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LIVER Publication-derived

Liver Spheroids for Drug Screening

Full SOP with Academic or a plan

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

Authors: 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 Duration: 12 days 4 phases Cell source: 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

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

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Attribution

This SOP was written by Organthis based on the published method in Coll et al., 2018. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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