Insulin-Producing Endocrine Cells Differentiated In Vitro From Human Embryonic Stem Cells Function in Macroencapsulation Devices In Vivo
Insulin-Producing Endocrine Cells Differentiated In Vitro From Human Embryonic Stem Cells Function in Macroencapsulation Devices In Vivo
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DOI：
10.5966/sctm.2015-0079
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发表时间：
2015-10-01
影响因子：
6
通讯作者：
D'Amour, Kevin A.
中科院分区：
文献类型：
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作者：
Agulnick, Alan D.;Ambruzs, Dana M.;D'Amour, Kevin A.
PEC-01细胞群由人胚胎干细胞（hESC）分化而来，含有胰腺祖细胞（PP），当将其装入大胶囊装置（以生产VC-01候选产品）并移植到小鼠体内时，可以成熟为葡萄糖反应性胰岛素分泌细胞和参与葡萄糖代谢的其他胰腺内分泌细胞。我们修改了制备 PEC-01 细胞的方案，使 73%-80% 的细胞群由 PDX1 阳性 (PDX1(+)) 和 NKX6.1(+) PP 组成。 PP 进一步分化为胰岛样细胞 (IC)，可重复地含有 73%-89% 的内分泌细胞，其中大约 40%-50% 表达胰岛素。这些胰岛素阳性细胞中的很大一部分是单一激素阳性，并表达转录因子 PDX1 和 NKX6.1。为了排除祖细胞对 IC 体内功能的显着贡献，我们使用简单的富集过程来去除剩余的 PP，产生含有 93%-98% 内分泌细胞和 1%-3% 祖细胞的聚集体。富集的 IC 在封装并植入小鼠体内时，其功能与 VC-01 候选产品类似，最终证明体外产生的 hESC 衍生的胰岛素生成细胞可以在设备中成熟并在体内发挥作用。使用我们的悬浮培养物的缩放版本，并且内分泌聚集体可以冷冻保存并保留功能。尽管 IC 表达多种重要的 (3 个细胞基因)，但细胞含有相对较低水平的几种成熟相关标记。与此相关，IC 发挥作用的时间与 PEC-01 细胞相似，表明 IC 在体内递送后需要细胞自主成熟，这将与移植物整合到宿主中同时发生。干细胞转化医学 2015;4:1214-1222
The PEC-01 cell population, differentiated from human embryonic stem cells (hESCs), contains pancreatic progenitors (PPs) that, when loaded into macroencapsulation devices (to produce the VC-01 candidate product) and transplanted into mice, can mature into glucose-responsive insulin-secreting cells and other pancreatic endocrine cells involved in glucose metabolism. We modified the protocol for making PEC-01 cells such that 73%-80% of the cell population consisted of PDX1-positive (PDX1(+)) and NKX6.1(+) PPs. The PPs were further differentiated to islet-like cells (ICs) that reproducibly contained 73%-89% endocrine cells, of which approximately 40%-50% expressed insulin. A large fraction of these insulin-positive cells were single hormone-positive and expressed the transcription factors PDX1 and NKX6.1. To preclude a significant contribution of progenitors to the in vivo function of ICs, we used a simple enrichment process to remove remaining PPs, yielding aggregates that contained 93%-98% endocrine cells and 1%-3% progenitors. Enriched ICs, when encapsulated and implanted into mice, functioned similarly to the VC-01 candidate product, demonstrating conclusively that in vitro-produced hESC-derived insulin-producing cells can mature and function in vivo in devices. A scaled version of our suspension culture was used, and the endocrine aggregates could be cryo-preserved and retain functionality. Although ICs expressed multiple important (3 cell genes, the cells contained relatively low levels of several maturity-associated markers. Correlating with this, the time to function of ICs was similar to PEC-01 cells, indicating that ICs required cell-autonomous maturation after delivery in vivo, which would occur concurrently with graft integration into the host. STEM CELLS TRANSLATIONAL MEDICINE 2015;4:1214-1222