Next Generation Sequencing Facilitates Quantitative Analysis of adipose tissue Treg and LN Treg Transcriptomes

The goal of this study is to reveal unique features of adipose Tregs and to study mechanisms underlying how mediator Med23 regulate adipose Treg function in aged mice. Therefore, mRNA profiles of LN Tregs (from 4-month old Foxp3Cre mice) and adipose Tregs (from 4-month old Foxp3Cre, 4-month old obese Foxp3Cre, 10-month old Foxp3Cre and 10-month old Med23fl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq2000. We compared transcriptome features between lymph node-derived Tregs and adipose Tregs from 4-month old lean or obese mice. Using unbiased comparative gene expression analyses, we found adipose Tregs display an up-signature of Insr (Insulin receptor) and Hif1a, while Pparg acts as a positive control. We next compared gene expression profiles of adipose Tregs from 10-month old Med23fl/fl;Foxp3Cre (MKO) and Foxp3Cre (WT) mice. adipose Med23-deltaTreg cells display impaired transcription of Pparg and Il1rl1 (ST2), and they simultaneously acquire the expression of Nt5e (CD73), while Entpd1 (CD39) expression was not dramatically altered. Our studies implicate protective roles of CD73hi adipose Tregs and offer new therapeutic strategies against age-associated metabolic syndrome. mRNA profiles of LN Tregs (from 4-month old Foxp3Cre mice) and adipose Tregs (from 4-month-old Foxp3Cre, 4-month-old obese Foxp3Cre, 10-month old Foxp3Cre and 10-month-old Med23fl/fl;Foxp3Cre mice) were generated by deep sequencing, single experiment, using Illumina HiSeq2000.本研究旨在揭示脂肪组织调节性T细胞（adipose Tregs）的独特特征，并阐明介导因子Med23调控老年小鼠脂肪Treg功能的潜在机制。为此，我们采用Illumina HiSeq2000平台开展单批次深度测序，获取了淋巴结调节性T细胞（LN Tregs，取自4月龄Foxp3Cre小鼠）及脂肪Treg（分别取自4月龄Foxp3Cre小鼠、4月龄肥胖Foxp3Cre小鼠、10月龄Foxp3Cre小鼠与10月龄Med23fl/fl;Foxp3Cre小鼠）的mRNA表达谱。我们对比了4月龄瘦型或肥胖小鼠的淋巴结来源Treg与脂肪Treg的转录组特征。通过无偏比较基因表达分析，我们发现脂肪Treg呈现胰岛素受体（Insr）与缺氧诱导因子1α（Hif1a）的高表达特征，而过氧化物酶体增殖物激活受体γ（Pparg）作为阳性对照。随后我们对比了10月龄Med23fl/fl;Foxp3Cre（MKO）小鼠与Foxp3Cre（WT，野生型）小鼠的脂肪Treg基因表达谱，结果显示，脂肪组织Med23缺失的Treg细胞中，Pparg与白细胞介素1受体样1（Il1rl1，又称ST2）的转录功能受损，同时获得了5'-核苷酸酶（Nt5e，又称CD73）的表达，而外核苷酸三磷酸二磷酸水解酶1（Entpd1，又称CD39）的表达未发生显著改变。本研究揭示了CD73高表达脂肪Treg的保护作用，并为年龄相关代谢综合征的治疗提供了全新策略。我们再次采用Illumina HiSeq2000平台开展单批次深度测序，获取了淋巴结调节性T细胞（LN Tregs，取自4月龄Foxp3Cre小鼠）及脂肪Treg（分别取自4月龄Foxp3Cre小鼠、4月龄肥胖Foxp3Cre小鼠、10月龄Foxp3Cre小鼠与10月龄Med23fl/fl;Foxp3Cre小鼠）的mRNA表达谱。