Efficient decellularization of equine tendon with preserved biomechanical properties and cytocompatibility for human tendon surgery indications

Abstract

Chronic and acute tendon injuries are frequent afflictions, for which treatment is often long and unsatisfactory. When facing extended injuries, matrices and scaffolds with sufficient biomechanical properties are required for surgical repair and could additionally serve as supports for cellular therapies to improve healing. In this study, protocols of either commonly used detergents only (SDS 1%, Triton 1%, TBP 1%, and Tween‐20 1%) or a combination of freeze/thaw (F/T) cycles with decellularization agents (NaCl 1M, ddH2O) were evaluated for the decellularization of horse equine superficial digital flexor tendon (SDFT) for hand flexor or extensor tendon reconstruction. Decellularization efficiency was assessed microscopically by histological staining (HE, DAPI) and DNA quantification. Macroscopical structure and biomechanical integrity of the tendon matrices were further assessed by gross observation, histological staining (SR), and mechanical testing (ultimate strain and stress, Young’s modulus, energy to failure) for select protocols. Decellularization with hypertonic NaCl 1M in association with F/T cycles produced the most robust tendon matrices, which were nontoxic after 10 days for subsequent recellularization with human fetal progenitor tendon cells (hFPTs). This standardized protocol uses a less aggressive decellularization agent than current practice, which allows subsequent reseeding with allogenic cells, therefore making them very suitable and bioengineered tendon matrices for human tendon reconstruction in the clinic.

Sterilization and disinfection methods for decellularized matrix materials: Review, consideration and proposal

“…Applications : Alcohol is commonly used in disinfection of skin and precision instruments, etc. There are many reports on the use of alcohol to disinfect dECM, which are mainly derived from pericardium [ 11 ], valve [ 123 , 124 ], blood vessel [ 12 , 121 , 125 , 126 ], liver [ [127] , [128] , [129] ], pancreas [ 130 ], trachea [ 131 ], bladder [ 130 , [132] , [133] , [134] , [135] ], kidney [ 48 , 136 ], bone [ 137 , 138 ], tendon [ [139] , [140] , [141] ] and nerve [ 142 ].…”

“…Tendons mainly come from bovine, horse and pig, and a small number of tendons are from human, rabbit, sheep, dog. The sterilization and disinfection methods of decellularized tendon mainly include ethanol [ [139] , [140] , [141] ], EO [ [78] , [79] , [80] ], PAA [ 95 , 96 ], irradiation [ 27 , [54] , [55] , [56] ], antibiotics [ 153 , 179 ] and ScCO 2 [ 55 ], etc.…”

Holistic Approach of Swiss Fetal Progenitor Cell Banking: Optimizing Safe and Sustainable Substrates for Regenerative Medicine and Biotechnology

“…The efficacy of tendon transfer is hindered by accelerated graft degeneration and would largely benefit from therapeutic cell stimulation, ideally leading to optimal elasticity, mobility, and tensile strength restoration (O'Brien, 1997;Kannus, 2000). Bioengineering scaffolds of interest, such as human cadaveric and equine decellularized tendons or artificial equivalents, enable optimal maintenance of biocompatibility, mechanical properties, and susceptibility for cell seeding, whereas autologous vestigial tendons remain as the standard of care (Wehbé, 1992;Chong et al, 2009;Jakubietz et al, 2011;Pridgen et al, 2011;Burk et al, 2016;Lovati et al, 2016;Valentin et al, 2016;Aeberhard et al, 2019). Vast arrays of potential therapeutic cell types have been investigated in tendon bioengineering for regeneration enhancement, including tendon sheath fibroblasts, adult tenocytes, stem cells, placenta cells, amniotic cells, and platelet-derivatives (Kadner et al, 2002;Kaviani et al, 2002Kaviani et al, , 2003Awad et al, 2003;Chen et al, 2009;Akhundov et al, 2012;Xu et al, 2013;Petrou et al, 2014).…”

A new decellularized tendon scaffold for rotator cuff tears – evaluation in rabbits

“…One approach to eliminating the risk of disease transmission or immune reactions and improving the availability of scaffolds is the decellularization process, which transforms a scaffold into a shelf product [ 14 – 16 ]. Decellularized tendon scaffolds (DTSs) are prepared from different protocols that include a combination of physical, chemical and enzymatic techniques [ 17 , 18 ].…”

“…Decellularized tendon scaffolds (DTSs) are prepared from different protocols that include a combination of physical, chemical and enzymatic techniques [ 17 , 18 ]. Recent studies have shown reproducible results in biomechanical and histological evaluations for protocols that include combinations of chemical and enzymatic decellularization agents [ 16 , 19 , 20 ].…”