Comparison and avoidance of toxicity of penetrating cryoprotectants

Edyta A. Szurek, Ali Eroglu

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

The objective of this study was to elucidate the toxicity of widely used penetrating cryoprotective agents (CPAs) to mammalian oocytes. To this end, mouse metaphase II (M II) oocytes were exposed to 1.5 M solutions of dimethylsulfoxide (DMSO), ethylene glycol (EG), or propanediol (PROH) prepared in phosphate buffered saline (PBS) containing 10% fetal bovine serum. To address the time- and temperature-dependence of the CPA toxicity, M II oocytes were exposed to the aforementioned CPAs at room temperature (RT, ~23°C) and 37°C for 15 or 30 minutes. Subsequently, the toxicity of each CPA was evaluated by examining post-exposure survival, fertilization, embryonic development, chromosomal abnormalities, and parthenogenetic activation of treated oocytes. Untreated oocytes served as controls. Exposure of MII oocytes to 1.5 M DMSO or 1.5 M EG at RT for 15 min did not adversely affect any of the evaluated criteria. In contrast, 1.5 M PROH induced a significant increase in oocyte degeneration (54.2%) and parthenogenetic activation (16%) under same conditions. When the CPA exposure was performed at 37°C, the toxic effect of PROH further increased, resulting in lower survival (15%) and no fertilization while the toxicity of DMSO and EG was still insignificant. Nevertheless, it was possible to completely avoid the toxicity of PROH by decreasing its concentration to 0.75 M and combining it with 0.75 M DMSO to bring the total CPA concentration to a cryoprotective level. Moreover, combining lower concentrations (i.e., 0.75 M) of PROH and DMSO significantly improved the cryosurvival of MII oocytes compared to the equivalent concentration of DMSO alone. Taken together, our results suggest that from the perspective of CPA toxicity, DMSO and EG are safer to use in slow cooling protocols while a lower concentration of PROH can be combined with another CPA to avoid its toxicity and to improve the cryosurvival as well.

Original languageEnglish (US)
Article numbere27604
JournalPLoS One
Volume6
Issue number11
DOIs
StatePublished - Nov 16 2011

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Cryoprotective Agents
cryoprotectants
dimethyl sulfoxide
Oocytes
Dimethyl Sulfoxide
Toxicity
oocytes
toxicity
Ethylene Glycol
ethylene glycol
Metaphase
metaphase
Fertilization
Chemical activation
Propylene Glycols
propanediols
Temperature
chromosome aberrations
Poisons
fetal bovine serum

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)

Cite this

Comparison and avoidance of toxicity of penetrating cryoprotectants. / Szurek, Edyta A.; Eroglu, Ali.

In: PLoS One, Vol. 6, No. 11, e27604, 16.11.2011.

Research output: Contribution to journalArticle

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abstract = "The objective of this study was to elucidate the toxicity of widely used penetrating cryoprotective agents (CPAs) to mammalian oocytes. To this end, mouse metaphase II (M II) oocytes were exposed to 1.5 M solutions of dimethylsulfoxide (DMSO), ethylene glycol (EG), or propanediol (PROH) prepared in phosphate buffered saline (PBS) containing 10{\%} fetal bovine serum. To address the time- and temperature-dependence of the CPA toxicity, M II oocytes were exposed to the aforementioned CPAs at room temperature (RT, ~23°C) and 37°C for 15 or 30 minutes. Subsequently, the toxicity of each CPA was evaluated by examining post-exposure survival, fertilization, embryonic development, chromosomal abnormalities, and parthenogenetic activation of treated oocytes. Untreated oocytes served as controls. Exposure of MII oocytes to 1.5 M DMSO or 1.5 M EG at RT for 15 min did not adversely affect any of the evaluated criteria. In contrast, 1.5 M PROH induced a significant increase in oocyte degeneration (54.2{\%}) and parthenogenetic activation (16{\%}) under same conditions. When the CPA exposure was performed at 37°C, the toxic effect of PROH further increased, resulting in lower survival (15{\%}) and no fertilization while the toxicity of DMSO and EG was still insignificant. Nevertheless, it was possible to completely avoid the toxicity of PROH by decreasing its concentration to 0.75 M and combining it with 0.75 M DMSO to bring the total CPA concentration to a cryoprotective level. Moreover, combining lower concentrations (i.e., 0.75 M) of PROH and DMSO significantly improved the cryosurvival of MII oocytes compared to the equivalent concentration of DMSO alone. Taken together, our results suggest that from the perspective of CPA toxicity, DMSO and EG are safer to use in slow cooling protocols while a lower concentration of PROH can be combined with another CPA to avoid its toxicity and to improve the cryosurvival as well.",
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