Deletion of miRNA processing enzyme Dicer in POMC-expressing cells leads to neurodegeneration and development of obesity

The activity of the endoribonuclease Dicer is crucial to produce the mature form of most microRNAs (miRNAs). Recent studies indicate that lack of miRNAs in different neuronal types results in a range of anatomical and behavioural phenotypes. In the present study we aimed to investigate the developmental and metabolic consequences of miRNA ablation in hypothalamic POMC neurons studying mice with a conditional deletion of Dicer in this population of neurons (POMCDicerKO). These mice exhibited a progressive obese phenotype characterized by hyperphagia, increased adiposity, hyperleptinemia, defective glucose metabolism and alterations in the pituitary-adrenal axis. The development of the obese phenotype was paralleled by a POMC neuron degenerative process that was complete by 6 weeks of age. Furthermore, immunohistochemistry and gene expression studies in control C57Bl/6 adult mice showed that Dicer was expressed in relevant hypothalamic areas and neurons implicated in energy balance, and that its expression was regulated by nutrient availability. Collectively, our results highlight a crucial role for miRNAs in POMC neuron survival and the consequent development of neurodegenerative obesity. Total RNA was extracted from hypothalamic microdissections of 2-week old control and POMCDicerKO mice using the RNeasy micro spin columns (Qiagen, Venlo, The Netherlands). Ten micrograms of total RNA were converted into cRNA, biotinylated, fragmented, and hybridized to GeneChip Mouse Genome 430 2.0 (Affymetrix, Santa Clara, CA). Six microarrays were hybridized with three independent samples from control and POMCDicerKO mice.核糖核酸内切酶Dicer (endoribonuclease Dicer) 的活性对于绝大多数microRNAs (miRNAs) 的成熟生成至关重要。近期研究表明，不同神经元类型中miRNAs的缺失会引发一系列解剖学与行为学表型异常。本研究旨在探究下丘脑POMC神经元中miRNA敲除所带来的发育与代谢后果，通过构建该神经元群体中Dicer条件性敲除的小鼠（POMCDicerKO）开展研究。该小鼠模型呈现出渐进性肥胖表型，具体表现为摄食过多、体脂堆积、高瘦素血症、糖代谢异常以及垂体-肾上腺轴功能紊乱。肥胖表型的发生与POMC神经元的退行性变化同步，该退行性过程在小鼠6周龄时完全完成。此外，对C57BL/6成年对照小鼠开展的免疫组织化学与基因表达分析显示，Dicer在与能量平衡相关的下丘脑区域及神经元中均有表达，且其表达水平受营养可获得性的调控。综上，本研究结果证实了miRNAs在POMC神经元存活中的关键作用，以及由此引发的退行性肥胖的发生发展。本研究使用RNeasy微型离心柱（Qiagen，荷兰芬洛），从2周龄对照小鼠与POMCDicerKO小鼠的下丘脑显微解剖组织中提取总RNA。取10μg总RNA反转录为cRNA，经生物素标记、片段化后，与GeneChip小鼠基因组430 2.0芯片（Affymetrix，美国加利福尼亚州圣克拉拉）进行杂交。本实验共使用对照组与POMCDicerKO组各3份独立样本，完成6次芯片杂交。