Laboratory of Chemical Genomics, Shenzhen Graduate School of Peking University, Shenzhen, People's Republic of China.
Diabetologia. 2011 Sep;54(9):2325-36. doi: 10.1007/s00125-011-2246-x. Epub 2011 Jul 14.
AIMS/HYPOTHESIS: The generation of induced pluripotent stem cells (iPSCs) provides a promising possibility for type 1 diabetes therapy. However, the generation of insulin-producing cells from iPSCs and evaluation of their efficacy and safety should be achieved in large animals before clinically applying iPSC-derived cells in humans. Here we try to generate insulin-producing cells from rhesus monkey (RM) iPSCs.
Based on the knowledge of embryonic pancreatic development, we developed a four-stage protocol to generate insulin-producing cells from RM iPSCs. We established a quantitative method using flow cytometry to analyse the differentiation efficiency. In addition, to evaluate the differentiation competence and function of RM iPSC-derived cells, transplantation of stage 3 and 4 cells into immunodeficient mice was performed.
RM iPSCs were sequentially induced to definitive endoderm (DE), pancreatic progenitors (PP), endocrine precursors (EP) and insulin-producing cells. PDX1(+) PP cells were obtained efficiently from RM iPSCs (over 85% efficiency). The TGF-β inhibitor SB431542 promoted the generation of NGN3(+) EP cells, which can generate insulin-producing cells in vivo upon transplantation. Finally, after this four-stage differentiation in vitro, insulin-producing cells that could secrete insulin in response to glucose stimulation were obtained. When transplanted into mouse models for diabetes, these insulin-producing cells could decrease blood glucose levels in approximately 50% of the mice.
CONCLUSIONS/INTERPRETATION: We demonstrate for the first time that RM iPSCs can be differentiated into functional insulin-producing cells, which will provide the basis for investigating the efficacy and safety of autologous iPSC-derived insulin-producing cells in a rhesus monkey model for type 1 diabetes therapy.
目的/假设：诱导多能干细胞（iPSCs）的产生为 1 型糖尿病治疗提供了有前景的可能性。然而，在将 iPSC 衍生细胞临床应用于人类之前，应该在大动物中从 iPSCs 产生胰岛素分泌细胞并评估其疗效和安全性。在这里，我们尝试从恒河猴（RM）iPSCs 中产生胰岛素分泌细胞。
基于对胚胎胰腺发育的认识，我们开发了一个四阶段方案，从 RM iPSCs 中产生胰岛素分泌细胞。我们建立了一种使用流式细胞术分析分化效率的定量方法。此外，为了评估 RM iPSC 衍生细胞的分化能力和功能，我们将第 3 期和第 4 期细胞移植到免疫缺陷小鼠中。
RM iPSCs 依次被诱导为确定的内胚层（DE）、胰腺祖细胞（PP）、内分泌前体细胞（EP）和胰岛素分泌细胞。从 RM iPSCs 中高效获得 PDX1(+) PP 细胞（效率超过 85%）。TGF-β 抑制剂 SB431542 促进 NGN3(+) EP 细胞的产生，这些细胞在体内移植后可产生胰岛素分泌细胞。最后，经过体外四步分化，获得了可响应葡萄糖刺激分泌胰岛素的胰岛素分泌细胞。当移植到糖尿病小鼠模型中时，这些胰岛素分泌细胞可使大约 50%的小鼠血糖水平降低。
结论/解释：我们首次证明 RM iPSCs 可分化为功能性胰岛素分泌细胞，这将为研究在 1 型糖尿病治疗的恒河猴模型中自体 iPSC 衍生的胰岛素分泌细胞的疗效和安全性提供基础。
