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 4

Glucose utilization and synthesis is altered in the human liver lipotoxic system.

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Glucose utilization and synthesis is altered in the human liver lipotoxi...
(A) Baseline levels of secreted glucose were measured from hepatocytes exposed to the healthy or lipotoxic milieu for 10 days and represented as fold change relative to healthy controls. n ≥ 12 experiments, 5 donors. (B) Hepatocyte expression of genes of the glycolysis and gluconeogenesis pathways are represented as log2-fold change of lipotoxic vs. healthy milieu (red = upregulation, blue = downregulation). n = 6 experiments, 3 donors. (C) Hepatocytes exposed to the healthy or lipotoxic milieu for 10 days were serum starved with or without 100 nM insulin for 3 hours and secreted glucose was measured and represented as fold change relative to baseline healthy controls. n ≥ 9 experiments, 5 donors. (D) Hepatocytes exposed to the healthy or lipotoxic milieu for 10 days were serum starved and then stimulated with or without 10 nM insulin for 10 minutes to measure Akt phosphorylation. Levels of phosphorylated Akt induced by insulin are represented as fold change relative to baseline phosphorylated Akt in the absence of insulin challenge. n = 5 experiments, 3 donors. *P < 0.05, **P < 0.01, Student’s 2-tailed t test. NS, not significant.