Combined obeticholic acid and elafibranor treatment promotes additive liver histological improvements in a diet-induced ob/ob mouse model of biopsy-confirmed NASH

Figures

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  Figure 1. Combined OCA and ELA treatment improves metabolic markers in AMLN ob/ob-NASH mice with biopsy-confirmed liver pathology. (A) Body weight (g), (B) Body weight gain relative to treatment start (day 0), (C) Whole-body composition, (D) Terminal liver weight; (E) Plasma total triglycerides (TG); (F) Plasma total cholesterol (TC); (G) Plasma alanine aminotransferase (ALT); (H) Plasma aspartate aminotransferase (AST).

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  Figure 2. Combined OCA and ELA treatment promotes additive effects on quantitative liver histopathology in AMLN ob/ob-NASH mice. Liver histomorphometric data are indicated before (baseline) and after treatment. (A) Fractional area (%) of fat; (B) Galectin-3; (C) Collagen 1a1. Data are expressed as % of total parenchymal area (subtraction of fat area). See Table 1 for corresponding numerical data and statistical evaluation.

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  Figure 3. Multivariate visualization of the anti-steatotic, anti-inflammatory, and anti-fibrotic effects of combined OCA and ELA treatment in AMLN ob/ob-NASH mice. Terminal liver histomorphometric data of fractional area (%) of collagen 1a1 plotted against corresponding fractional area (%) of steatosis (panel A, r2 = 0.54) and galectin-3 (panel B, r2 = 0.60), respectively. Linear regression lines are indicated.

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  Figure 4. Representative histological effects of OCA and ELA monotherapy vs. combined treatments in AMLN ob/ob-NASH mice. Representative photomicrographs depicting the anti-steatotic (HE staining), anti-inflammatory (Galectin-3 immunostaining), and anti-fibrotic (Collagen 1a1 immunostaining) effects of OCA and ELA monotherapy vs. combined treatment in AMLN ob/ob-NASH mice.

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  Figure 5. OCA and ELA exert additive effects on hepatic global gene expression and NAFLD/fibrosis-associated candidate genes in AMLN ob/ob-NASH mice. (A) Venn diagram depicting the total number of shared and separate differentially expressed genes (DEGs) in AMLN ob/ob-NASH mice receiving OCA (30 mg/kg), ELA (3 mg/kg), or combined OCA (30 mg/kg) + ELA (3 mg/kg) treatment as compared to vehicle controls; (B) Relative gene expression levels (z-scores) of differentially expressed candidate genes associated with NASH and fibrosis (see Supplementary Table S1 for complete list of candidate genes); (C) Expression levels (RPKM values) of genes represented in most significantly regulated pathways associated with lipid metabolism/FXR signaling, monocyte recruitment/inflammation signaling, and stellate cell activation/extracellular matrix (ECM) organization. *p < 0.05, **p < 0.01, ***p < 0.001 (vs. vehicle); #p < 0.05, ##p < 0.01, ###p < 0.01 (vs. OCA alone); ¤p < 0.01, ¤¤p < 0.01, ¤¤¤p < 0.001 (vs. Elafibranor alone).

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  Figure 6. Global analysis of pathway perturbations following combined OCA and ELA treatment in AMLN ob/ob-NASH mice. (A) Venn diagram depicting the total number of shared and separate Reactome pathway perturbations in ob/ob-NASH mice receiving OCA (30 mg/kg), ELA (3 mg/kg), or combined OCA (30 mg/kg) + ELA (3 mg/kg) treatment as compared to vehicle controls; (B) Significantly regulated Reactome pathways grouped according to biological pathway. Further resolution of selected Reactome pathways, including (C) Innate immune system, (D) Adaptive immune system, and (C) Metabolism of lipids. Upregulated (red color gradient) and downregulated (blue color gradient) gene expression in individual Reactome pathways are ranked according to statistical significance level.