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LIVER Publication-derived

A High-Throughput Microphysiological Liver Chip System to Model Drug-Induced Liver Injury Using Human Liver Organoids

Source Meyer et al., 2024 · University of Michigan, Ann Arbor · 10.1016/j.gastha.2024.08.004

👤 Sophia R. Meyer, Charles J. Zhang, Max A. Garcia, Megan C. Procario, Sanghee Yoo, Amber L. Jolly, Sumin Kim, Jiho Kim, Kyusuk Baek, Roland D. Kersten, Robert J. Fontana, Jonathan Z. Sexton ⏱ 55 days 📋 9 phases 🧫 Human iPSC

Abstract

This protocol describes the differentiation of human iPSCs into liver organoids (HLOs) and their culture on Curio Barrier Liver Chips (Curiochips) to model drug-induced liver injury. The system maintains hepatocyte function and CYP450 metabolic activity for up to 28 days and enables rapid detection of hepatotoxicity at physiologically relevant drug concentrations, offering a high-throughput platform for DILI risk assessment in drug development.

Cell source
Human iPSC
Application
Disease modeling; Drug-induced liver injury (DILI) screening and risk prediction

Protocol overview

35 steps across 9 phases

Human iPSC Culture and Definitive Endoderm (DE) Generation Days 0–6
  1. 1 Seed iPSCs on Matrigel-coated plates
  2. 2 Treat iPSCs with Activin A for 3 days
  3. 3 Treat iPSCs with FGF4 for 3 additional days
DE Spheroid Maturation to Liver Organoids (Ex Situ) Days 7–26
  1. 1 Embed DE spheroids in Matrigel droplets
  2. 2 Treat embedded spheroids with retinoic acid for 4 days
  3. 3 Culture spheroids in hepatocyte growth media for 16 days
HLO Recovery from Matrigel and Cell Dissociation Day 26–27
  1. 1 Wash organoids to remove Matrigel
  2. 2 Dissociate organoids with trypsin
  3. 3 Count cells for downstream seeding
Curiochip Preparation and HLO Patterning Day 27
  1. 1 Prepare hydrogel cell suspension
  2. 2 Load cell-containing hydrogel into middle channel
  3. 3 Load acellular hydrogel into side 1 channel
  4. 4 Load culture medium into side 2 channel
  5. 5 Establish media gradient for osmotically driven flow
Curiochip Culture and Functional Assessment (Days 1–28) Days 27–55 (28 days on chip)
  1. 1 Culture HLO-seeded Curiochips with routine media changes
  2. 2 Collect media for albumin secretion assessment
  3. 3 Extract RNA for CYP450 gene expression analysis
CYP450-Mediated Substrate Metabolism Assessment Day 7 (or as specified)
  1. 1 Prepare substrate-containing media
  2. 2 Incubate Curiochips with substrates
  3. 3 Collect media for substrate analysis
  4. 4 Stop reactions with cold methanol
  5. 5 Analyze samples by LC/MS/MS
Drug-Induced Liver Injury (DILI) Assessment Days 7–21 of culture (7-day drug treatment)
  1. 1 Pre-treat Curiochips and establish baseline
  2. 2 Treat Curiochips with DILI-causing agents
  3. 3 Collect media for ALT activity and albumin quantification
  4. 4 Measure alanine transaminase (ALT) activity
In Situ Differentiation (Alternative Protocol) Days 0–28
  1. 1 Prepare definitive endoderm spheroids on day 7
  2. 2 Load acellular hydrogel and culture medium
  3. 3 Complete hepatocyte maturation on chip
  4. 4 Assess in situ differentiation and perform DILI studies
Immunofluorescence Imaging and Phenotypic Assessment Days 7–28 (as specified)
  1. 1 Prepare Curiochips for imaging
  2. 2 Stain for hepatocyte and stellate cell markers
  3. 3 Stain for neutral lipid droplets (DILI injury marker)
  4. 4 Acquire high-content confocal images
  5. 5 Quantify biomarker intensity

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Meyer et al., 2024. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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