Development of an in vitro human liver system for interrogating nonalcoholic steatohepatitis
Ryan E. Feaver, … , Brian R. Wamhoff, Ajit Dash
Ryan E. Feaver, … , Brian R. Wamhoff, Ajit Dash
Published December 8, 2016
Citation Information: JCI Insight. 2016;1(20):e90954. https://doi.org/10.1172/jci.insight.90954.
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Technical Advance Hepatology Therapeutics

Development of an in vitro human liver system for interrogating nonalcoholic steatohepatitis

Abstract

A barrier to drug development for nonalcoholic steatohepatitis (NASH) is the absence of translational preclinical human-relevant systems. An in vitro liver model was engineered to incorporate hepatic sinusoidal flow, transport, and lipotoxic stress risk factors (glucose, insulin, free fatty acids) with cocultured primary human hepatocytes, hepatic stellate cells (HSCs), and macrophages. Transcriptomic, lipidomic, and functional endpoints were evaluated and compared with clinical data from NASH patient biopsies. The lipotoxic milieu promoted hepatocyte lipid accumulation (4-fold increase, P < 0.01) and a lipidomics signature similar to NASH biopsies. Hepatocyte glucose output increased with decreased insulin sensitivity. These changes were accompanied by increased inflammatory analyte secretion (e.g., IL-6, IL-8, alanine aminotransferase). Fibrogenic activation markers increased with lipotoxic conditions, including secreted TGF-β (>5-fold increase, P < 0.05), extracellular matrix gene expression, and HSC activation. Significant pathway correlation existed between this in vitro model and human biopsies. Consistent with clinical trial data, 0.5 μM obeticholic acid in this model promoted a healthy lipidomic signature, reduced inflammatory and fibrotic secreted factors, but also increased ApoB secretion, suggesting a potential adverse effect on lipoprotein metabolism. Lipotoxic stress activates similar biological signatures observed in NASH patients in this system, which may be relevant for interrogating novel therapeutic approaches to treat NASH.

Authors

Ryan E. Feaver, Banumathi K. Cole, Mark J. Lawson, Stephen A. Hoang, Svetlana Marukian, Brett R. Blackman, Robert A. Figler, Arun J. Sanyal, Brian R. Wamhoff, Ajit Dash

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Figure 6

Evidence for extracellular matrix remodeling in the human liver lipotoxic system.

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Evidence for extracellular matrix remodeling in the human liver lipotoxi...
(A, C, and D) Secreted TGF-β (A), osteopontin (OPN) (C), and procollagen 1α1 (D) were measured in the media effluent from devices at the indicated days. n ≥ 4 experiments, 4 donors. (B) Hepatocyte expression of genes of the collagen formation pathway are represented as log2-fold change of lipotoxic vs. healthy milieu (red = upregulation, blue = downregulation). n = 6 experiments, 3 donors. (E) Representative photomicrographs (original magnification, ×20) of nonparenchymal cells (NPCs) are shown. Macrophages (CD68+, red), nuclei (DAPI+, blue), and hepatic stellate cells (smooth muscle α-actin+ (SMAA+), green). Scale bars: 100 μm. SMAA staining intensity from immunofluorescent NPCs images were quantified and represented as fold change relative to healthy controls. n ≥ 4 experiments, 3 donors. Triangles indicate samples that were below the lower limit of quantification. *P < 0.05, **P < 0.01, Student’s 2-tailed t test.