Skip to content
Back to browse
LIVER Publication-derived

Liver Organoids for Drug Screening (2023)

Full SOP with Academic or a plan

Source Wu et al., 2023 · Shanghai Institute of Nutrition and Health, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences · 10.1186/s13619-022-00148-1

Authors: Xiaoshan Wu, Dacheng Jiang, Yi Yang, Shuang Li, Qiurong Ding Duration: 25 days 10 phases Cell source: Human iPSC

Abstract

This protocol describes the generation of functional human liver organoids (HLOs) from human induced pluripotent stem cells (hiPSCs) containing multiple hepatic cell types including hepatocytes, stellate cells, and Kupffer cells. HLOs are used to model diverse phenotypes of drug-induced liver injury (steatosis, fibrosis, and immune responses) and to enable high-throughput screening for anti-fibrotic compounds.

Cell source
Human iPSC
Application
Disease modeling and drug screening

Protocol overview

38 steps across 10 phases

Phase 1 · Foregut differentiation from hiPSCs Day 0-6
  1. a Cell seeding and plate preparation
  2. b Day 1 medium change
  3. c Day 2 medium change
  4. d Day 3 medium change
  5. e Day 4-6 foregut specification medium
Phase 2 · HLO generation - standard protocol (HLO) Day 6-20
  1. a Harvest foregut cells
  2. b Embed foregut cells in Matrigel with RA medium
  3. c Medium switch to liver maturation medium
  4. d Culture until day 20
Phase 3 · HLO generation - 5-factor expansion protocol (5F-HLO) Day 6-25
  1. a Harvest and dissociate foregut cells
  2. b Embed cells in Matrigel with 5-factor expansion medium
  3. c Switch to RA medium
  4. d Re-embed organoids in liver maturation medium
  5. e Final maturation culture
Phase 4 · Drug treatment - APAP (Acetaminophen) 7 days
  1. a Prepare APAP treatment concentrations
  2. b Incubate HLOs with APAP
  3. c Assess toxicity readouts
Phase 5 · Drug treatment - FIAU (Fialuridine) 10 days
  1. a Prepare FIAU treatment concentrations
  2. b Incubate HLOs with FIAU
  3. c Assess steatosis and mitochondrial function
Phase 6 · Drug treatment - MTX (Methotrexate) 7 days
  1. a Prepare MTX treatment concentrations
  2. b Incubate HLOs with MTX
  3. c Assess steatosis, fibrosis, and gene expression
Phase 7 · Drug treatment - TAK-875 (Fasiglifam) 7 days
  1. a Prepare TAK-875 treatment concentrations
  2. b Incubate HLOs with TAK-875
  3. c Assess oxidative stress and immune response
Phase 8 · Fibrosis induction - TGFβ or LPS treatment 3 days
  1. a Prepare TGFβ or LPS treatment solutions
  2. b Incubate HLOs with fibrogenic factors
  3. c Assess fibrosis and collect samples
Phase 9 · Anti-fibrotic drug screening - high-throughput 3 days (TGFβ induction + compound treatment)
  1. a Prepare compound library and treatment solutions
  2. b Incubate HLOs with compounds and TGFβ
  3. c Fix and stain organoids for Collagen I
  4. d High-content image acquisition and analysis
  5. e Candidate compound validation
Phase 10 · Validation of anti-fibrotic compounds in MTX and LPS models 9 days (MTX); 3 days (LPS)
  1. a Prepare MTX + compound treatment
  2. b Prepare LPS + compound treatment
  3. c Assess compound efficacy in MTX model
  4. d Assess compound efficacy in LPS model

Full SOP

Full SOPs need an account and a plan

Create an account, then get free Academic access (for verified academics) or choose a plan. Plans include step-by-step procedures, reagent concentrations, QC checklists and batch record templates.

Create an account

Already registered? Log in

Attribution

This SOP was written by Organthis based on the published method in Wu et al., 2023. Once the originating laboratory has confirmed it, the SOP is marked Verified.

Something wrong with this entry? Report an issue with this protocol

Need a commercial licence?
Use this protocol in your therapeutic or diagnostic pipeline.
Request Commercial Licence