Mechanism of Intracellular Ice Formation and the Importance of Rapid Warming during Vitrification

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  • 細胞内氷晶形成メカニズムの解明と 細胞内氷晶形成回避における融解速度の重要性
  • サイボウ ナイヒョウショウケイセイ メカニズム ノ カイメイ ト サイボウ ナイヒョウショウケイセイ カイヒ ニ オケル ユウカイ ソクド ノ ジュウヨウセイ

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Abstract

When cells are vitrified with a minimum degree of supercooling, i.e., in near equilibrium, no ice would form in the cell. However, in most cases the degree of supercooling is not minimal, and minute invisible ice crystals will form in cells during the vitrification process. During warming, differences in surface free energy can cause them to recrystallize into larger lethal crystals, which will be lethal to the cells. It is commonly believed that both the cooling rate and the concentration of glass-inducing solutes (cryoprotectants) has to be high during the vitrification process. However, the results that we present here, using mouse oocytes, contradict these beliefs. First, rather than maintaining a high cooling rate during vitrification, our data actually demonstrate that it is the warming rate that should be high. Moreover, our results also contradict the second belief that a high concentration of cryoprotectant is preferred. We show that if the warming rate is extremely rapid, oocyte survival is high even when the vitrification solution contains only a half the normal concentration of cryoprotectants. Therefore, we conclude that rapid warming, rather than cooling or high concentration of cryoprotectant, is essential to prevent intracellular ice formation (recrystallization).

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