Hepatic gene expression in CDAHFD-induced non-alcoholic steatohepatitis model mouse

A choline-deficient amino acid defined high-fat diet (CDAHFD)-induced NASH model is a well-established and reproducible experimental animal model. However, the cause of hepatic fibrogenesis on this model remains unclear. This study aimed to clarify the phenotypic changes and causality between metabolic changes with hepatic fibrogenesis in the CDAHFD-fed mice. To characterize the phenotypic changes in the CDAHFD-fed mouse liver, we conducted transcriptomic analyses (RNA-seq). C57BL/6J male mice were given a choline-deficient L-amino acid defined (0.1% methionine) 60% kcal high-fat diet (CDAHFD; A06071302, Research Diets) for 0, 1, 4, 8, and 12 weeks. The age- and sex-matched acclimatized mice fed a standard chow diet (CRF-1; Oriental Yeast, Japan) were assigned to the control group (0 weeks). Total RNA was extracted from frozen liver samples using the RNeasy Mini kit (Qiagen). After RNA quantification and quality control, libraries were constructed using the TruSeq Stranded mRNA LT Sample Prep kit (Illumina). The NovaSeq 6000 (Illumina) sequence paired-end read length was set to 2 × 100 bp; raw data were processed and checked using the Galaxy system. The filtered data were mapped onto the Gencode reference genome (GRCm38.p6) and annotated using Gencode (M24).

胆碱缺乏氨基酸限定型高脂饮食（choline-deficient amino acid defined high-fat diet，缩写CDAHFD）诱导的非酒精性脂肪性肝炎（non-alcoholic steatohepatitis，NASH）模型是一类成熟且可重复的实验动物模型。然而，该模型中肝纤维化的致病机制仍未明确。本研究旨在明确CDAHFD喂养小鼠体内代谢变化与肝纤维化之间的因果关联及表型变化。为解析CDAHFD喂养小鼠肝脏的表型特征变化，本研究开展了转录组测序（RNA-seq）。选取C57BL/6J雄性小鼠，分别喂食含0.1%甲硫氨酸的胆碱缺乏L-氨基酸限定型60%热量高脂饮食（CDAHFD；货号A06071302，Research Diets公司），并分别在造模0、1、4、8及12周时取样；同时设置对照组，喂食标准维持饲料（CRF-1；日本Oriental Yeast公司），其取样时间点对应造模0周。采用RNeasy Mini试剂盒（Qiagen公司）从冻存肝脏样本中提取总RNA。完成RNA定量与质量控制后，使用TruSeq Stranded mRNA LT样本制备试剂盒（Illumina公司）构建测序文库。使用NovaSeq 6000测序仪（Illumina公司）进行双端测序，测序读长设置为2×100 bp；原始数据通过Galaxy系统完成处理与质控。将过滤后的测序数据比对至Gencode参考基因组（GRCm38.p6），并通过Gencode（M24）完成基因注释。