Theoretical and experimental study of the antifreeze protein AFP752, trehalose and dimethyl sulfoxide cryoprotection mechanism: correlation with cryopreserved cell viability

Abstract

In this work the physico-chemical properties of selected cryoprotectants (antifreeze protein TrxA-AFP752, trehalose and dimethyl sulfoxide) were correlated with their impact on the constitution of ice and influence on frozen/thawed cell viability. The freezing processes and states of investigated materials solutions were described and explained from a fundamental point of view using ab-initio modelling (molecular dynamics, DFT), Raman spectroscopy, Differential Scanning Calorimetry and X-Ray Diffraction. For the first time, in this work we correlated the microscopic view (modelling) with the description of the frozen solution states and put these results in the context of human skin fibroblast viability after freezing and thawing. DMSO and AFP had different impacts on their solution's freezing process but in both cases the ice crystallinity size was considerably reduced. DMSO and AFP treatment in different ways improved the viability of frozen/thawed cells.

Chromatin architecture changes and DNA replication fork collapse are critical features in cryopreserved cells that are differentially controlled by cryoprotectants

“…First, we used the same gates in all experiments, and second, in all cases, we excluded cell debris more precisely (as compared to RSC Adv.) 3 . The same gating and the same style of debris exclusion is extremely important to correctly evaluate viability of freeze-thawed cells because many of these cells are more or less fragmented.…”

“…However, extensive damage occurs to a percentage of frozen and thawed cells and tissues. Though the freeze-thaw process can be greatly affected by the use of cryoprotective additives to improve cell viability 3 , 4 , the effects of freezing and cryoprotectants per se on the complex status of cell nuclei (and the genetic information contained therein) remain controversial 4 – 7 . Contradictory results in the literature have prevented a consensus on the fundamental question of the extent of DNA and chromatin fragmentation that occurs during freezing and thawing 8 – 11 .…”

“…In our previous work 3 , we focused on the formation of ice during freezing as an important parameter that strongly influences cellular destruction and examined specific properties of selected cryoprotectant solutions during freezing, including dimethyl sulfoxide (DMSO), trehalose and a recombinant antifreeze fusion protein (AFP) that was originally isolated from the Anatolica polita desert beetle 2 , 3 . Building on this knowledge, here, we used these cryoprotectants to investigate the importance and extent of chromatin damage in freeze-thawed cells, specifically fragmentation and structural changes of chromatin.…”

Simple Syntheses of New Pegylated Trehalose Derivatives as a Chemical Tool for Potential Evaluation of Cryoprotectant Effects on Cell Membrane

“…Cryopreservation of cells plays an essential role in many areas [1][2][3][4][5]. The main difficulty in cryopreservation is that ice crystallization, which appears throughout the freezing process, can significantly damage the cells and cause the loss of viability after the cells are thawed [6].…”

Antioxidative stress of Chitosan-Gallium Nanoparticles and low doses of γ-irradiation on mice bearing Ehrlich carcinoma

“…Researchers have considered the impacts of low-dose radiation, given as a single dose or fractionated or prolonged over time, with regard to altering cancer progression or resolution as well as their impacts on the systemic immune system [23] .It was found that low-dose radiation increases cellular antioxidant activity; facilitates DNA damage repair; reduces malignant transformation and mutagenesis, and stimulates immune surveillance [24]. The cell nucleus and DNA are susceptible to several stressors[25, 26,27,28] and can be greatly harmed with relatively low doses of ionizing radiation [29,30]. As adverse effects of ionizing radiation on (cancer) cells are mainly mediated by fragmentation of nuclear chromatin via inserting double strand breaks (DSBs) into the DNA molecule [31].…”