Breakthrough Tissue and Organ Preservation and Transplantation Using Scaled-Up Nanowarming Technology

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ABSTRACT: Rewarming biomaterials from the vitrified state is a critical step in obtaining successful cryopreservation. Successful techniques for rescuing cryopreserved bulk biomaterials and organs would not only provide critical improvements for donor-organ transport, supply, and matching, but is also a missing link in the potential supply chain for engineered tissues. Typical freezing processes cause significant damage to biomaterials through ice crystal formation and cellular dehydration. However, with the aid of cryoprotectant (CPA) solutions, biospecimens can be stabilized in the vitreous (i.e. “glass” or “amorphous”) state, allowing for long-term cryopreservation. A number of groups have employed successful techniques for cooling bulk systems to the vitreous state (including entire rabbit kidneys). Rewarming these vitrified biomaterials is a greater engineering challenge, due to the critical warming rates (hundreds of oC/min) necessary to avoid devitirification (i.e. crystallization) during thaw. In addition, non-uniformity in temperature field produces thermal stresses that can crack the brittle material, and so both speed and uniformity of thaw are of critical importance. Here we propose to investigate the ability of radiofrequency heated magnetic nanoparticles, or “nanowarming,” to overcome this major limitation hindering further development of bulk cryopreservation approaches. Although electromagnetic rewarming has been tried, the direct coupling of the waves to tissue inherently results in non-uniformity in heating, which leads to cracking and differential viability. At lower radiofrequencies (RF < 1 MHz) alternating magnetic fields (AMFs) can uniformly penetrate tissues without attenuation and negligible dielectric coupling. Although these lower frequency fields will be unable to rapidly heat the tissue on their own, they are ability to produce significant heating through coupling with magnetic (e.g. iron-oxide) nanoparticles. We have already demonstrated that this approach is able to generate heating rates rapid enough to avoid devitirification (greater than 200 oC/min) and should scale independent of sample size. The objective of this study is to refine this novel nanowarming technology for use in cryopreserving biologic tissues and intact organs for transplant. To this end, in Aim 1 we will scale up the nanoparticle production process and the size of the RF heating device. In Aim 2 we will optimize CPA and nanoparticle composition and loading/unloading conditions for vitrification and nanowarming of cells and tissues (arteries). In Aim 3 we will test these optimized conditions in heart transplant models of increasing size and complexity. In summary, the focus of this proposal will be to leverage our breakthrough nanowarming technology by optimizing CPA composition and nanoparticle delivery in a scaled up system capable of vitrifying and recovering cells, arteries, and intact organs with an eye on future application for cryopreserving tissues and organs for use in human transplantation.