Jafarian Arefeh, Taghikani Mohammad, Abroun Saeid, Allahverdi Amir, Lamei Maryam, Lakpour Niknam, Soleimani Masoud
Department of Clinical Biochemistry, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
Department of Hematology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran.
PLoS One. 2015 Jun 5;10(6):e0128650. doi: 10.1371/journal.pone.0128650. eCollection 2015.
MicroRNAs (miRNAs) are a group of endogenous small non-coding RNAs that regulate gene expression at the post-transcriptional level. A number of studies have led to the notion that some miRNAs have key roles in control of pancreatic islet development and insulin secretion. Based on some studies on miRNAs pattern, the researchers in this paper investigated the pancreatic differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) by up-regulation of miR-375 and down-regulation of miR-9 by lentiviruses containing miR-375 and anti-miR-9.
After 21 days of induction, islet-like clusters containing insulin producing cells (IPCs) were confirmed by dithizone (DTZ) staining. The IPCs and β cell specific related genes and proteins were detected using qRT-PCR and immunofluorescence on days 7, 14 and 21 of differentiation. Glucose challenge test was performed at different concentrations of glucose so extracellular and intracellular insulin and C-peptide were assayed using ELISA kit. Although derived IPCs by miR-375 alone were capable to express insulin and other endocrine specific transcription factors, the cells lacked the machinery to respond to glucose.
It was found that over-expression of miR-375 led to a reduction in levels of Mtpn protein in derived IPCs, while treatment with anti-miR-9 following miR-375 over-expression had synergistic effects on MSCs differentiation and insulin secretion in a glucose-regulated manner. The researchers reported that silencing of miR-9 increased OC-2 protein in IPCs that may contribute to the observed glucose-regulated insulin secretion. Although the roles of miR-375 and miR-9 are well known in pancreatic development and insulin secretion, the use of these miRNAs in transdifferentiation was never demonstrated. These findings highlight miRNAs functions in stem cells differentiation and suggest that they could be used as therapeutic tools for gene-based therapy in diabetes mellitus.
微小RNA（miRNA）是一类内源性小非编码RNA，可在转录后水平调控基因表达。多项研究表明，一些miRNA在胰腺胰岛发育和胰岛素分泌的控制中起关键作用。基于对miRNA模式的一些研究，本文的研究人员通过含有miR-375和抗miR-9的慢病毒上调miR-375并下调miR-9，研究了人骨髓间充质干细胞（hBM-MSCs）的胰腺分化。
诱导21天后，用双硫腙（DTZ）染色确认含有胰岛素产生细胞（IPC）的胰岛样簇。在分化的第7、14和21天，使用qRT-PCR和免疫荧光检测IPC以及β细胞特异性相关基因和蛋白质。在不同浓度的葡萄糖下进行葡萄糖激发试验，因此使用ELISA试剂盒测定细胞外和细胞内胰岛素及C肽。尽管单独由miR-375衍生的IPC能够表达胰岛素和其他内分泌特异性转录因子，但这些细胞缺乏对葡萄糖作出反应的机制。
发现miR-375的过表达导致衍生的IPC中Mtpn蛋白水平降低，而在miR-375过表达后用抗miR-9处理对MSCs分化和胰岛素分泌具有葡萄糖调节的协同作用。研究人员报告说，miR-9的沉默增加了IPC中的OC-2蛋白，这可能有助于观察到的葡萄糖调节的胰岛素分泌。尽管miR-375和miR-9在胰腺发育和胰岛素分泌中的作用是众所周知的，但从未证明这些miRNA在转分化中的用途。这些发现突出了miRNA在干细胞分化中的功能，并表明它们可作为糖尿病基因治疗的治疗工具。
