Analysis of gene expression patterns in brown adipose tissue. Analysis of gene expression patterns in brown adipose tissue

Brown adipose tissue is a specialized fat tissue involved in heat generation by using a process termed thermogenesis. This ability to convert nutrient energy into heat sets brown adipocytes apart from the more common type of white adipocytes, which are mainly involved in energy storage. Brown adipose depots occur as a large interscapular depot in mice. Brown adipocytes are multilocular, i.e. contain many small lipid-filled vacuoles and uniquely express the uncoupling protein 1. In this study, the gene expression patterns in brown adipose tissue samples of young, healthy mice was analyzed. Overall design: Male C57Bl6J mice were housed under controlled conditions at 22 ±2 °C in an 12/12-hour light/dark cycle maintained on a standard chow diet (Ssniff, Soest, Germany). The animals were killed at 10 weeks of age by cervical dislocation. Brown adipose tissue depots were isolated and immediately frozen in liquid nitrogen. RNA was isolated from 50 mg of tissue. NGS sequencing was conducted using Illumina HiSeq 2000 with 2 x 100 bp reads (LGC genomics, Berlin, Germany). For data processing and analysis, FASTQC v0.11.8 was used for quality assessment and trimmgalore v0.6.4 was applied for filtering adapter sequences and small reads. The clean reads were mapped to a reference genome (Genome Reference Consortium Mouse Build 38 mm10) using the alignment tools HISAT2 v2.1.0 and Bowtie2 v2.3.5.1 Expression levels were quantified Cufflinks v.2.2.1.

棕色脂肪组织（Brown adipose tissue）是一类特化的脂肪组织，通过被称为产热作用（thermogenesis）的过程参与热量产生。这种将营养物质能量转化为热量的能力，使棕色脂肪细胞（brown adipocytes）区别于更为常见的主要负责能量储存的白色脂肪细胞（white adipocytes）。在小鼠体内，棕色脂肪沉积多以肩胛间大块沉积的形式存在。棕色脂肪细胞呈多室形态，即包含多个被脂质填充的小型液泡，且特异性表达解偶联蛋白1（uncoupling protein 1）。
本研究对年轻健康小鼠的棕色脂肪组织样本的基因表达模式进行了分析。
总体实验设计：将雄性C57Bl6J小鼠饲养于可控环境中，环境温度为22±2℃，采用12小时光照/12小时黑暗的光周期，并饲喂标准维持饲料（Ssniff，德国措斯特）。在小鼠10周龄时通过颈椎脱臼法处死动物。分离棕色脂肪沉积组织，并立即置于液氮中速冻保存。从50mg组织中提取RNA。采用Illumina HiSeq 2000平台进行NGS测序，测序读长为2×100 bp（LGC genomics，德国柏林）。在数据处理与分析环节，使用FASTQC v0.11.8进行质量评估，采用trimmgalore v0.6.4过滤接头序列与短读段。将质控后的干净读段比对至参考基因组（基因组参考联盟小鼠构建版本38 mm10，Genome Reference Consortium Mouse Build 38 mm10），比对工具选用HISAT2 v2.1.0与Bowtie2 v2.3.5.1。使用Cufflinks v.2.2.1对基因表达水平进行定量分析。