Modelling Liver Molecular Fibrogenesis In Human Liver Slices Culture.

Background: To study the first steps of liver molecular fibrogenesis in our ex vivo model of human liver slices culture. Methods: Human fibrotic liver samples (F0 to F4 fibrosis stage according to the METAVIR score) were collected after liver resection. Human liver slices of 350 µm (2.7x106cells per slice) were cultivated for up to 21 days. Fibrotic liver slices F2 to F4 (FLS) were infected (or not) with HCVcc supernatant infection [Con1/C3 (genotype1b)] (MOI = 0.1) (INF FLS). F0-F1 HLS (HLS) were used as controls. At day 0, either ursodeoxycholic acid (UDCA, only choleretic and hepatoprotective properties) and/or atocopherol (Toco, anti-oxydant properties which could reduce fibrosis progression) were added to standard of care concentrations on HLS and FLS. The following fibrosis markers expression were assayed in HLS, in FLS and in INF FLS, [tumor growth factor b (TGFβ), Heat shock protein 47 (Hsp47), Alpha smooth muscle actin (α-Sma), Procollagen1 A1 (Procl1A1), Matrix metalloproteinases 2, 9 (MMP-2, 9), Vascular Endothelial Growth Factor (VEGF)] and checked by RT-qPCR and the triglyceride production by ELISA assays. Results: The TGFβ, Hsp47,α-Sma, Procl1A1, MMP-2, MMP-9, VEGF expression and the triglyceride production increased significantly in both HLS and FLS but to a higher extent in FLS as compared to HLS, with a triglyceride production higher [~x 3.2] in FLS. HCV infection increased significantly the fibrotic markers expression and triglyceride production in non-infected (NI) FLS and INF FLS to a higher extent in INF FLS as compared to controls. The induced molecular fibrogenesis significantly increased, particularly true for the advanced stage of fibrosis, including cirrhosis. Under a-tocopherol exposure, a significant reduction of TGF-β RNA expression was observed in NINF and INF FLS but only for the stage F4. UDCA did not impact TGFβ RNA expression in NINF F0 to F4 liver slices, but significantly reduces its expression in F2F3F4 infected liver slices. Combination of UDCA and Toco treatments did not result in an additive effect and only reduced the TGFβ RNA expression in F2-F4 INF HLS but not in F2-F4 NI HLS. Conclusion: Our ex vivo model of human liver slices culture, supporting hepatocyte-specific gene expression for 21 days, provides an easy tool for studying human molecular liver fibrogenesis which could accelerate the evaluation of the potency of new anti-fibrotic therapies, alone or in combination.