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LIVER Publication-derived

Generation of expandable human pluripotent stem cell-derived hepatocyte-like liver organoids

Source Mun et al., 2019 · Korea Research Institute of Bioscience and Biotechnology (KRIBB) · 10.1016/j.jhep.2019.06.030

👤 Seon Ju Mun, Jae-Sung Ryu, Mi-Ok Lee, Ye Seul Son, Soo Jin Oh, Hyun-Soo Cho, Mi-Young Son, Dae-Soo Kim, Su Jung Kim, Hyun Ju Yoo, Ho-Joon Lee, Janghwan Kim, Cho-Rok Jung, Kyung-Sook Chung, Myung Jin Son ⏱ 25 days 📋 6 phases 🧫 Human iPSC, Human ESC

Abstract

This protocol describes the generation of expandable, functionally mature human hepatocyte-like liver organoids from pluripotent stem cells (iPSCs and hESCs). The organoids exhibit self-renewal capacity over long-term culture (>24 passages), maintain mature hepatic functions including drug metabolism and serum protein production, and can be used for disease modeling (hepatic steatosis), drug toxicity prediction, and drug efficacy screening.

Cell source
Human iPSC, Human ESC
Application
Disease modeling, Drug screening, Toxicity prediction

Protocol overview

16 steps across 6 phases

Definitive Endoderm Differentiation Day 0–3
  1. 1 Seed PSCs on Matrigel-coated dishes
  2. 2 Exchange medium to definitive endoderm induction medium
  3. 3 Culture in definitive endoderm medium
Hepatic Endoderm Differentiation Day 9–13
  1. 1 Exchange medium to hepatic endoderm induction medium
  2. 2 Culture under hypoxia
Hepatic Maturation Day 13–25
  1. 1 Exchange medium to hepatic maturation medium
  2. 2 Culture under hypoxia
  3. 3 Continue culture under normoxia
3D Organoid Formation and Embedding Day 22–25
  1. 1 Collect floating cysts
  2. 2 Embed organoids in Matrigel dome
Organoid Maintenance and Routine Culture Day 25 onwards
  1. 1 Routine medium replacement
  2. 2 Mechanical passaging
  3. 3 Alternative: Chemical passaging
  4. 4 Cryopreservation
Optional: Further Differentiation Protocol (Expansion and Differentiation Media) Variable (2–3 days EM, then 6 days DM)
  1. 1 Switch to expansion medium
  2. 2 Switch to differentiation medium

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Mun et al., 2019. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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