In vitro drug screening for type 1 diabetes therapies has largely been conducted on human organ donor islets for proof of efficacy. While native islets are the ultimate target of these drugs (either in situ or for transplantation), significant benefit can be difficult to ascertain due to the highly heterogeneous nature of individual donors and the overall scarcity of human islets for research. We present an in vitro coculture model based on immortalized insulin-producing beta-cell lines with human endothelial cells in 3D spheroids that aims to recapitulate the islet morphology in an effort towards developing a standardized cell model for in vitro diabetes research. Human insulin-producing immortalized EndoC-βH5 cells are cocultured with human endothelial cells in varying ratios to evaluate 3D cell culture models for type 1 diabetes research. Insulin secretion, metabolic activity, live cell fluorescence staining, and gene expression assays were used to compare the viability and functionality of spheroids composed of 100% beta-cells, 1 : 1 beta-cell/endothelial, and 1 : 3 beta-cell/endothelial. Monoculture and βH5/HUVEC cocultures formed compact spheroids within 7 days, with average diameter ~140 μm. This pilot study indicated that stimulated insulin release from 0 to 20 mM glucose increased from ~8-fold for monoculture and 1 : 1 coculture spheroids to over 20-fold for 1 : 3 EndoC-βH5/HUVEC spheroids. Metabolic activity was also ~12% higher in the 1 : 3 EndoC-βH5/HUVEC group compared to other groups. Stimulating monoculture beta-cell spheroids with 20 mM glucose +1 μg/mL glycine-modified INGAP-P increased the insulin stimulation index ~2-fold compared to glucose alone. Considering their availability and consistent phenotype, EndoC-βH5-based spheroids present a useful 3D cell model for in vitro testing and drug screening applications.

中文翻译：

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用于 1 型糖尿病研究的可再生人类细胞模型：EndoC-βH5/HUVEC 共培养球体

1 型糖尿病疗法的体外药物筛选主要在人体器官供体胰岛上进行，以证明疗效。虽然天然胰岛是这些药物的最终目标（无论是原位还是移植），但由于个体供体的高度异质性以及用于研究的人类胰岛的总体稀缺性，可能很难确定显着的益处。我们提出了一种基于永生化产生胰岛素的 β 细胞系与 3D 球体中的人内皮细胞的体外共培养模型，旨在重现胰岛形态，努力开发用于体外糖尿病研究的标准化细胞模型。将产生人胰岛素的永生化 EndoC- β H5 细胞与人内皮细胞以不同比例共培养，以评估 1 型糖尿病研究的 3D 细胞培养模型。使用胰岛素分泌、代谢活性、活细胞荧光染色和基因表达测定来比较由 100% β 细胞、1:1 β 细胞/内皮细胞和 1:3 β 细胞/内皮组成的球体的活力和功能。内皮细胞。单一培养和β H5/HUVEC共培养在7天内形成致密的球体，平均直径~  140μm。该初步研究表明，0 至 20 mM 葡萄糖刺激的胰岛素释放从单一培养和 1:1 共培养球体的约 8 倍增加到 1:3 EndoC- β H5/HUVEC 球体的 20 倍以上。与其他组相比， 1:3 EndoC- β H5/HUVEC 组的代谢活性也高出约 12% 。用 20 mM 葡萄糖 +1 μg /mL 甘氨酸修饰的 INGAP-P刺激单一培养的 β 细胞球体， 与单独使用葡萄糖相比，胰岛素刺激指数增加约 2 倍。考虑到其可用性和一致的表型，基于 EndoC- β H5 的球体为体外测试和药物筛选应用提供了有用的 3D 细胞模型。