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LIVER Publication-derived

Generation of Hepatic Stellate Cells from Human Pluripotent Stem Cells

Source Coll et al., 2018 · Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Barcelona, Spain · 10.1016/j.stem.2018.05.027

👤 Mar Coll, Luis Perea, Ruben Boon, Sofia B. Leite, Julia Vallverdú, Inge Mannaerts, Ayla Smout, Adil El Taghdouini, Delia Blaya, Daniel Rodrigo-Torres, Isabel Graupera, Beatriz Aguilar-Bravo, Christophe Chesne, Mustapha Najimi, Etienne Sokal, Juan José Lozano, Leo A. van Grunsven, Catherine M. Verfaillie, Pau Sancho-Bru ⏱ 12 days 📋 4 phases 🧫 Human iPSC

Abstract

This protocol describes the differentiation of human pluripotent stem cells (iPSCs and hESCs) into hepatic stellate cell-like cells (iPSC-HSCs) over 12 days using sequential growth factor stimulation. iPSC-HSCs closely resemble primary human HSCs and can be assembled into 3D spheroids with HepaRG hepatocytes to model liver fibrosis and assess drug-induced hepatotoxicity.

Cell source
Human iPSC
Application
Disease modeling, Drug screening, Fibrosis modeling

Protocol overview

13 steps across 4 phases

Mesodermal Progenitor Induction Day 0–4
  1. 1 Prepare culture plates and cell seeding
  2. 2 Replace medium with HSC differentiation medium
  3. 3 Add BMP4 to induce mesodermal commitment
  4. 4 Continue BMP4 treatment and change medium every 48 hours
Liver Mesenchymal Submesothelial Phenotype Induction Day 4–8
  1. 1 Add FGF1 and FGF3 and reduce BMP4
  2. 2 Continue medium changes with BMP4, FGF1, and FGF3
  3. 3 Add retinol and palmitic acid from day 6
HSC Phenotype Maturation Day 8–12
  1. 1 Discontinue BMP4 and FGF signaling
  2. 2 Continue retinol and palmitic acid treatment to day 12
  3. 3 Harvest differentiated iPSC-HSCs on day 12
Optional: iPSC-HSC Expansion and Characterization Day 12 onwards
  1. 1 Plate differentiated cells for expansion (Optional)
  2. 2 Culture iPSC-HSCs in expansion medium
  3. 3 Assess HSC phenotype by flow cytometry (Optional)

Full SOP

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Attribution

This SOP was authored by Organthis based on the published method in Coll et al., 2018. The originating laboratory holds no rights in this SOP and has not endorsed it unless marked Verified.

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