Rich S J, Armitage W J
Department of Ophthalmology, Bristol University, United Kingdom.
Cryobiology. 1992 Apr;29(2):153-64. doi: 10.1016/0011-2240(92)90016-u.
Equilibration of corneas with sufficiently high concentrations of cryoprotectants to inhibit potentially damaging ice formation during cryopreservation has not yet been achieved. This study examined the effects on the structure and function of rabbit corneal endothelium of the low toxicity cryoprotectant glycerol. Corneas were exposed to concentrations ranging from 2.0 to 6.8 M glycerol in a Hepes-buffered Ringer's solution containing glutathione, adenosine, 5 mM sodium bicarbonate and 6% w/v bovine serum albumin. Endothelial function was assessed by monitoring corneal thickness during perfusion of the endothelial surface at 34 degrees C for 6 h. Endothelial structure was observed using specular microscopy during perfusion and scanning electron microscopy after perfusion. Corneas tolerated exposure to 2.0 and 3.4 M glycerol for 20 min at 4 and -5 degrees C, respectively. Tolerance of 4.8 M glycerol for 10 min at -10 degrees C was improved by decreasing the dilution temperatures. Ten-minute exposure to 6.1 and 6.8 M glycerol was tolerated at -15 degrees C. In all cases corneas initially showed signs of damage but endothelial function was regained following structural repair. Corneas exposed to 6.8 M glycerol and cooled below the glass transition temperature were nonfunctional after warming. Ice formation during warming was believed to be the cause of injury.
在冷冻保存过程中，使角膜与足够高浓度的冷冻保护剂达到平衡以抑制潜在的破坏性冰晶形成尚未实现。本研究考察了低毒性冷冻保护剂甘油对兔角膜内皮结构和功能的影响。将角膜暴露于含有谷胱甘肽、腺苷、5 mM碳酸氢钠和6% w/v牛血清白蛋白的Hepes缓冲林格氏溶液中，甘油浓度范围为2.0至6.8 M。通过在34℃下对内皮表面灌注6小时期间监测角膜厚度来评估内皮功能。在灌注期间使用镜面显微镜观察内皮结构，并在灌注后使用扫描电子显微镜观察。角膜分别在4℃和-5℃下耐受暴露于2.0 M和3.4 M甘油20分钟。通过降低稀释温度，提高了角膜在-10℃下对4.8 M甘油10分钟的耐受性。在-15℃下，角膜耐受暴露于6.1 M和6.8 M甘油10分钟。在所有情况下，角膜最初均显示出损伤迹象，但在结构修复后内皮功能得以恢复。暴露于6.8 M甘油并冷却至低于玻璃化转变温度的角膜在复温后无功能。复温期间的冰晶形成被认为是损伤的原因。
