Shimoda M, Itoh T, Sugimoto K, Takita M, Chujo D, Iwahashi S, SoRelle J A, Naziruddin B, Levy M F, Grayburn P A, Matsumoto S
Baylor Research Institute, Dallas, Texas, USA.
Transplant Proc. 2011 Nov;43(9):3161-6. doi: 10.1016/j.transproceed.2011.09.098.
Islet purification is mainly performed by the density gradient method. However, purification of the embedded islets that are surrounded by exocrine tissue should be difficult, because their density is similar to exocrine tissue. In this study, we performed chart review to assess the relationship between the ratio of embedded islets and efficacy of purification. Then, we tested several conditions of a new method to free the islets from surrounded exocrine tissues using high osmolality solution with gentle agitation.
First, we performed chart review of our human islet isolation. Second, embedded islet-enriched human islet fractions (embedded islets >50%) were suspended in University of Wisconsin (UW) solution (UW group, 320 mOsm/kg/H(2)0) or osmolality-adjusted UW solution (400, 500, and 600 mOsm/kg/H(2)0; 400 group, 500 group, and 600 group, respectively). Each tube was gently shaken at 4°C. The tissue samples were taken before shaking and after 15, 30, and 60 minutes. Islet yield, percentage of embedded islets, and viabilities were assessed.
The chart review revealed that high ratio of embedded islets deteriorated the efficacy of islet purification. The islet yield in all groups except for the 600 group did not change at 15 minutes, but it decreased in all groups at 60 minutes. The average percentage of embedded islets before shaking was 62.6%. Although percentage of embedded islets were decreasing in all groups, it was < 20% at 15 minutes in the 500 and 600 groups whereas it was >44% in the UW group, which indicated that higher osmolality would have a greater effect. Viability was >95% in all groups at 30 minutes.
The embedded islets deteriorated the efficacy of islet purification. Gentle agitation of embedded islets in high osmolality (500 mOsm/kg/H(2)O, 15 minutes) could release islets from surrounded exocrine tissue.
胰岛纯化主要通过密度梯度法进行。然而，对于被外分泌组织包围的包埋胰岛进行纯化应该很困难，因为它们的密度与外分泌组织相似。在本研究中，我们进行了图表回顾以评估包埋胰岛的比例与纯化效果之间的关系。然后，我们测试了一种新方法的几种条件，该方法使用高渗溶液并轻柔搅拌以使胰岛从周围的外分泌组织中游离出来。
首先，我们对我们的人胰岛分离进行了图表回顾。其次，将富含包埋胰岛的人胰岛组分（包埋胰岛>50%）悬浮于威斯康星大学（UW）溶液（UW组，320 mOsm/kg/H₂O）或渗透压调整后的UW溶液（400、500和600 mOsm/kg/H₂O；分别为400组、500组和600组）中。每个试管在4°C下轻柔摇晃。在摇晃前以及摇晃15、30和60分钟后采集组织样本。评估胰岛产量、包埋胰岛的百分比和活力。
图表回顾显示，包埋胰岛的高比例会降低胰岛纯化的效果。除600组外，所有组在15分钟时胰岛产量未发生变化，但在60分钟时所有组的胰岛产量均下降。摇晃前包埋胰岛的平均百分比为62.6%。尽管所有组中包埋胰岛的百分比都在下降，但500组和600组在15分钟时<20%，而UW组>44%，这表明较高的渗透压会产生更大的影响。所有组在30分钟时活力>95%。
包埋胰岛会降低胰岛纯化的效果。在高渗（500 mOsm/kg/H₂O，15分钟）条件下轻柔搅拌包埋胰岛可使胰岛从周围的外分泌组织中释放出来。
