An Integrated Approach to Optimize Vitrification of Articular Cartilage

• Vitrification of Articular Cartilage

• Author / Creator

  Wu, Kezhou

• Articular cartilage (AC) is a few-millimetres-thick hyaline cartilage tissue covering the bone end in articulating joints. AC is composed primarily of cells called “chondrocytes” and abundant surrounding cartilage matrix components. AC is a vital structure in daily joint mobility function. Thus, it is important to maintain an intact AC structure. Due to the avascular and aneural anatomical nature, AC has a poor self-repair capacity after injuries. AC defects over 1 cm2 often progress into osteoarthritis if not properly treated. While osteochondral graft transplantation is an effective method of AC defect repair, it is limited by several factors such as donor availability and perioperative preparations, which make AC defect repair a challenging clinical issue. Cryopreservation of chondrocytes and AC for long-term storage is a promising solution that can provide clinical surgeons with a long and flexible timeframe for performing cartilage transplantation. However, this requires a well-designed tissue banking system with appropriate cryopreservation protocols to store donated AC tissues.

  Cryoprotectants (CPAs) are capable of reducing ice formation within cells and tissues during the cooling–warming procedures, which plays an essential role in AC cryopreservation. However, they can exhibit undesirable effects such as osmotic or oxidative stress on chondrocytes which leads to cell dysfunction and damage. Additionally, the long CPA permeation process required for AC cryopreservation might aggravate the severity of CPA toxicity on the chondrocytes. It remains challenging to eliminate all adverse effects induced by the CPA compounds and the CPA permeation procedures. In the past decades, several promising cryopreservation approaches emerged as tools to guide the design of cryopreservation protocols for preserving AC at cryogenic temperatures. The proposed approaches include slow graded freezing (stepwise cooling), multiple steps of cryoprotectant addition and removal, modelling of cryoprotectant transport, etc. With the emergence of vitrification, an ‘ice-free’ method to preserve tissue in a “glassy” solid that avoids ice crystal formation at −196 °C, more approaches have been developed including liquidus tracking of cryoprotectants, minimization of cryoprotectant toxicity by using cryoprotectant mixtures, mathematical calculation of cryoprotectant permeation and vitrifiability, etc. All of these approaches have provided information to promote the advancement of AC vitrification. Furthermore, this knowledge has been applied to intact human AC.

  In this thesis, different strategies have been investigated to improve AC cryopreservation protocols from several perspectives: 1) to validate the efficiency of vitrification protocols generated via engineering optimization of CPA concentration, permeation temperature, duration of CPA permeation; 2) to improve heat transfer of AC during cooling and warming processes by the optimization of container size and packaging method; 3) to mitigate the toxicity of CPAs by inclusion of additives in the CPA mixtures composed of multiple types of CPAs.

  The research presented in this thesis focuses on the investigation of a combination of different strategies within the continuously evolving cryopreservation protocols for porcine and human AC, and the evaluation of their protective as well as damaging mechanisms on the chondrocytes. The findings from these studies provide us better understanding and new knowledge to establish successful cryopreservation protocols for intact human AC.

• Subjects / Keywords

  • Vitrification
  • Articular Cartilage
  • Approach

• Graduation date

  Fall 2020

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-c2v0-zd32

• License

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