Cryopreservation with glycerol improves the in vitro biomechanical characteristics of human patellar tendon allografts
To evaluate the in vitro biomechanical characteristics of patellar tendon ligaments (BTB) when stored as fresh frozen or as glycerol cryopreserved allografts.
Seventy patellar tendons were harvested from 35 cadaveric human donors and randomly assigned into seven groups. Grafts in group FRESH were mechanically tested within 2 h of harvesting. FROZ-3, FROZ-6, and FROZ-9 were deep-frozen to −80°C for 3, 6, and 9 months, respectively. Grafts in groups CRYO-3, CRYO-6, and CRYO-9 were initially incubated with 10 % glycerol in a phosphate-buffered saline for 1 h and then stored in glycerol solution (10 % glycerol in PBS) at −80°C for 3, 6, and 9 months, respectively. Grafts were mechanically tested with two cycling modes (50–250°N and 150–500°N) and then loaded to failure.
Cryopreserved grafts demonstrated more consistent results and expressed lower elongation rates after both cycling loading protocols compared to their frozen counterparts at all storage times. During load-to-failure analysis, ultimate stiffness levels were predominantly higher (23.9–61.5 %) in cryopreserved grafts compared with frozen grafts, and ultimate stress levels were 26 % (13.3–47.7 %) higher, regardless of the storage time. Moreover, cryopreserved grafts revealed similar ultimate elongation and uniformly higher ultimate stiffness and ultimate stress levels compared to fresh grafts.
The results of this in vitro study demonstrated superior mechanical properties of cryopreserved grafts compared to frozen grafts within a preservation period of 9 months. Cryopreservation with glycerol solution might be used to further improve the quality of preserved soft-tissue allografts.
KeywordsAnterior cruciate ligament Allograft Cryopreservation Glycerolisation Human bone-patellar tendon-bone graft Mechanical testing
- 2.American Association of Tissue Banks: Standards for Tissue Banking—12th edn. McLean: American Association of Tissue Banks, 2008. Available at: http://www.aatb.org
- 14.Hoburg A, Keshlaf S, Schmidt T, Smith M, Gohs U, Perka C, Pruss A, Scheffler S (2011) Fractionation of high-dose electron beam irradiation of BPTB grafts provides significantly improved viscoelastic and structural properties compared to standard gamma irradiation. Knee Surg Sports Traumatol Arthrosc 19:1955–1961PubMedCrossRefGoogle Scholar
- 15.Honl M, Carrero V, Hille E, Schneider E, Morlock MM (2002) Bone-patellar tendon-bone grafts for anterior cruciate ligament reconstruction: an in vitro comparison of mechanical behavior under failure tensile loading and cyclic submaximal tensile loading. Am J Sports Med 30:549–557PubMedGoogle Scholar
- 18.Kamiński A, Gut G, Marowska J, Lada-Kozłowska M, Biwejnis W, Zasacka M (2008) Mechanical properties of radiation-sterilised human bone-tendon-bone grafts preserved by different methods. Cell Tissue Bank 45:122–129Google Scholar
- 27.Paulos LE, Karistinos A, Walker J (2006) “Criteria”-based rehabilitation of surgically reconstructed and nonsurgically treated anterior cruciate ligament injuries. In: Scott WN (ed) Insall & scott surgery of the knee, 4th edn. Elsevier Churchill Livingstone, Philadelphia, pp 693–714Google Scholar
- 29.Rodrigo JJ, Jackson DW, Simon TM (1993) The immune response to freeze dried bone tendon bone allografts in humans. Am J Knee Surg 6:347–353Google Scholar
- 32.Scheffler SU, Sudkamp NP, Gockenjan A, Hoffmann RF, Weiler A (2002) Biomechanical comparison of hamstring and patellar tendon graft anterior cruciate ligament reconstruction techniques: the impact of fixation level and fixation method under cyclic loading. Arthroscopy 18:304–315PubMedCrossRefGoogle Scholar
- 37.Wingenfeld C, Egli RJ, Hempfing A, Ganz R, Leunig M (2002) Cryopreservation of osteochondral allografts: dimethyl sulfoxide promotes angiogenesis and immune tolerance in mice. J Bone Joint Surg Am 84-A:1420–1429 (Erratum in: J Bone Joint Surg Am 84-A:1855)Google Scholar