Precision-cut liver slice (PCLS) is a model that allows preservation of animal study the liver lobule architecture by maintaining cell diversity and interactions. PCLSs prepared from fasted mice were incubated with [14C]-palmitate to measure 14CO2 release during incubation. By this method, we found that the livers of DEF mice presented lower fatty acid oxidation compared to those of CT mice (Figure 2B). Incubation of PCLSs from fed CT and DEF mice with [14C]-acetate or [14C]-palmitate showed a 40% increase in TG synthesis and FA-esterification into TG in DEF mice compared to CT mice (Figure 2C, D). Figure 2 n-3 PUFA depletion leads to decreased hepatic fatty acid oxidation and increased TG secretion and synthesis.

Microarray analysis confirms a metabolic shift in favour of fatty acid and cholesterol synthesis at the expense of fatty acid oxidation in the livers of n-3 PUFA depleted mice The gene expression profiles in the livers of CT and DEF mice were analysed by microarray in both the fasted and fed state. Increased expression of all of the enzymes involved in fatty acid synthesis and desaturation was observed in DEF mice compared to CT mice in the fed and fasted states (Figure 3A and Table S2). The statistical significance of this observation was confirmed by analysing gene ontology with the bioinformatics tool DAVID (Table S3). DEF mice exhibited a lower expression of several markers of fatty acid oxidation, including PPAR�� and its target genes (Figure 3A and Table S2). The bioinformatics evaluation of transcription factor target genes among the list of genes regulated using TFactS [19] supported the inhibition of the PPAR�� pathway (Table S4).

Figure 3 n-3 PUFA-depleted mice exhibited higher mRNA content of enzymes involved in fatty acid and cholesterol synthesis. In agreement with the physiological data, phospholipid transfer protein (PLTP) and Apolipoprotein B (Apo B), two factors involved in fatty acid secretion, were increased in DEF mice when compared to CT mice (Table S2). Increased expression of several enzymes involved in cholesterol synthesis (from HMGCoAs to Dhcr7) and esterification for hepatic secretion (ACAT2) also occurred in fed and fasted DEF mice compared to CT mice (Figure 3B and Table S2). With regard to bile acid metabolism, a higher expression of Cyp7a1 and a lower expression of Cyp7b1 suggest a stimulation of the classic bile acid synthesis pathway AV-951 at the expense of the alternative pathway through n-3 PUFA depletion (Figure 3B and Table S2). SREBP-1c is involved in the metabolic alterations occurring in the livers of n-3 PUFA depleted mice TFactS analysis of microarray results revealed activation of SREBPs (Table S4) but did not distinguish between SREBP-2, SREBP-1c and SREBP-1a.