 2017;50:207-19. DOI: 10.1016/j.actbio.2016.12.033

Ramm R, Goecke T, Theodoridis K, Hoeffler K, Sarikouch S, Findeisen K, et al. Decellularization combined with enzymatic removal of N-linked glycans and residual DNA reduces inflammatory response and improves the performance of porcine xenogeneic pulmonary heart valves in an ovine in vivo model. Xenotransplantation. 2020;27(2):1-12. DOI: 10.1111/xen.12571

Helliwell JA, Thomas DS, Papathanasiou V,Homer-Vanniasinkam S, Desai A, Jennings LM, et al. Development and characterisation of a low-concentration sodium dodecyl sulphate decellularised porcine dermis. J Tissue Eng. 2017;8:1-12. DOI: 10.1177/2041731417724011

Schaner PJ, Martin ND, Tulenko TN, Shapiro IM, Tarola NA, Leichter RF, et al. Decellularized vein as a potential scaffold for vascular tissue engineering. J Vasc Surg. 2004;40(1):146-53. DOI: 10.1016/j.jvs.2004.03.033

Jelev L, Surchev L. A novel simple technique for en face endothelial observations using water-soluble media -’thinned-wall’ preparations. J Anat. 2008;212(2):192-7. DOI: 10.1111/j.1469-7580.2007.00844.x

Vunjak-Novakovic G, Freshney RI, editors. Culture of cells for tissue engineering. Wiley; 2006. 536 p.

Blazhevich OV. Cultivation of cells. Minsk: BSU; 2004. 78 p.

Golubev DB, Sominina AA, Medvedeva MN. Guide to the use of cell cultures in virology. Leningrad: Medicine; 1976. 224 p.

Roosens A, Somers P, DE Somer F, Carriel V, Van Nooten G, Cornelissen R. Impact of detergent-based decellularization methods on porcine tissues for heart valve engineering. Ann Biomed Eng. 2016 Sep;44:2827-39. DOI: 10.1007/s10439-016-1555-0

Korzhevsky DE. Application of hematoxylin in histological technique. Morphology. 2007;132(6):77-82.

Alturkistani HA, Tashkandi FM, Mohammedsaleh ZM. Histological stains: A literature review and case study. Global J Health Sci. 2016;8(3):72-9. DOI: 10.5539/gjhs.v8n3p72

Horobin RW. Biological staining: mechanisms and theory. Biotech Histochem. 2002;77(1):3-13.

Lilly R. Pathohistological technique and practical histochemistry. Moscow; 1969. 122 p.

Starodubtsev VB, Karpenko AA, Ignatenko PV, Kim IN, Zoloev DG, Gorbatykh VN. Immediate and long-term results of various methods of carotid bifurcation reconstruction. Angiol Sosud Khir. 2014;20(4):93-4.

Mueller C, Dave H, Prêtre R. Surgical repair of aorto-ventricular tunnel. Multimed Man Cardiothorac Surg. 2012;2012:1093-7. DOI: 10.1093/mmcts/mms006

Chesnov UM. Biocompatibility of xenopericardium fixed with epoxy compounds in in vitro and in vivo experiments. In: Manak NA, editor. Current issues of cardiology. vol. 2. Minsk: Encyclopedia; 2002. p. 188-90.

Nonaka M, Iwakura A, Yamanaka K. Technique to treat extensive abscesses in double valve replacement for prosthetic valve endocarditis. J Heart Valve Dis. 2013;22(4):575-7.

Akhmedov SD, Afanasyev SA, Dyakova ML, Fatkhutdinov TX, Kaktursky LV. Use of a cell-free matrix for the formation of new blood vessels and heart by tissue engineering. Cell Transplant Tissue Eng. 2009;4(2):32-9.

Gilbert WT, Sellaro LT, Badylak FS. Decellularization of tissues and organs. Biomaterials. 2006;27:3675-83. DOI: 10.1016/j.biomaterials.2006.02.014

Rieder E, Kasimir MT, Silberhumer G, Seebacher G, Wolner E, Simon P, et al. Decellularization protocols of porcine heart valves differ significantly in efficiency of cell removal and susceptibility of the matrix to recellularization with human vascular cells. J Thorac Cardiovasc Surg. 2004;127(2):399-405. DOI: 10.1016/j.jtcvs.2003.06.017

Hudson TW, Zawko S, Deister C, Lundy S, Hu CY, Lee K, et al. Optimized acellular nerve graft is immunologically tolerated and supports regeneration. Tissue Eng. 2004;10(11-12):1641-51. DOI: 10.1089/ten.2004.10.1641

Grauss RW, Hazekamp MG, van Vliet S, Gittenberger-de Groot AC, DeRuiter MC. Decellularization of rat aortic valve allografts reduces leaflet destruction and extracellular matrix remodeling. J Thorac Cardiovasc Surg. 2003 Dec;126(6):2003-10. DOI: 10.1016/s0022-5223(03)00956-5

Keane TJ, Swinehart IT, Badylak SF. Methods of tissue decellularization used for preparation of biologic scaffolds and in vivo relevance. Methods. 2015;84:25-34. DOI: 10.1016/j.ymeth.2015.03.005

Tran HLB, Dinh TTH, Nguyen MTN, To QM, Pham ATT. Prepa­ration and characterization of acellular porcine pericardium for cardiovascular surgery. Turk J Biol. 2016;40:1243-50. DOI: 10.3906/biy-1510-44

Oswal D, Korossis S, Mirsadraee S, Wilcox H, Watterson K, Fisher J, et al. Biomechanical characterization of decellularized and cross-liked bovine pericardium. J Heart Valve Dis. 2007;16:165-74.

Andrée B, Bela K, Horvath T, Lux M, Ramm R, Venturini L, et al. Successful re-endothelialization of a perfusable bio­logical vascularized matrix (BioVaM) for the generation of 3D artificial cardiac tissue. Basic Res Cardiol. 2014;109(6):441. DOI: 10.1007/s00395-014-0441-x

Crapo PM, Gilbert TW, Badylak SF. An overview of tissue and whole organ decellularization processes. Biomaterials. 2011;32(12):3233-43. DOI: 10.1016/j.biomaterials.2011.01.057

Schmitt A, Csiki R, Tron A, Saldamli B, Tübel J, Florian K, et al. Optimized protocol for whole organ decellularization. Eur J Med Res. 2017;22:31. DOI: 10.1186/s40001-017-0272-y

Hashemi SS, Jowkar S, Mahmoodi M,Rafati AR,Mehrabani D, Zarei M, et al. Biochemical methods in production of three-dimensional scaffolds from human skin: A window in aesthetic surgery. World J Plast Surg. 2018;7(2):204-11.

Elder BD, Eleswarapu SV, Athanasiou KA. Extraction techniques for the decellularization of tissue engineered articular cartilage constructs. Biomaterials. 2009;30(22):3749-56. DOI: 10.1016/j.biomaterials.2009.03.050

Xing S, Liu C, Xu B, Chen J, Yin D, Zhang C. Effects of various decellularization methods on histological and biomechanical properties of rabbit tendons. Exp Ther Med. 2014;8(2):628-34. DOI: 10.3892/etm.2014.1742

Caralt M, Uzarski JS, Iacob S, Obergfell KP, Berg N, Bijonowski BM, et al. Optimization and critical evaluation of decellularization strategies to develop renal extracellular matrix scaffolds as biological templates for organ engineering and transplantation. Am J Transplant. 2015;15(1):64-75. DOI: 10.1111/ajt.12999

Manalastas TM, Dugos N, Ramos G, Mondragon JM. Effect of decellularization parameters on the efficient production of kidney bioscaffolds. Appl Biochem Biotechnol. 2020;1:1. DOI: 10.1007/s12010-020-03338-2

He M, Callanan A., Lagaras K, Steele JAM, Stevens MM. Optimization of SDS exposure on preservation of ECM characteristics in whole organ decellularization of rat kidneys. J Biomed Mater Res B Appl Biomater. 2017;105(6):1352-60. DOI: 10.1002/jbm.b.33668

GOST Style Citations

Copyright (c) 2020 The Author(s)

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.