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LIVER Publication-derived

Ginsenoside Rb1 improves human nonalcoholic fatty liver disease with liver organoids-on-a-chip

Source Wang et al., 2024 · Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences · 10.1016/j.engreg.2024.06.002

👤 Hui Wang, Yue Zhu, Pengcheng Shi, Xiangyang Li, Qingyun Bu, Yachun Li, Xiaoyan You, Guoping Zhao ⏱ 30 days 📋 6 phases 🧫 Human iPSC

Abstract

This protocol describes the generation of hiPSC-derived liver organoids (LOs) on a microfluidic chip platform, modeling free fatty acid (FFA)-induced nonalcoholic fatty liver disease (NAFLD), and evaluating the therapeutic effects of ginsenoside Rb1. The LOs-on-a-chip system recapitulates key pathological features of NAFLD including steatosis, inflammation, and fibrosis, and allows high-throughput assessment of drug efficacy.

Cell source
Human iPSC
Application
Disease modeling and drug screening

Protocol overview

34 steps across 6 phases

Microfluidic chip fabrication and preparation Pre-culture
  1. 1 PDMS mixture preparation
  2. 2 PDMS curing
  3. 3 Layer bonding via oxygen plasma treatment
  4. 4 Chip sterilization option 1: Autoclave
  5. 5 Chip sterilization option 2: UV exposure
  6. 6 Chip surface hydrophobization
HiPSC culture and expansion Days -5 to 0
  1. 1 Matrigel coating plate preparation
  2. 2 Matrigel coating incubation
  3. 3 HiPSC seeding on Matrigel-coated plates
  4. 4 Daily medium changes
  5. 5 HiPSC passage at 80–90% confluence
  6. 6 HiPSC subculture at dilution ratio
  7. 7 Y27632 treatment post-passage
  8. 8 Medium replacement after Y27632 treatment
HiPSC differentiation to liver organoids (Days 0–10) Days 0–10
  1. 1 Initial hiPSC seeding at 1:6 dilution
  2. 2 Definitive endoderm induction (Days 0–5)
  3. 3 Liver-specific differentiation (Days 5–8)
  4. 4 Hepatic progenitor amplification (Days 8–10)
  5. 5 Cell dissociation and harvesting (Day 10)
Liver organoid formation and maturation on chip (Days 10–30) Days 10–30
  1. 1 Cell seeding into microfluidic chip
  2. 2 3D spheroid formation (Day 10–11, overnight)
  3. 3 Perfusion initiation (Days 10–13)
  4. 4 Hepatocyte maturation medium introduction (Day 13)
  5. 5 OSM removal (Day 18)
  6. 6 Physiological LO maturation (Days 18–30)
NAFLD disease modeling with free fatty acids (Days 20–25) Days 20–25
  1. 1 FFA preparation
  2. 2 FFA exposure initiation (Day 20)
  3. 3 Control group preparation
  4. 4 FFA exposure duration (Days 20–25)
Drug treatment with ginsenoside Rb1 and controls (Days 25–30) Days 25–30
  1. 1 Drug treatment initiation (Day 25)
  2. 2 Positive control drug treatment
  3. 3 Control groups during drug treatment
  4. 4 Drug exposure duration (Days 25–30)
  5. 5 Organoid harvest (Day 30)

Full SOP

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Attribution

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

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