AUTHORS

Sophie Cazanave1, Alexei Podtelezhnikov2, Mulugeta Seneshaw1, Robert Vincent1, Amon Asgharpour1, Kristian Jensen2,Keith Q. Tanis2, Andrea L. Webber2, Liangsu Wang2, Pierre Bedossa3, Faridoddin Mirshahi1 and Arun J. Sanyal1.

1 Division of Gastroenterology, Hepatology and Nutrition, Department of Internal Medicine, Virginia Commonwealth University School of Medicine, Richmond, VA, USA, 2 Merck Research Laboratories, Kenilworth, NJ, USA, 3 Department of Pathology, Hôpital Beaujon, Clichy, France.

BACKGROUND

  • Nonalcoholic fatty liver disease (NAFLD), especially its aggressive form nonalcoholic steatohepatitis (NASH), can progress to cirrhosis.
  • Changes in the hepatic transcriptome during disease progression have not been fully characterized and limit the development of precision medicine-base therapeutics.
  • We have recently Developed a diet-induced animal model of NAFLD (DIAMOND model) that closely mimics progressive human NAFLD in terms of physiological, metabolic biochemical and histologic parameter.

To gain further insights into the molecular progression of NAFLD, we conduct an in-depth longitudinal transcriptome study of DIAMOND mice at 8, 24 and 52 weeks of diet.

AIM

To model the molecular progression of NAFLD

MATERIALS AND METHODS

  • Animals and Diets: Male C57Bl/6J/129Sl/SvlmJ mouse strain (B6/129) were fed a high-fat diet with 42 energy% from fat (Western Diet, WD, Harlan TD.88137) and with high fructose-glucose solution (SW, 23.1g/l d-fructose + 18.9 g/l d-glucose) or a standard chow diet for 8, 24 or 52 weeks (n=4-5/group). At 52 weeks, 90% of WD SW-fed mice develop tumors.
  • Microarray: Total RNA of reported integrity numbers (RIN) >8 were extracted. Mouse whole-genome profiling was performed using the Illumina mouse WG6 Expression BeadChip kits (Mouse WG-6 2.0). Following quantile normalization, 2 groups differences were evaluated by 2-way ANOVA. Biological enrichment among differentially expressed genes
    (false discovery rate <0.1) was evaluated using Ingenuity
    Pathway Analysis (IPA).

TABLE 1: TOP CANONICAL PATHWAYS

FIGURE 1: LIPID METABOLISM

Heat maps resulting from hierarchical clustering for genes implicated in (A) Fatty acid Oxidation or (B) Sterol Biosynthesis pathways with a fold-change greater than ±1.5 from WD SW liver samples as compared to CD NW or from liver tumors at 52 wks as compared to WD SW 52 wks with a false discovery rate (FDR) < 0.1. Whiskers show the 5th to 95th percentile with outliers (red cross). Red dashed lines show ±1.5 fold-change, and blue dash line shows no change compared with CD NW at 8, 24 and 52 wks or compared to WD SW 52 wks for liver tumors.

FIGURE 2: CELL DEATH/INFLAMMATION

Heat maps resulting from hierarchical clustering for genes implicated in (A) Unfolded Response Protein, (B) Apoptosis, (C) Fcg-mediated Phagocytosis and (D) Toll-like Receptor pathways with a fold-change greater than ±1.5 from WD SW liver samples as compared to CD NW or from liver tumors at 52 wks as compared to WD SW 52 wks with a false discovery rate (FDR) < 0.1.

FIGURE 3: FIBROSIS/CELL
PROLIFERATION/ONCOGENIC PATHWAYS

heat maps resulting from hierarchical clustering for genes implicated in (A) Hepatic Fibrosis, (B) Cell Proliferation or (C) Cancer pathways with a fold-change greater than ±1.5 from WD SW liver samples as compared to CD NW or from liver tumors at 52 wks as compared to WD SW 52 wks with a false discovery rate (FDR) < 0.1.

CONCLUSION

The molecular progression of NAFLD involves:

  • First, a metabolic perturbation which triggers subsequent cell
    stress and inflammation driving cell death and turnover.
  • Over time, inflammation and fibrogenic pathways become
    dominant while in advanced disease an inflammatory-oncogenic
    profile dominates.

REFERENCES

  1. Asgharpour A, Cazanave SC et al., J Hepatol. 2016 Sep;65(3):579-88. A
    diet-induced animal model of non-alcoholic fatty liver disease and hepatocellular
    cancer.