title: A liquidus tracking approach to the cryopreservation of human cartilage allografts.
abstract: In the "liquidus tracking" (LT) approach to cryopreservation both the temperature and the concentration of cryoprotectant (CPA) are controlled such that solution composition "tracks" the liquidus (melting point) line for that system. Ice crystal formation is prevented but the tissue is not exposed to CPA concentrations exceeding those experienced by cells during conventional cryopreservation. This approach is particularly appropriate for articular cartilage because chondrocytes in situ are exquisitely susceptible to damage by the crystallisation of ice. This project aimed to develop a suitable process for tissue to be used in the surgical repair of damaged human knee joints. A high proportion of the chondrocytes should be alive. Human articular cartilage was obtained from deceased donors and dimethyl sulphoxide (DMSO) was used as the CPA, cooling was at 0.14°C/min and warming at 0.42°C/min. The vehicle solution was CPTes2. A program of increasing DMSO concentration was developed for cooling and this gave satisfactory tissue concentrations but reduction of DMSO concentration during warming was inadequate, resulting in higher tissue concentrations than required. Biomechanical testing indicated a compressive modulus of 9.5±1.3 MPa in LT-processed cartilage, with control values of 11.6±0.8 MPa (p>0.05, Student's t-test). Measurement of GAG synthesis sometimes approached 65% or 85% of control, but the variability of replicate data prevented firm conclusions. Ideally allograft tissue should score 1A or above on the Noyes scale and the donor age should be less than 46 years but the cartilage used in this study did not meet these standards.
authors.affiliation: R&D Tissue Services, NHS Blood and Transplant, 14 Estuary Banks, Speke, Liverpool L24 8RB, UK; Leopold Muller Arthritis Research Centre, Robert Jones and Agnes Hunt Orthopaedic Hospital Foundation Trust, Oswestry, Shropshire SY10 7AG, UK; Biology Department, University of York, York YO10 5DD, UK. Electronic address: A.G.Kay@keele.ac.uk.
authors.forename: A G
authors.initials: AG
authors.lastname: Kay
authors.affiliation: Centre For Tissue Injury and Repair, Institute of Inflammation and Repair, Faculty of Medical and Human Sciences, The University of Manchester, Stopford Building, Oxford Road, Manchester M13 9PT, UK.
authors.forename: J A
authors.initials: JA
authors.lastname: Hoyland
authors.affiliation: R&D Tissue Services, NHS Blood and Transplant, 14 Estuary Banks, Speke, Liverpool L24 8RB, UK.
authors.forename: P
authors.initials: P
authors.lastname: Rooney
authors.affiliation: R&D Tissue Services, NHS Blood and Transplant, 14 Estuary Banks, Speke, Liverpool L24 8RB, UK.
authors.forename: J N
authors.initials: JN
authors.lastname: Kearney
authors.affiliation: Biology Department, University of York, York YO10 5DD, UK.
authors.forename: D E
authors.initials: DE
authors.lastname: Pegg
mesh_terms: D000328:Adult; D000368:Aged; D064591:Allografts / Q000601:surgery*; D002358:Cartilage, Articular / Q000601:surgery*; D019902:Chondrocytes / Q000502:physiology; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D003460:Crystallization; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D005260:Female; D006801:Humans; D007718:Knee Injuries / Q000601:surgery; D007719:Knee Joint / Q000601:surgery*; D008297:Male; D008875:Middle Aged; D013696:Temperature
chemical_list: D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide
keywords: Articular cartilage; Chondrocyte; Cryobiology; Cryopreservation; Liquidus tracking; Vitrification