Woods E J, Zieger M A, Lakey J R, Liu J, Critser J K
Cryobiology Research Institute, Methodist Hospital of Indiana, Inc., Indianapolis, Indiana, USA.
Cryobiology. 1997 Sep;35(2):106-13. doi: 10.1006/cryo.1997.2029.
Cryopreservation protocols for pancreatic islets may be optimized by modeling the changes in volume that occur during cryoprotectant addition/dilution and cooling/warming. Modeling may be facilitated if it can be demonstrated that isolated islets behave as ideal osmometers where the equilibrium volume is a linear function of the reciprocal of the osmolality of the extracellular medium (the Boyle van't Hoff relationship). The present study was performed to test the hypotheses that: (i) human and canine islets exhibit a linear osmotic response during exposure to hypo-, iso-, and hyperosmotic solutions and to determine the human and canine islets osmotically inactive fraction (Vbp); (ii) to determine if the ionic properties of a concentrated electrolyte solution have an effect on the osmotic response; and (iii) to determine if the osmotic response of pancreatic islets varies between species (human and canine). Islets were isolated from four human and four canine pancreases using standard collagenase digestion and EuroFicoll purification. After 12 h incubation at 37 degrees C, islets (n = 4 from each isolation) were exposed to varying salt or sucrose concentrations (75-1200 mOsm/kg). Islets were held by a micropipette system attached to an inverted microscope, and solutions were exchanged while videotaping. An imaging system was used to determine the cross-sectional area of the islet which was then used to estimate the spherical volume. The results of linear regression analysis indicated that both human and canine islets respond ideally to osmotic conditions within a range of 150-1200 mOsm/kg in both the electrolyte (salt) and the nonelectrolyte (sucrose) solutions. The Vbp value (mean +/- SD) of the four donors was 0.513 +/- 0.068 for canine islets in sucrose solutions, 0.474 +/- 0.068 for canine islets in salt solutions, 0. 425 +/- 0.073 for human islets in sucrose solutions, and 0.39 +/- 0. 073 for human islets in salt solutions. There was a significant difference between human and canine islets (P < 0.05) but not between experimental solutes (NaCl or sucrose) (P < 0.05). These data may be used in conjunction with osmotic tolerance data to develop improved methods for the cryopreservation of isolated islets.
通过对添加/稀释冷冻保护剂以及冷却/升温过程中发生的体积变化进行建模，可以优化胰岛的冷冻保存方案。如果能够证明分离的胰岛表现为理想的渗透压计，即平衡体积是细胞外介质渗透压倒数的线性函数（玻意耳-范特霍夫关系），那么建模过程可能会更加容易。本研究旨在验证以下假设：（i）人和犬的胰岛在暴露于低渗、等渗和高渗溶液时表现出线性渗透反应，并确定人和犬胰岛的非渗透活性部分（Vbp）；（ii）确定浓缩电解质溶液的离子特性是否对渗透反应有影响；（iii）确定胰腺胰岛的渗透反应在不同物种（人和犬）之间是否存在差异。使用标准胶原酶消化和EuroFicoll纯化方法从四个人和四只犬的胰腺中分离胰岛。在37℃孵育12小时后，将胰岛（每次分离n = 4）暴露于不同的盐或蔗糖浓度（75 - 1200 mOsm/kg）。胰岛通过连接到倒置显微镜的微量移液器系统固定，在录像的同时更换溶液。使用成像系统确定胰岛的横截面积，然后用于估计球体体积。线性回归分析结果表明，人和犬的胰岛在电解质（盐）和非电解质（蔗糖）溶液中，在150 - 1200 mOsm/kg的范围内对渗透条件均表现出理想反应。四只供体犬胰岛在蔗糖溶液中的Vbp值（平均值±标准差）为0.513±0.068，在盐溶液中为0.474±0.068；人胰岛在蔗糖溶液中的Vbp值为0.425±0.073，在盐溶液中为0.39±0.073。人和犬胰岛之间存在显著差异（P < 0.05），但实验溶质（NaCl或蔗糖）之间无显著差异（P < 0.05）。这些数据可与渗透耐受性数据结合使用，以开发改进的分离胰岛冷冻保存方法。
