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

Abstract

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.

Diffusion kinetics and perfusion times in tissue models obtained by bioorthogonal Raman μ-spectroscopy

“…Diffusion kinetics in biological tissues have been investigated for decades ( 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 ). Quantification of such transport phenomena is pivotal to pharmacology ( 9 , 10 , 11 ), cryopreservation ( 12 , 13 , 14 ), metabolism and cell signaling studies ( 1 , 2 , 3 , 5 , 6 ), food science ( 15 , 16 ), sample preparation ( 11 ), and medical diagnostics ( 17 , 18 , 19 ). On the macroscopic scale, magnetic resonance imaging ( 20 , 21 ), attenuated total reflection spectroscopy ( 14 , 22 ), and permeation into acceptor reservoirs ( 6 , 9 ) are the predominant analytical techniques.…”

Bioinspired Cryoprotectants Enabled by Binary Natural Deep Eutectic Solvents for Sustainable and Green Cryopreservation

“…However, DMSO shows non-negligible cytotoxicity and has side effects on cell genes, , extremely restraining its widespread application in some areas with safety concerns such as the food industry. Currently, glycerol is still the mainstream cryoprotectant for cryopreserving food-related microorganisms, while glycerol is a relatively weak cryoprotectant and shows relatively poor penetrability even if it is classified as a typical penetrating cryoprotectant, , affecting its cryoprotective effects. Besides, glycerol also exerts cytotoxicity at higher concentrations, causing potential food safety issues.…”