Bo Li1, Carol Lee1, Marissa Cadete1, Hiromu Miyake1, Dorothy Lee1, Agostino Pierro2. 1. Division of General and Thoracic Surgery, Translational Medicine Program, The Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, M5G 1X8, Canada. 2. Division of General and Thoracic Surgery, Translational Medicine Program, The Hospital for Sick Children, 555 University Avenue, Toronto, Ontario, M5G 1X8, Canada. agostino.pierro@sickkids.ca.
Abstract
BACKGROUND: Adult intestinal organoids have been used to study ex vivo intestinal injury in adulthood. However, the neonatal intestinal epithelium has many unique features that are different from adult mature intestine. Establishing a neonatal ex vivo organoid model is essential to study the epithelial physiology in early postnatal development and to investigate derangements associated with disease processes during the neonatal period like necrotizing enterocolitis (NEC). METHODS: Fresh and frozen terminal ileum was harvested from mice pups on postnatal day 9. Crypts were isolated and organoids were cultured. Organoids were exposed to hypoxia and lipopolysaccharide (LPS) for 48 h to induce epithelial injury. Inflammatory cytokines and tight junction proteins were evaluated. RESULTS: Robust intestinal organoids can be formed from both fresh and frozen intestinal tissue of neonatal mice pups. Hypoxia and LPS administration induced intestinal inflammation and disrupted tight junctions in these neonatal intestinal organoids. CONCLUSIONS: We have established a novel method to grow organoids from neonatal intestine. We demonstrated that these organoids respond to the injury occurring during neonatal intestinal diseases such as NEC by increasing the organoid inflammation and by disrupting the organoid barrier function. Organoids provide an ex vivo platform to study intestinal physiology and pathology during the neonatal period.
BACKGROUND: Adult intestinal organoids have been used to study ex vivo intestinal injury in adulthood. However, the neonatal intestinal epithelium has many unique features that are different from adult mature intestine. Establishing a neonatal ex vivo organoid model is essential to study the epithelial physiology in early postnatal development and to investigate derangements associated with disease processes during the neonatal period like necrotizing enterocolitis (NEC). METHODS: Fresh and frozen terminal ileum was harvested from mice pups on postnatal day 9. Crypts were isolated and organoids were cultured. Organoids were exposed to hypoxia and lipopolysaccharide (LPS) for 48 h to induce epithelial injury. Inflammatory cytokines and tight junction proteins were evaluated. RESULTS: Robust intestinal organoids can be formed from both fresh and frozen intestinal tissue of neonatal mice pups. Hypoxia and LPS administration induced intestinal inflammation and disrupted tight junctions in these neonatal intestinal organoids. CONCLUSIONS: We have established a novel method to grow organoids from neonatal intestine. We demonstrated that these organoids respond to the injury occurring during neonatal intestinal diseases such as NEC by increasing the organoid inflammation and by disrupting the organoid barrier function. Organoids provide an ex vivo platform to study intestinal physiology and pathology during the neonatal period.
Authors: Bo Li; Carol Lee; Marissa Cadete; Haitao Zhu; Yuhki Koike; Alison Hock; Richard Y Wu; Steven R Botts; Adam Minich; Mashriq Alganabi; Lijun Chi; Elke Zani-Ruttenstock; Hiromu Miyake; Yong Chen; Annika Mutanen; Bo Ngan; Kathene C Johnson-Henry; Paolo De Coppi; Simon Eaton; Pekka Määttänen; Paul Delgado-Olguin; Philip M Sherman; Augusto Zani; Agostino Pierro Journal: Cell Death Dis Date: 2019-10-03 Impact factor: 8.469