Transcription profiling of mouse liver from wild type and Wrn mutant animals with or without vitamin C treatment

Werner syndrome (WS) is a rare disorder characterized by the premature onset of a number of age-related diseases. The gene responsible for WS is believed to be involved in different aspects of transcription, replication, and/or DNA repair. We generated a mouse model with a deletion in the helicase domain of the murine WRN homologue that recapitulates most of the WS phenotypes including an abnormal hyaluronic acid excretion, higher reactive oxygen species (ROS) levels, increased genomic instability and cancer incidence resulting in a 10-15% decreased life span expectancy. In addition, WS patients and Wrn mutant mice show hallmarks of a metabolic syndrome including premature visceral obesity, hypertriglyceridemia, insulin-resistant diabetes type 2 and associated cardiovascular diseases. In this study, we compared the expression profile of liver tissues from 9 months old Wrn mutant and wild type animals. Gene set enrichment analysis of the microarray data indicated that Wrn mutant mice exhibited down-regulation of genes normally decreased in several transgenic mouse models of hepatoma and during caloric restriction. Wrn mutant mice also altered the expression of genes involved in inflammation as well as in glutathione and xenobiotic metabolisms. These results indicate that Wrn mutant mice respond to the observed oxidative stress by altering the necessary pathways to survive. Vitamin C supplementation rescued the life span expectancy of Wrn mutant mice and reversed several age-related abnormalities in adipose, cardiac, and liver tissues, genomic integrity and inflammatory status. Finally, gene set enrichment analyses revealed that vitamin C decreased genes normally up regulated in human WS fibroblasts and cancers and it increased genes involved in tissue injury response and adipocyte dedifferentiation in obese mice. Experiment Overall Design: Microarray analyses were performed on the liver tissues of 9 months old mice. Four independent biological replicates of this experiment (wild type vs Wrn mutant or wild type vs vitamin C treated mutant mice) were carried out with a dye swap on two replicates of each genotype.沃纳综合征（Werner syndrome, WS）是一种罕见的遗传病，其特征为多种年龄相关性疾病提前发作。目前认为，导致沃纳综合征的致病基因参与转录、复制及/或DNA修复等多个生物学过程。本研究构建了一种小鼠模型，该模型在小鼠WRN同源基因的解旋酶结构域存在缺失，可重现沃纳综合征的多数表型，包括透明质酸排泄异常、活性氧（reactive oxygen species, ROS）水平升高、基因组不稳定性增加以及癌症发生率上升，最终导致寿命缩短10%~15%。此外，沃纳综合征患者与Wrn突变小鼠均表现出代谢综合征的典型特征，包括早发性内脏型肥胖、高甘油三酯血症、胰岛素抵抗型2型糖尿病以及相关心血管疾病。本研究对比了9月龄Wrn突变型与野生型小鼠的肝脏组织基因表达谱。对微阵列（microarray）数据进行基因集富集分析（gene set enrichment analysis）后发现，Wrn突变小鼠体内，在多种肝癌转基因小鼠模型及热量限制状态下通常发生下调的基因，其表达水平进一步降低。此外，Wrn突变小鼠还改变了与炎症、谷胱甘肽代谢及外源性物质代谢相关的基因的表达水平。上述结果表明，Wrn突变小鼠通过调控必要的生存相关通路，以应对观测到的氧化应激。补充维生素C可恢复Wrn突变小鼠的预期寿命，并逆转脂肪、心脏及肝脏组织的多种年龄相关性异常，改善基因组完整性与炎症状态。最后，基因集富集分析显示，维生素C可下调人类沃纳综合征成纤维细胞及癌症组织中通常呈高表达的基因，同时上调肥胖小鼠体内与组织损伤应答及脂肪细胞去分化相关的基因。实验整体设计：本研究对9月龄小鼠的肝脏组织开展微阵列分析。本实验共设置4组独立生物学重复（野生型 vs Wrn突变型，以及野生型 vs 维生素C处理的突变型小鼠），针对每种基因型的2个重复样本进行了染料互换实验。