Hierachical Study on Heat Transfer and Viability of Cell during Freezing of Biological Tissue

冷冻可以减缓或停止一些生物反应，以保存生物组织，解冻可以让它复活。冰冻和解冻对生命系统来说也是致命的。在冷冻过程中，细胞内外结冰、渗透水渗入细胞膜、细胞变形等行为在微观上发生，给细胞带来严重的损伤，关系到活细胞的生死。这些微观行为至少发生在细胞大小的水平上，似乎是由于生物材料的物理化学复杂性。在本研究项目中，首先提出了简化的细胞冻结模型，讨论了细胞在冻结过程中细胞内外结冰、渗透水透过细胞膜等微尺度行为与温度的关系；其次，研究了细胞活力与冻融过程中微观行为的关系。在实验中，对小麦原生质体进行了显微镜观察，并通过荧光试验检测了细胞的活力。在冻融过程中遇到的渗透条件也是通过渗透操作实现的。综合上述结果，综合考虑“溶液效应损伤”和“细胞内冰冻损伤”的影响，估算了细胞在冷冻过程中的损伤程度。在这些结果的基础上，结合热传递过程讨论了细胞在生物组织冷冻过程中的生存能力。

Freezing can slow down or stop some biological reactions for preserving the biological tissue, and thawing can bring it back to life. It is also true that freezing and thawing are lethal to the living system. During freezing, the extra- and intra-cellular ice formation, osmotic water permeation through cell membrane, deformation of cell and other behaviors occur at microscale and they bring serious injuries connecting to the life-and-death of living cells. These micro behaviors occurred on at least cell size level seem to be due to the physicochemical complexity of biological materials. Water is chemically restrained in the states of aqueous solution and colloidal solution, and those are physically confined in small spaces or compartmentalized by the cross-linked network structures.In this research project, firstly, simplified freezing model of cell element was proposed, and the extra- and intra-cellular ice formation, osmotic water permeation through cell membrane and other microscale behavior during the freezing of biological cell were discussed as a function of temperature.Secondly, viability of the cell was studied in relation with the micro-behaviors during freezing and thawing. In experiments, microscopic observation of wheat protoplasts was performed, and the cell viability was inspected by the fluorescence test. The osmotic conditions encountered during freeze-thaw process were also realized by osmotic manipulations. Summarizing these results, the cell damage during freezing process was estimated taking account of "solution effect injury" and "intracellular ice injury".Thirdly, the freezing process of biological tissue was numerically simulated using heat transfer and thermodynamic model. On the basis of these results, viability of the cell during freezing of biological tissue was discussed in conjunction with the heat transfer process.