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SKIN Publication-derived

Atopic Dermatitis Skin Organoids

Full SOP with Academic or a plan

Source Jung et al., 2022 · Adult Stem Cell Research Center and Research Institute for Veterinary Science, College of Veterinary Medicine, Seoul National University · 10.1016/j.isci.2022.105150

Authors: Song-yi Jung, Hyun Ju You, MinJi Kim, GwangPyo Ko, Seunghee Lee, Kyung-Sun Kang Duration: 125 days 6 phases Cell source: Human iPSC

Abstract

This protocol describes the generation of air-liquid interface (ALI)-cultured human skin organoids from iPSCs with Wnt signaling activation to model atopic dermatitis. The organoids feature a stratified squamous epithelium resembling adult human skin and are used to study Staphylococcus aureus colonization and infection, demonstrating direct causality between S. aureus and atopic dermatitis pathologies. The model also evaluates the protective effect of commensal Cutibacterium acnes bacteria.

Cell source
Human iPSC
Application
Disease modeling (atopic dermatitis)

Protocol overview

22 steps across 6 phases

Phase 1 · Embryoid body (EB) formation and initial differentiation Day 0-6
  1. a hiPSC dissociation and EB formation
  2. b Y-27632 supplementation for EB formation
  3. c Transfer EBs to differentiation medium
  4. d Wnt activation and CNCC induction
Phase 2 · Skin organoid maturation and self-assembly Day 11-85
  1. a Transition to skin maturation medium
  2. b Transfer to low-attachment 6-well plates
  3. c Orbital shaking culture
  4. d Regular medium replacement
Phase 3 · Air-liquid interface (ALI) culture setup and initiation Day 85-119
  1. a Prepare collagen-coated Transwell inserts
  2. b Add basal culture medium to Transwell wells
  3. c Cut organoids into quarters and place on Transwell
  4. d Initial ALI culture in humidified incubator
Phase 4 · ALI-skin organoid maturation under dry conditions Day 119-125
  1. a Transfer to dry culture conditions
  2. b Daily medium replacement during dry culture
  3. c Culture in dry conditions for 6 days
Phase 5 · Bacterial preparation and co-culture Variable (co-culture initiated after ALI maturation)
  1. a Prepare S. aureus inoculum
  2. b Prepare C. acnes inoculum
  3. c Wash ALI-organoids with antibiotic-free medium
  4. d Inoculate bacteria on organoid surface
  5. e Co-culture under dry conditions
Phase 6 · C. acnes pre-treatment protocol (optional) Variable
  1. a Pre-treat organoids with C. acnes
  2. b Subsequent S. aureus challenge

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Attribution

This SOP was written by Organthis based on the published method in Jung et al., 2022. Once the originating laboratory has confirmed it, the SOP is marked Verified.

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