Wang Wenjing, Upshaw Lisa, Zhang Guangming, Strong D Michael, Reems Jo-Anna
Islet and Cell Processing Laboratory, Puget Sound Blood Center/Northwest Tissue Center, 921 Terry Avenue, Seattle, WA 98104, USA.
Cell Tissue Bank. 2007;8(3):187-94. doi: 10.1007/s10561-006-9029-5. Epub 2006 Nov 9.
Despite improvements and recent attempts to standardize techniques to isolate islets from human donor pancreata, there still exists the problem of consistently recovering sufficient quantities of high quality islets. Moreover, achieving consistent recoveries of high numbers of good quality islets becomes even more challenging from marginal grade human donor pancreata with prolonged cold ischemic times. In this study, we investigate whether addition of Pefabloc SC, a serine protease inhibitor, in combination with Pulmozyme, a recombinant human DNase I, to Liberase HI improves islet isolation outcome from marginal grade human donor pancreata (cold ischemic time > 12 h). Twenty-three marginal grade human donor pancreata were randomly digested using four different enzyme preparations: (1) Liberase alone (n = 6), (2) +Pefabloc (n = 7), (3) +Pefabloc/Pulmozyme (n = 5), and (4) +Pulmozyme (n = 5). Overall, there were no significant differences in donor age, body mass index (BMI), pancreas weight, and cold ischemic time. After purification, significantly higher islet yields (3,281 +/- 590 IE/g) were obtained with the Pefabloc/Pulmozyme group as compared to the Liberase alone (1,615 +/- 305 IE/g) or the Pefabloc group (1,255 +/- 261 IE/g) (P < 0.05). Significant improvements in islet viability were also noted from the Pefabloc/Pulmozyme group (87.3 +/- 4.4%) as opposed to islets isolated from the Pefabloc group (75.2 +/- 3.9%) (P < 0.05). No significant differences in insulin secretory response to glucose stimulation among the four groups were observed, which indicates that the addition of Pefabloc and/or Pulmozyme does not have a detrimental effect on the functionality of islets. It is concluded that the addition of Pefabloc in combination with Pulmozyme to the Liberse HI significantly improves islet isolation outcome and potentially impacts the viability and morphology of the islets obtained from marginal grade human donor pancreata with prolonged cold ischemic times.
尽管在改进以及近期尝试将从人类供体胰腺中分离胰岛的技术标准化方面取得了进展，但始终存在持续获取足量高质量胰岛的问题。此外，对于冷缺血时间延长的边缘质量等级的人类供体胰腺，要始终获得大量高质量的胰岛更是一项挑战。在本研究中，我们探究了在 Liberase HI 中添加丝氨酸蛋白酶抑制剂 Pefabloc SC 并联合重组人脱氧核糖核酸酶 I（Pulmozyme）是否能改善从边缘质量等级的人类供体胰腺（冷缺血时间>12 小时）中分离胰岛的结果。23 个边缘质量等级的人类供体胰腺被随机采用四种不同的酶制剂进行消化：（1）仅使用 Liberase（n = 6），（2）+Pefabloc（n = 7），（3）+Pefabloc/Pulmozyme（n = 5），以及（4）+Pulmozyme（n = 5）。总体而言，供体年龄、体重指数（BMI）、胰腺重量和冷缺血时间方面无显著差异。纯化后，与仅使用 Liberase 组（1,615 ± 305 胰岛当量/g）或 Pefabloc 组（1,255 ± 261 胰岛当量/g）相比，Pefabloc/Pulmozyme 组获得的胰岛产量显著更高（3,281 ± 590 胰岛当量/g）（P < 0.05）。与从 Pefabloc 组分离的胰岛（75.2 ± 3.9%）相比，Pefabloc/Pulmozyme 组的胰岛活力也有显著提高（87.3 ± 4.4%）（P < 0.05）。四组之间在对葡萄糖刺激的胰岛素分泌反应方面未观察到显著差异，这表明添加 Pefabloc 和/或 Pulmozyme 对胰岛的功能没有不利影响。得出的结论是，在 Liberse HI 中添加 Pefabloc 并联合 Pulmozyme 可显著改善胰岛分离结果，并可能影响从冷缺血时间延长的边缘质量等级的人类供体胰腺中获得的胰岛的活力和形态。
