The dominance of warming rate over cooling rate in the survival of mouse oocytes subjected to a vitrification procedure.
The formation of more than trace amounts of ice in cells is lethal. The two contrasting routes to avoiding it are slow equilibrium freezing and vitrification. The cryopreservation of mammalian oocytes by either method continues to be difficult, but there seems a slowly emerging consensus that vitrification procedures are somewhat better for mouse and human oocytes. The approach in these latter procedures is to load cells with high concentrations of glass-inducing solutes and cool them at rates high enough to induce the glassy state. Several devices have been developed to achieve very high cooling rates. Our study has been concerned with the relative influences of warming rate and cooling rate on the survival of mouse oocytes subjected to a vitrification procedure. Oocytes suspended in an ethylene glycol-acetamide-Ficoll-sucrose solution were cooled to -196 degrees C at rates ranging from 37 to 1827 degrees C/min between 20 and -120 degrees C, and for each cooling rate, warmed at rates ranging from 139 to 2950 degrees C/min between -70 and -35 degrees C. The results are unambiguous. If the samples were warmed at the highest rate, survivals were >80% over cooling rates of 187-1827 degrees C/min. If the samples were warmed at the lowest rate, survivals were near 0% regardless of the cooling rate. We interpret the lethality of slow warming to be a consequence of it allowing time for the growth of small intracellular ice crystals by recrystallization.
Fundamental and Applied Cryobiology Group, Department of Biochemistry and Cellular and Molecular Biology, The University of Tennessee, Knoxville, TN 37996-0840, USA.
Shinsuke
S
Seki
Peter
P
Mazur
D000818:Animals; D002152:Calorimetry, Differential Scanning / Q000379:methods; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D003460:Crystallization; D005260:Female; D005615:Freezing; D006358:Hot Temperature; D006801:Humans; D007053:Ice; D051379:Mice; D008813:Mice, Inbred ICR; D009865:Oocytes / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors
D003451:Cryoprotective Agents; D007053:Ice