P12-01 Moderate intake of beer improves nonalcoholic fatty liver disease (NAFLD) in a high fat diet (HFD)-induced mouse model

Purpose Both beer and some of its components, particularly polyphenols and iso-alpha-acids, have proven to be able to attenuate hepatic lipid accumulation or perturbed blood parameters in different rodent models through different putative mechanisms [1, 2, 3]. The current study was carried out within the NUTRATGE project (https://nutrage.it/) and aimed to evaluate the anti-steatotic capacity of beer in an HFD-induced NAFLD mouse model. Methods The beer was characterized for bioactive molecules content and individual phenolic compounds using UHPLC-ESI-MS/MS. In the in vivo study, forty-eight six-weeks-old male mice (C56BL/6) were randomly divided into four groups and supplemented daily during 10 weeks as follows: 1) normal diet (CTR); 2) a CTR diet and 0.14 ml/day beer (CTR+Beer); 3) a HFD (HFD); 4) a HFD and 0.14 ml/day beer (HFD+Beer). Prior to sacrifice, the weight of each animal was recorded, and blood was collected. We quantified liver lipids, performed histopathological evaluation using hematoxylin and eosin staining, and analyzed biomarkers of oxidative stress. Additionally, analysis of gene expression and DNA methylation of hepatic tissue was performed by RNA-Seq and Reduced Representation Bisulfite Sequencing. Results The beer displayed a good content in total phenols (25.01±1.27 mg GAE/100 ml), flavonoids (3.17±0.17 mg CE/100 ml) and flavonols (3.07±0.23 mg QE/100 ml). Among the single phenolic compounds, isoquercetin emerged as the predominant polyphenol (14.68±2.68 mg/100 ml). Compared to CTR, HFD group showed significantly higher levels of AST, ALT, TC, LDL-C, glucose, body weight and liver lipids, indicating the presence of steatosis, confirmed also by histological analysis. In HFD+beer group all the parameters returned to levels similar to those of CTR. All groups exhibited comparable levels of both protein carbonylation and lipid peroxidation in the liver, suggesting that our model represents an early stage of NAFLD with no oxidative stress. Analysis of transcriptomic and CpG methylation profile showed a clear separation between CTR and HFD groups. Beer consumption only partially affected gene expression whereas specifically changed the DNA methylation profile. RNA-Seq revealed 162 differentially expressed genes (DEGs) between CTR and HFD, whose biological function was related to cellular inflammatory processes and regulation of lipid metabolism. Beer consumption ameliorated the HDF effect (CTRvsHFD+ beer, DEGs=43) showing alteration in the inflammatory response but not in the lipid homeostasis. RRBS profile identified 562 (CTRvsHFD), 429 (CTRvsHFD+beer), 469 (CTRvsCTR+beer) and 860 (HFDvsHFD+ beer) differentially methylated cytosines (DMCs). DMCs target genes related to acyl glycerol and lipid biosynthetic process for CTRvsHFD+beer and insulin signaling for CTRvsCTR+beer comparisons. In summary, beer was capable to improve NAFLD likely due to the ability of polyphenols to modulate lipid metabolism.