Lakey J R, Rajotte R V, Warnock G L, Kneteman N M
Department of Surgery, University of Alberta, Edmonton, Canada.
Transplantation. 1995 Mar 15;59(5):689-94. doi: 10.1097/00007890-199503150-00008.
We have retrospectively evaluated islet isolation records collected from 230 consecutive adult pancreases with 146 pancreases fulfilling all criteria for our evaluation. Fifty-six pancreases procured by our local organ procurement team (33 before in situ vascular flush and 23 after in situ vascular flush with University of Wisconsin [UW] solution) were compared with 90 pancreases received from distant centers that were shipped in UW solution after in situ vascular flushing. Cold storage of 3-26 hr preceded islet isolation using collagenase digestion and Ficoll purification. Recoveries of islets were assessed by duplicate counts of dithizone-stained aliquots before and after purification. Isolations were considered successful if > 100,000 viable islets (islet equivalents to 150 microns) were recovered after purification. Islet function was assessed by in vitro glucose-stimulated perifusion and islets were considered viable if the stimulation index (glucose stimulated over basal insulin secretion) was > 2. Eighty-three percent of the isolations from locally procured, UW-flushed pancreases were successful, as compared with 86% for pancreases that were stored for 3 to 8 hr, 73% for 8 to 16 hr of storage, and 38% for isolations following > 16 hr of cold storage. Increasing the duration of cold storage prior to islet isolation was associated with an increased proportion of failed isolations and decreased measures of islet viability. The actual numbers of islets per gram of pancreas before and after purification were significantly reduced if the hypothermic cold storage was > 16 hr. In vitro viability of isolated islets during glucose-stimulated perifusion showed a higher percentage of viable islets from pancreases with shorter durations of cold storage. For pancreases with > 16 hr of cold storage prior to islet isolation, islet viability was significantly reduced. We conclude that, with the current methods available to recover and store cadaver donor pancreases and the methods currently used to isolate human islets, yields of purified islets decline significantly from human pancreases that have been subjected to cold storage of > 16-18 hr.
我们回顾性评估了从230例连续的成人胰腺中收集的胰岛分离记录，其中146例胰腺符合我们评估的所有标准。将我们当地器官采购团队获取的56例胰腺（33例在原位血管冲洗前，23例在使用威斯康星大学（UW）溶液进行原位血管冲洗后）与从远处中心接收的90例胰腺进行比较，这些胰腺在原位血管冲洗后用UW溶液运送。在使用胶原酶消化和Ficoll纯化进行胰岛分离之前，进行了3 - 26小时的冷藏。通过对纯化前后双硫腙染色等分试样的重复计数来评估胰岛回收率。如果纯化后回收的活胰岛（胰岛当量为150微米）超过100,000个，则认为分离成功。通过体外葡萄糖刺激灌流评估胰岛功能，如果刺激指数（葡萄糖刺激的胰岛素分泌量超过基础胰岛素分泌量）大于2，则认为胰岛存活。从当地采购的、经UW冲洗的胰腺中进行的分离，83%成功，相比之下，冷藏3至8小时的胰腺分离成功率为86%，冷藏8至16小时的为73%，冷藏超过16小时后的分离成功率为38%。胰岛分离前冷藏时间的延长与分离失败比例的增加以及胰岛活力指标的降低相关。如果低温冷藏时间超过16小时，每克胰腺纯化前后的实际胰岛数量会显著减少。在葡萄糖刺激灌流期间，分离胰岛的体外活力显示，冷藏时间较短的胰腺中活胰岛的百分比更高。对于胰岛分离前冷藏时间超过16小时的胰腺，胰岛活力显著降低。我们得出结论，采用目前可用于回收和储存尸体供体胰腺的方法以及目前用于分离人胰岛所使用的方法，来自冷藏时间超过16 - 18小时的人胰腺的纯化胰岛产量会显著下降。
