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LIVER Publication-derived

IL-17 Signaling in Primary Sclerosing Cholangitis Patient-Derived Extrahepatic Cholangiocyte Organoids

Source Moreno et al., 2024 · Mayo Clinic, Division of Gastroenterology and Hepatology, Rochester, Minnesota, USA · 10.1097/hc9.0000000000000454

👤 Ana S. Garcia Moreno, Maria E. Guicciardi, Alexander Q. Wixom, Erik Jessen, Jingchun Yang, Sumera I. Ilyas, Jackie K. Bianchi, Filippo Pinto e Vairo, Konstantinos N. Lazaridis, Gregory J. Gores ⏱ 15 days 📋 4 phases 🧫 Patient-Derived Cholangiocytes (Primary Sclerosing Cholangitis and Non-PSC Controls)

Abstract

This protocol describes the generation and IL-17A stimulation of extrahepatic cholangiocyte organoids (ECO) derived from patients with primary sclerosing cholangitis (PSC) and non-PSC controls. The ECO are cultured from cholangiocytes obtained via endoscopic retrograde cholangiography, expanded in Matrigel-based organoid culture, and treated with recombinant human IL-17A to assess transcriptomic, secretomic, and genomic responses.

Cell source
Patient-Derived Cholangiocytes (Primary Sclerosing Cholangitis and Non-PSC Controls)
Application
Disease modeling; Immunological pathway interrogation in cholestatic liver disease

Protocol overview

18 steps across 4 phases

Patient Sample Collection and Cholangiocyte Isolation Day 0
  1. 1 Endoscopic Retrograde Cholangiography (ERC) and Sample Acquisition
  2. 2 Bile Sample Preparation
  3. 3 Cytology Brush Processing
  4. 4 Cell Pellet Preparation
Organoid Initiation and Expansion Days 0–14 (initial expansion)
  1. 1 Matrigel Embedding and Seeding
  2. 2 Initial Organoid Culture (First 3 Days)
  3. 3 Medium Transition to Expansion Medium
  4. 4 Organoid Passage
IL-17A Stimulation and Analysis Day 14–Day 15 (experimental treatment)
  1. 1 IL-17A Treatment Preparation
  2. 2 IL-17A Treatment of ECO
  3. 3 Sample Collection for Transcriptomic Analysis
  4. 4 Supernatant Collection for Secretome Analysis
Characterization by Immunofluorescence Day 7–Day 14 (parallel to expansion)
  1. 1 Whole-Mount Immunofluorescence Sample Preparation
  2. 2 Permeabilization and Blocking
  3. 3 Primary Antibody Staining
  4. 4 Washing and Secondary Antibody Staining
  5. 5 Nuclear Counterstaining and Mounting
  6. 6 Confocal Microscopy Imaging

Full SOP

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Attribution

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

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