A method of isolating viable chondrocytes with proliferative capacity from cryopreserved human articular cartilage
- PDF / 631,678 Bytes
- 10 Pages / 547.087 x 737.008 pts Page_size
- 43 Downloads / 165 Views
ORIGINAL PAPER
A method of isolating viable chondrocytes with proliferative capacity from cryopreserved human articular cartilage Zhidao Xia • Xin Duan • David Murray James T. Triffitt • Andrew J. Price
•
Received: 18 May 2012 / Accepted: 6 July 2012 / Published online: 17 July 2012 Ó Springer Science+Business Media B.V. 2012
Abstract This study aimed to optimise methods of cryopreserving human articular cartilage (AC) tissue for the isolation of late chondrocytes. Human AC specimens from osteoarthritis patients who had undergone total knee replacement were used to optimise the chondrocyte isolation process and the choice of cryoprotective agent (CPA). For AC tissue cryopreservation, intact cored cartilage discs (5 mm diameter) and diced cartilage (0.2–1 mm cubes) from the same sized discs were step cooled and stored in liquid nitrogen for up to 48 h before chondrocyte isolation and in vitro assay of cell viability and proliferative potential. The results showed that 10 % dimethyl sulphoxide in 90 % foetal bovine serum was a successful CPA for chondrocyte cryopreservation. Compared with intact cored discs, dicing of AC tissue into 0.2–1 mm cubes significantly increased the viability and proliferative capacity of surviving
chondrocytes after cryopreservation. In situ crosssection imaging using focused ion beam microscopy revealed that dicing of cored AC discs into small cubes reduced the cryo-damage to cartilage tissue matrix. In conclusion, modification of appropriate factors, such as the size of the tissue, cryoprotective agent, and isolation protocol, can allow successful isolation of viable chondrocytes with high proliferative capacity from cryopreserved human articular cartilage tissue. Further studies are required to determine whether these cells may retain cartilage differentiation capacity and provide sufficient chondrocytes for use as implants in clinical applications. Keywords Cryopreservation Human articular cartilage Cryoprotective agent Focused ion beam (FIB) microscopy Viability Proliferation
Introduction Z. Xia D. Murray J. T. Triffitt A. J. Price Nuffield Department of Orthopaedic Surgery, Botnar Research Centre, Oxford University Institute of Musculoskeletal Sciences, University of Oxford, Oxford OX3 7LD, UK Z. Xia (&) X. Duan Institute of Life Science, College of Medicine, Swansea University, Singleton Park, Swansea SA2 8PP, UK e-mail: [email protected] X. Duan Orthopaedics Department, The Second People’s Hospital of Chengdu, Wucheng Road, Chengdu 610017, China
Articular cartilage damage through injury or disease results in significant disability for a large number of individuals (Jomha et al. 2002). Surgical techniques such as arthroscopic debridement (Johnson 2001), microfracture techniques (Steadman et al. 2001), and use of stem cell implants (Magne et al. 2005), fresh/ frozen articular cartilage allografts (Gross et al. 2001; Clatworthy et al. 2001), fresh autografts (Whiteside et al. 2005; Andres et al. 2003) and autologous chondrocyte implantation (ACI) with or witho
Data Loading...
