Monomeric glutaraldehyde fixation and amino acid detoxification of decellularized bovine pericardium for production of biocompatible tissue with tissue-guided regenerative potential

Angélique Lewies; Lezelle Botes; Johannes Jacobus van den Heever; Pascal Maria Dohmen; Francis Edwin Smit

doi:10.1016/j.heliyon.2023.e19712

Monomeric glutaraldehyde fixation and amino acid detoxification of decellularized bovine pericardium for production of biocompatible tissue with tissue-guided regenerative potential

Heliyon. 2023 Sep 1;9(9):e19712. doi: 10.1016/j.heliyon.2023.e19712. eCollection 2023 Sep.

Authors

Angélique Lewies¹, Lezelle Botes², Johannes Jacobus van den Heever¹, Pascal Maria Dohmen^{1

3}, Francis Edwin Smit¹

Affiliations

¹ Department of Cardiothoracic Surgery, Faculty of Health Sciences, University of the Free State, Bloemfontein, South Africa.
² Department of Health Sciences, Central University of Technology, Free State, Bloemfontein, South Africa.
³ Department of Cardiac Surgery, Heart Centre Rostock, University of Rostock, Germany.

Abstract

The effect of monomeric glutaraldehyde fixation and amino acid detoxification on biocompatibility and tissue-guided regenerative potential of decellularized bovine pericardium was evaluated. The degree of cross-linking, porosity, enzymatic degradation, alpha-galactosyl content, the efficacy of detoxification, and cytotoxicity towards human epithelial cells were assessed. Tissue was subcutaneously implanted for eight weeks in male juvenile Sprague-Dawley rats, and mechanical properties, host cell infiltration, and calcification were evaluated. Three groups were compared i) decellularized tissue, ii) decellularized, monomeric glutaraldehyde fixed and amino acid detoxified tissue, and iii) commercial glutaraldehyde fixed non-decellularized tissue (Glycar®) (n = 6 rats per group). The fixation process gave a high degree of cross-linking (>85%), and was resistant to enzymatic degradation, with no significant effect on porosity. The detoxification process was effective, and the tissue was not toxic to mammalian cells in vitro. Tissue from both decellularized groups had significantly higher (p < 0.05) porosity and host cell infiltration in vivo. The process mitigated calcification. A non-significant decrease in the alpha-galactosyl content was observed, which increased when including the alpha-galactosidase enzyme. Mechanical properties were maintained. The fixation and detoxification process adequately removes free aldehyde groups and reduces toxicity, preventing enzymatic degradation and allowing for host cell infiltration while mitigating calcification and retaining the mechanical properties of the tissue. This process can be considered for processing decellularized bovine pericardium with tissue-guided regeneration potential for use in cardiovascular bioprostheses; however, methods of further reducing antigenicity, such as the use of enzymes, should be investigated.

Keywords: Amino acids; Bioprosthetic valves; Bovine pericardium; Calcification; Decellularization; Detoxification; Glutaraldehyde fixation; Host cell infiltration.