Partial freezing of rat livers extends preservation time by 5-fold

Tessier, Shannon N.; Vries, Reinier J. de; Pendexter, Casie A.; Cronin, Stephanie E.J.; Özer, Sinan; Hafiz, Ehab; Raigani, Siavash; Oliveira-Costa, João Paulo; Wilks, Benjamin T.; Higuita, Manuela Lopera; Gulik, Thomas M. van; Usta, O. Berk; Stott, Shannon L.; Yeh, Heidi; Yarmush, Martin L.; Uygun, Korkut; Toner, Mehmet

doi:10.1038/s41467-022-31490-2

Cited by 33 publications

(33 citation statements)

References 67 publications

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“…2,10,17,18 Partial freezing is a preservation method inspired by freeze-tolerant animals in nature and aims to achieve significant metabolic suppression by coupling high subzero storage temperatures (−10°C) with cellular dehydration, making it particularly difficult to test in classic in vitro models. 10 Therefore, by taking advantage of the versatility of zebrafish as a screening tool, both adult and larvae models were utilized for screening and viability assessments after partial freezing. In tandem, fifty combinations (CPAs cocktails) of twelve different CPAs were tested for their ability to preserve heart viability and functionality after extended partial freezing in a scalable, high-throughput format with the aim of identifying potential CPAs cocktail candidates for the use in long-term storage of cardiac allografts.…”

Section: Introductionmentioning

confidence: 99%

“…The purpose of this study is, therefore, to innovate in solid organ preservation research by investigating the usability of zebrafish as a high‐throughput model organism, which to our knowledge, has not been done before. As the case study, this work focuses on heart preservation via partial freezing due to (i) the urgency to improve heart preservation protocols, (ii) the ability of zebrafish heart to recapitulate human cardiac function, and (iii) the promising results seen previously in partial freezing of rodent and human liver, likely to translate into heart preservation 2,10,17,18 . Partial freezing is a preservation method inspired by freeze‐tolerant animals in nature and aims to achieve significant metabolic suppression by coupling high subzero storage temperatures (−10°C) with cellular dehydration, making it particularly difficult to test in classic in vitro models 10 .…”

Section: Introductionmentioning

confidence: 99%

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Zebrafish as a high throughput model for organ preservation and transplantation research

Da Silveira Cavalcante,

Lopera Higuita,

González‐Rosa

et al. 2023

The FASEB Journal

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Despite decades of effort, the preservation of complex organs for transplantation remains a significant barrier that exacerbates the organ shortage crisis. Progress in organ preservation research is significantly hindered by suboptimal research tools that force investigators to sacrifice translatability over throughput. For instance, simple model systems, such as single cell monolayers or co‐cultures, lack native tissue structure and functional assessment, while mammalian whole organs are complex systems with confounding variables not compatible with high‐throughput experimentation. In response, diverse fields and industries have bridged this experimental gap through the development of rich and robust resources for the use of zebrafish as a model organism. Through this study, we aim to demonstrate the value zebrafish pose for the fields of solid organ preservation and transplantation, especially with respect to experimental transplantation efforts. A wide array of methods were customized and validated for preservation‐specific experimentation utilizing zebrafish, including the development of assays at multiple developmental stages (larvae and adult), methods for loading and unloading preservation agents, and the development of viability scores to quantify functional outcomes. Using this platform, the largest and most comprehensive screen of cryoprotectant agents (CPAs) was performed to determine their toxicity and efficiency at preserving complex organ systems using a high subzero approach called partial freezing (i.e., storage in the frozen state at −10°C). As a result, adult zebrafish cardiac function was successfully preserved after 5 days of partial freezing storage. In combination, the methods and techniques developed have the potential to drive and accelerate research in the fields of solid organ preservation and transplantation.

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Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Zebrafish as a high throughput model for organ preservation and transplantation research

Da Silveira Cavalcante,

Lopera Higuita,

González‐Rosa

et al. 2023

The FASEB Journal

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“…At such temperatures, biological activity is reduced and both structure and function can be protected for several days. Whilst such a short timeframe is limiting, this can be sufficient to overcome extreme time constraints associated with, for example, transport, quality checks and organ transplants [11,62,[66][67][68].…”

Section: Ice Nucleation and Direct Ice Damagementioning

confidence: 99%

Scaling up Cryopreservation from Cell Suspensions to Tissues: Challenges and Successes

Kilbride¹,

Meneghel²,

Chawda³

et al. 2023

Biomedical Engineering

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This chapter covers the key physical, biological and practical challenges encountered when developing cryopreservation protocols for larger biological structures and examines areas where cryopreservation has been successful in scaling to larger structures. Results from techniques being used in attempts to overcome these challenges are reviewed together with the indicators for future development that arise from them. The scale-up of cryopreservation to tissues with diverse functions and cell types makes the control of freezing and thawing more challenging. Technology may—or may not—be available depending on the size of the material involved. To meet the challenge there must be innovation in technology, techniques and understanding of damage-limiting strategies. Diversity of cell structure, size, shape and expected function means a similarly diverse response to any imposed cryopreservation conditions and interaction with ice crystals. The increasing diffusion distances involved, and diversity of permeability properties, will affect solutes, solvents, heat and cryoprotectant (CPA) transfer and so add to the diversity of response. Constructing a single protocol for cryopreservation of a larger sample (organoids to whole organs) becomes a formidable challenge.

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“…A variety of techniques for supercooling preservation without ice were developed throughout the years. To list a few: emulsification (2), elevated pressure (3), electromagnetic fields (4), cryoprotectant solutions at temperatures at which they do not freeze (5) (6), antifreeze proteins (7), partial freezing to mimic survival of freeze tolerant species (8) which recently reported major achievements (9), deep-supercooling of large volumes with surface sealing with immiscible fluids (10) which also reported major achievements (11).…”

Section: Introductionmentioning

confidence: 99%

A two-compartment system for subfreezing temperatures preservation of large volumes of organic matter in an isochoric system

Năstase

Botea

Beșchea

et al. 2022

Preprint

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This is a technology paper on the design of and test results from an 11-liter isochoric (constant volume) chamber, for the preservation of large, organs in a supercooled state. Details of the design are given, as well as a proof that the chamber is isochoric. Five repeats show that in this large chamber, ice nucleation of steam distilled water occurs at -2 degree Celsius within less than 12 hours, in all the repeats. An examination of the experimental results suggests that the ice nucleation starts on the inner walls of the isochoric chamber. A new two compartment isochoric chamber was designed to reduces the probability of ice nucleation on the walls of the chamber. In the two-compartment system, the biological matter and the preservation fluid are introduced in a sealed low-density polyethylene bag, and placed in the center of the isochoric chamber, in such a way that the bag does not touch the walls. The space between the inner walls of the isochoric chamber and the outer walls of the bag are filled with a fluid with a composition that does not freeze at the storage temperature. Three repeat experiments with steam distilled water and with in vitro pig liver show that with this technique, the system remained supercooled, without any ice nucleation for the duration of the experiments. Experiments were voluntary terminated at 48 hours of supercooling. This new technology may hold promise for long term preservation of large biological organs in a supercooled state, without the use of any chemical additives.

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Partial freezing of rat livers extends preservation time by 5-fold

Cited by 33 publications

References 67 publications

Zebrafish as a high throughput model for organ preservation and transplantation research

Zebrafish as a high throughput model for organ preservation and transplantation research

Scaling up Cryopreservation from Cell Suspensions to Tissues: Challenges and Successes

A two-compartment system for subfreezing temperatures preservation of large volumes of organic matter in an isochoric system

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