Assessment of the cryoprotectant concentration inside a bulky organ for cryopreservation using X-ray computed tomography

Corral, Ariadna; Balcerzyk, Marcin; Parrado-Gallego, Ángel; Fernández-Gómez, I.; Lamprea, David R.; Olmo, Alberto; Risco, Ramón

doi:10.1016/j.cryobiol.2015.09.007

Cited by 21 publications

(12 citation statements)

References 42 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…In general, it would be valuable to observe CPA tissue concentrations in real-time to monitor equilibration status, as this would make it possible to adjust both equilibration duration and also the initial CPA concentrations in the equilibration solution. In the case of Me 2 SO, such real-time observation is possible using CT, due to the high electron density of the sulfur atom included [19]. Herein, we could confirm that real-time CT-imaging is a useful tool to assess Me 2 SO tissue distribution.…”

Section: Discussionsupporting

confidence: 65%

“…Most of the perfusion-based protocols for organ cryopreservation that have been published share the common feature that the CPA equilibration durations used range from approximately 25 minutes up to a couple of hours. Passive equilibration using diffusion alone would increase the recommended equilibration duration even further: Corral et al [19] postulated that passive whole-organ equilibration of a rabbit kidney with Me 2 SO would take 7-9 days. None of the above studies carried out real-time monitoring of the equilibration process; they either estimated CPA tissue concentrations based on indirect reference values ( [10]), or evaluated the CPA concentration after assumed equilibration [15,19]leaving open the question of whether the Me 2 SO distribution actually reaches an adequate level much earlier.…”

Section: Discussionmentioning

confidence: 99%

“…Passive equilibration using diffusion alone would increase the recommended equilibration duration even further: Corral et al [19] postulated that passive whole-organ equilibration of a rabbit kidney with Me 2 SO would take 7-9 days. None of the above studies carried out real-time monitoring of the equilibration process; they either estimated CPA tissue concentrations based on indirect reference values ( [10]), or evaluated the CPA concentration after assumed equilibration [15,19]leaving open the question of whether the Me 2 SO distribution actually reaches an adequate level much earlier. The present study shows that all cells and extracellular spaces in rat heart tissue are 95% equilibrated after � 35 s perfusion with 15% (v/v) Me 2 SO; thus, Me 2 SO tissue distribution completes much more rapidly upon organ perfusion than previously expected.…”

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Me2SO perfusion time for whole-organ cryopreservation can be shortened: Results of micro-computed tomography monitoring during Me2SO perfusion of rat hearts

et al. 2020

View full text Add to dashboard Cite

Cryopreservation of whole organs and specific tissues is an important and continually expanding field of medicine. The protocols currently used for organ preservation do not ensure survivability and functionality; the protocols for ovarian tissue lead to acceptable outcomes, but these are still capable of further improvement. In general, cryopreservation protocols need to be optimized. One important approach to improving cryopreservation protocols in general involves reducing exposure to cytotoxic cryoprotective agents prior to freezing. This study, therefore, evaluated the real-time tissue penetration of dimethyl sulfoxide, a cryoprotective agent that is widely used in cryopreservation. Dimethyl sulfoxide penetration in rat hearts perfused with a 15% (v/v) dimethyl sulfoxide solution was examined in real-time using dynamic contrast-enhanced micro-computed tomography imaging. Viability of cardiomyocytes was not significantly affected by the dimethyl sulfoxide perfusion procedure. Two different perfusion rates were evaluated and compared with perfusion using a common iodine-based contrast agent (iomeprol). The dynamic contrast-enhanced microcomputed tomography imaging data showed that dimethyl sulfoxide flushes both the extracellular and intracellular spaces in rat heart tissue to 95% equilibration after � 35 s via perfusion. Subsequent wash-out via perfusion is completed to 95% within � 49 s. The equilibration duration routinely used in dimethyl sulfoxide-based protocols for cryopreservation should therefore be questioned. Shorter incubation duration would perhaps be sufficient, as well as being beneficial in relation to cell survivability. It would be helpful to have techniques for non-invasive real-time monitoring of the penetration of cryoprotective agents and such techniques should be used to revise cryopreservation protocols. Switching to perfusionbased equilibration procedures might be beneficial, if feasible.

show abstract

Section: Discussionsupporting

confidence: 65%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Me2SO perfusion time for whole-organ cryopreservation can be shortened: Results of micro-computed tomography monitoring during Me2SO perfusion of rat hearts

et al. 2020

View full text Add to dashboard Cite

show abstract

“…To determine CPA perfusion and removal efficiency, samples from three different patients were analyzed by computed tomography (CT) in order to ascertain DMSO concentrations in ovarian tissue after vitrification. A NanoCT scanner (Bioscan, USA; Mediso, Hungary) was used for this purpose, as previously reported 15,43 . The equipment was set up to maintain the samples at temperatures below −140 °C during vitrified tissue analysis.…”

Section: Methodsmentioning

confidence: 99%

Stepped vitrification technique for human ovarian tissue cryopreservation

Leonel

Corral

Risco

et al. 2019

Sci Rep

Self Cite

View full text Add to dashboard Cite

The advantage of stepped vitrification (SV) is avoiding ice crystal nucleation, while decreasing the toxic effects of high cryoprotectant concentrations. We aimed to test this method for human ovarian tissue cryopreservation. Ovarian cortex was taken from 7 fertile adult women. Samples were subjected to an SV protocol performed in an automatic freezer, which allowed sample transfer to ever higher concentrations of dimethyl sulfoxide (DMSO) as the temperature was reduced. Histological evaluation of the vitrified-warmed tissue showed large numbers of degenerated follicles after 24 hours of in vitro culture. We therefore evaluated DMSO perfusion rates by X-ray computed tomography, ice crystal formation by freeze-substitution, and cell toxicity by transmission electron microscopy, seeking possible reasons why follicles degenerated. Although cryoprotectant perfusion was considered normal and no ice crystals were formed in the tissue, ultrastructural analysis detected typical signs of DMSO toxicity, such as mitochondria degeneration, alterations in chromatin condensation, cell vacuolization and extracellular matrix swelling in both stromal and follicular cells. The findings indicated that the method failed to preserve follicles due to the high concentrations of DMSO used. However, adaptations can be made to avoid toxicity to follicles caused by elevated levels of cryoprotectants.

show abstract

“…To rationally design cryopreservation protocols for tissue vitrification, insights in permeation kinetics of CPAs into tissues are needed to ensure maximum permeation and homogeneous distribution of CPAs while minimizing the exposure time and toxicity effects. Various methods have been applied to study CPA permeation in tissues, including nuclear magnetic resonance 22,23 and X-ray computer tomography 24,25 . Furthermore, osmometer measurements of solutions in which CPA-permeated tissue was equilibrated have been employed to derive CPA diffusion coefficients 26,27 .…”

Section: Introductionmentioning

confidence: 99%

Spectroscopic monitoring of transport processes during loading of ovarian tissue with cryoprotective solutions

Han¹,

Sydykov

Yang³

et al. 2019

Sci Rep

View full text Add to dashboard Cite

There is an increasing demand for female fertility preservation. Cryopreservation of ovarian cortex tissue by means of vitrification can be done ad-hoc and for pre-pubertal individuals. Obtaining a homogeneous distribution of protective agents in tissues is one of the major hurdles for successful preservation. Therefore, to rationally design vitrification strategies for tissues, it is needed to determine permeation kinetics of cryoprotective agents; to ensure homogeneous distribution while minimizing exposure time and toxicity effects. In this study, Fourier transform infrared spectroscopy (FTIR) was used to monitor diffusion of different components into porcine ovarian cortex tissue. Water fluxes and permeation kinetics of dimethyl sulfoxide (DMSO), glycerol (GLY), ethylene glycol (EG), and propylene glycol (PG) were investigated. Diffusion coefficients derived from FTIR data, were corroborated with differential scanning calorimetry and osmometer measurements. FTIR allowed real-time spectral fingerprinting of tissue during loading with mixtures of protective agents, while discriminating between different components and water. Exposure to vitrification solutions was found to cause drastic initial weight losses, which could be correlated with spectral features. Use of heavy water allowed distinguishing water fluxes associated with dehydration and permeation, both of which were found to precede permeation of cryoprotective agents. Overall, DMSO and EG were found to permeate faster than GLY and PG. In mixtures, however, solutes behave differently. The non-invasive spectroscopic method described here to study permeation of vitrification solution components into ovarian tissue can be applied to many other types of engineered constructs, tissues, and possibly organs.

show abstract

Assessment of the cryoprotectant concentration inside a bulky organ for cryopreservation using X-ray computed tomography

Cited by 21 publications

References 42 publications

Me2SO perfusion time for whole-organ cryopreservation can be shortened: Results of micro-computed tomography monitoring during Me2SO perfusion of rat hearts

Me2SO perfusion time for whole-organ cryopreservation can be shortened: Results of micro-computed tomography monitoring during Me2SO perfusion of rat hearts

Stepped vitrification technique for human ovarian tissue cryopreservation

Spectroscopic monitoring of transport processes during loading of ovarian tissue with cryoprotective solutions

Contact Info

Product

Resources

About