Ice-free vitrification and nanowarming of meniscal grafts for transplantation

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ABSTRACT: Limited availability of fresh viable donor meniscal grafts reduces size-matching efficiency for meniscal allograft transplantation, restricting surgical accessibility and patient outcomes. Current preservation methods with retention of meniscal viability are inadequate. Fibrochondrocyte viability of transplanted menisci is accepted as one of the determinants of outcome following allograft transplantation. We have previously developed an ice-free vitrification method of cryopreservation that maintains excellent chondrocyte viability in animal models and human articular cartilage. We present preliminary data here demonstrating that fibrochondrocytes in menisci can also survive vitrification due to the absence of ice formation during cooling and warming of small 1-3mL samples. However, it has not been possible to rewarm larger samples due to ice nucleation during rewarming that results in loss of cell viability. The innovation in this proposal relates to a new rewarming method that does not have the limitations of boundary convection warming that should be effective for samples up to 50mL in volume. This rewarming method utilizes radio frequency induced heating of magnetic iron nanoparticles. We will optimize nanowarming of full thickness meniscal fibrocartilage plugs for maintenance of cell viability, including gene expression, and extracellular matrix integrity. This objective will be developed in two specific aims to optimize nanowarming variables and scaleup from 5mL employing tissue plugs to 30mL with intact menisci. Menisci will be vitrified and stored for 1-4 weeks and then nanowarmed meniscal cell viability, chemistry, and biomaterial properties will be compared with untreated fresh control tissue. The nanowarming conditions that provides the best preservation of cells with minimal if any biomaterial changes will be selected for further investigation in vivo and translation to human menisci in a subsequent, future, Phase II SBIR application.