Lahmy Reyhaneh, Soleimani Masoud, Sanati Mohammad H, Behmanesh Mehrdad, Kouhkan Fatemeh, Mobarra Naser
Department of Genetics, Faculty of Biology Sciences, Tarbiat Modares University, Tehran, Iran.
Department of Haematology, School of Medicine, Tarbiat Modares University, Tehran, Iran.
J Tissue Eng Regen Med. 2016 Jun;10(6):527-34. doi: 10.1002/term.1787. Epub 2013 Jul 30.
Development of stem cell-based therapies for the treatment of type 1 diabetes would provide a renewable supply of human β-cells. Human embryonic stem cells (ESCs) are considered to be one of the stem cell populations with sufficient proliferative capacity to achieve this goal. Currently, differentiation protocols directing ESCs toward a pancreatic fate employ a variety of expensive cytokines and inhibitors. With the known significance of microRNAs in islet development, we present a novel and cost-effective strategy in which miR-375 overexpression promotes pancreatic endocrine differentiation in hESCs in the absence of any extrinsic factors. miR-375 has been shown to be a key regulator of pancreatic development and function in zebrafish, mouse and human. In this study, hESCs were transduced with lentiviral vectors containing human miR-375 precursor and aggregated to form human embryoid bodies (hEBs) for up to 21 days. Morphological assessment, immunocytochemistry and DTZ staining confirmed that miR-375-induced hEBs have similar characteristics to those of mature islets. In addition, the dynamic expression profile of endodermal marker Foxa2 and endocrine-specific genes, including HNF4α, Pdx1, Pax6, Nkx6.1, Glut2 and insulin, were detected by quantitative real-time PCR. Finally, insulin release upon glucose stimulation was detected in our differentiated clusters. The data presented here demonstrate the feasibility of using microRNAs to direct differentiation into the pancreatic lineage. Copyright © 2013 John Wiley & Sons, Ltd.
开发基于干细胞的疗法来治疗1型糖尿病将提供可再生的人类β细胞来源。人类胚胎干细胞（ESCs）被认为是具有足够增殖能力以实现这一目标的干细胞群体之一。目前，将ESCs诱导分化为胰腺细胞命运的方案使用了多种昂贵的细胞因子和抑制剂。鉴于微小RNA在胰岛发育中的已知重要性，我们提出了一种新颖且具有成本效益的策略，即miR-375过表达在无任何外源性因子的情况下促进人胚胎干细胞向胰腺内分泌细胞分化。在斑马鱼、小鼠和人类中，miR-375已被证明是胰腺发育和功能的关键调节因子。在本研究中，用人miR-375前体的慢病毒载体转导人胚胎干细胞，并聚集形成人胚状体（hEBs）长达21天。形态学评估、免疫细胞化学和DTZ染色证实，miR-375诱导的hEBs具有与成熟胰岛相似的特征。此外，通过定量实时PCR检测了内胚层标志物Foxa2和内分泌特异性基因（包括HNF4α、Pdx1、Pax6、Nkx6.1、Glut2和胰岛素）的动态表达谱。最后，在我们分化的细胞簇中检测了葡萄糖刺激后的胰岛素释放。本文提供的数据证明了使用微小RNA指导分化为胰腺谱系的可行性。版权所有© 2013约翰威立父子有限公司。
