Alberta Diabetes Institute, Alberta, Canada.
Department of Surgery, University of Alberta, Edmonton, Alberta, Canada.
Transplantation. 2018 Oct;102(10):e413-e423. doi: 10.1097/TP.0000000000002354.
There is a strong rationale to pursue the use of neonatal porcine islets (NPIs) as an unlimited source of islets for clinical xenotransplantation. Because NPIs are composed of immature insulin producing beta (ß) cells and ductal precursor cells, they provide an ideal model to examine culture conditions to enhance ß cell proliferation and/or ß cell neoformation from ductal cells. In an attempt to optimize the potential of NPIs as a source of ß cell grafts, we used an in vitro differentiation protocol and measured its effect on the functional maturation and differentiation of NPIs.
Pancreata from 1- to 3-day-old neonatal pigs were digested and cultured in standard Ham's F10 media for 5 days. Each independent preparation was then further cultured in Dulbecco's modified Eagle medium nutrient mixture-F12 differentiation media containing growth factors added in a stepwise fashion, or cultured in control Ham's F10 media. After 20 days in culture, islets were assessed for insulin secretory capacity, cellular composition, gene expression, and metabolic activity after transplantation in immunodeficient mice with diabetes.
Compared with control islets, differentiated islets exhibited a significantly higher proportion of endocrine cells, proliferating cell nuclear antigen double positive ß cells, and an enhanced glucose-stimulated insulin secretory activity. Mice transplanted with differentiated islets had significantly lower blood glucose values at weeks 18 and 20 compared with nondifferentiated controls and were shown to be more glucose tolerant.
Culturing NPIs in a 20-day stepwise differentiation media increases the proportion of endocrine cells and augments both in vitro and in vivo function of the islets.
利用新生猪胰岛（NPIs）作为临床异种移植的无限胰岛来源具有充分的理由。因为 NPIs 由不成熟的胰岛素产生β（ß）细胞和导管前体细胞组成，所以它们提供了一个理想的模型来研究培养条件，以增强ß细胞增殖和/或从导管细胞形成新的ß细胞。为了优化 NPIs 作为ß细胞移植物来源的潜力，我们使用了体外分化方案，并测量了其对 NPIs 的功能成熟和分化的影响。
从 1 至 3 天大的新生猪的胰腺中消化并在标准的 Ham's F10 培养基中培养 5 天。每个独立的制剂然后进一步在含有逐步添加生长因子的 Dulbecco 改良 Eagle 培养基营养混合物-F12 分化培养基中培养，或在对照 Ham's F10 培养基中培养。在培养 20 天后，评估胰岛的胰岛素分泌能力、细胞组成、基因表达和在患有糖尿病的免疫缺陷小鼠中的代谢活性。
与对照胰岛相比，分化胰岛表现出更高比例的内分泌细胞、增殖细胞核抗原双阳性ß细胞，以及增强的葡萄糖刺激胰岛素分泌活性。与未分化对照相比，移植分化胰岛的小鼠在第 18 和 20 周的血糖值明显更低，并且表现出更高的葡萄糖耐量。
在 20 天的逐步分化培养基中培养 NPIs 增加了内分泌细胞的比例，并增强了胰岛的体外和体内功能。
