*2.7. RNA-Seq Analysis*

Pre-processing and normalization. The count matrix was derived from FASTQ files of wild-type, CCl4 plus olive oil and pure olive oil samples using the web application BioJupies [23]. The samples were normalized using the R package edgeR (version 3.25.8) [24].

Differential gene expression analysis. Differential gene expression analysis was performed with the R package limma (version 3.39.18) [25]. To extract the effect of chronic CCl4 intoxication, we computed contrasts comparing the treated samples (CCl4 plus olive oil) versus matched olive oil samples. While there are matched oil samples for month 2 and 12, there were no oil samples for month 6 available. As the oil effect was relatively constant across time we imputed oil expression values for month 6 by taking the arithmetic mean of month 2 and month 12. A gene was considered as differentially expressed with abs(logFC) ≥ 1.5 and FDR ≤ 0.05.
