Impact of Hypothermia upon Chondrocyte Viability and Cartilage Matrix Permeability after 1 Month of Refrigerated Storage

Abstract

Background: The purpose of this research was to assess the extracellular matrix and chondrocytes of articular cartilage during refrigerated storage and to determine whether changes could be detected in the time frame that cartilage is stored for clinical use. Methods: Porcine cartilage was stored as either bisected femoral heads with bone attached or plugs without the underlying bone in culture medium with fetal bovine serum for 1 month at 4 °C. Metabolic activity was tested using a resazurin reduction method on intact tissue and viable cell recovery after enzymatic tissue digestion at each time point. Cartilage plug permeability was evaluated by measuring electrical conductivity. Results: Storage in culture medium provided good cartilage viability and metabolic function for 7 days; however, significant changes were observed in femoral heads (p < 0.05). All mean chondrocyte assessment values were <30% of fresh controls at 28 days. Cartilage plugs tended to perform better after 7 days of storage than the femoral heads and retained significantly higher metabolic activity (mean = 94.5% vs. 70.5%; p < 0.05). Cartilage plugs demonstrated consistent changes in electrical conductivity after 28 days of storage (p < 0.05). Conclusion: Refrigerated storage of cartilage results in both loss of chondrocyte viability and matrix permeability.

Impact of Storage Solution Formulation during Refrigerated Storage upon Chondrocyte Viability and Cartilage Matrix

“…Then conductivity was tested, first in isotonic PBS and then after swelling in hypotonic saline (0.2xPBS). The electrical conductivity was measured based on the principle of a four wire resistance test using a Keithley Sourcemeter (Model 2400, Keithley Instruments, Inc., Cleveland, OH) and a custom designed conductivity chamber reported previously [Gu et al, 2002; Brockbank et al, 2011b]. Briefly, the conductivity apparatus consists of two stainless steel current electrodes coaxial to two Teflon-coated Ag/AgCl voltage electrodes placed on the top and bottom of a cylindrical nonconductive Plexiglass chamber (5mm diameter).…”

“…There are, however, reports supporting chondrocyte survival for days or weeks of hypothermic storage in their natural extracellular matrix in humans [Williams et al, 2003; Ball et al, 2004; Allen et al, 2005; Malinin et al, 2006] and several animal species [Black et al, 1979; Rodrigo et al, 1980; Wayne et al, 1990; Oates et al, 1995; Kim et al, 1996; Rohde et al, 2004; Williams et al, 2004; Malinin et al, 2006; Teng et al, 2008; Onuma et al, 2009]. Because such survival was considered unusual, compared with other tissue types, we previously initiated research to assess the impact of 4°C storage in DMEM culture medium on cartilage cell viability and establish whether or not cartilage plugs were an acceptable model compared with bisected femoral heads with both cartilage and bone tissue present [Brockbank et al, 2011b]. We also included extracellular matrix (ECM) permeability evaluation during storage because prolonged storage of cartilage with viable cells might promote the release of enzymes that impact tissue material properties [Brockbank et al, 2011b].…”

“…Because such survival was considered unusual, compared with other tissue types, we previously initiated research to assess the impact of 4°C storage in DMEM culture medium on cartilage cell viability and establish whether or not cartilage plugs were an acceptable model compared with bisected femoral heads with both cartilage and bone tissue present [Brockbank et al, 2011b]. We also included extracellular matrix (ECM) permeability evaluation during storage because prolonged storage of cartilage with viable cells might promote the release of enzymes that impact tissue material properties [Brockbank et al, 2011b]. We found that storage in DMEM culture medium with 10% FBS provides good cartilage viability for 7 days.…”

“…In addition, there was a marked tendency for cartilage plugs to demonstrate higher viability values than the femoral heads. However, 1 month storage of cartilage resulted in loss of both chondrocyte viability and ECM permeability [Brockbank et al, 2011b]. …”

Tissue Banking and Transfusion Medicine – a Suitable Cooperation Model

“…Brockbank et al [7] concern themselves with the question of the consequences of long-term storage of cartilage tissue at 4 °C and provided evidence of negative effects on chondrocyte viability and matrix permeability. George Galea [8] reports on the main reasons for the exclusion of femoral head donors in Scotland and discusses the results by comparing them with the regional blood donor collective.…”