Toll-like receptor 4-mediated endoplasmic reticulum stress in intestinal crypts induces necrotizing enterocolitis

Amin Afrazi, Maria F. Branca, Chhinder P. Sodhi, Misty Good, Yukihiro Yamaguchi, Charlotte E. Egan, Peng Lu, Hongpeng Jia, Shahab Shaffiey, Joyce Lin, Congrong Ma, Garrett Vincent, Thomas Prindle, Samantha Weyandt, Matthew D. Neal, John A. Ozolek, John Wiersch, Markus Tschurtschenthaler, Chiyo Shiota, George K. GittesTimothy R. Billiar, Kevin Mollen, Arthur Kaser, Richard Blumberg, David J. Hackam*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

136 Scopus citations


The cellular cues that regulate the apoptosis of intestinal stem cells (ISCs) remain incompletely understood, yet may play a role in diseases characterized by ISC loss including necrotizing enterocolitis (NEC). Toll-like receptor-4 (TLR4) was recently found to be expressed on ISCs, where its activation leads to ISC apoptosis through mechanisms that remain incompletely explained.Wenow hypothesize that TLR4 induces endoplasmic reticulum (ER) stress within ISCs, leading to their apoptosis in NECpathogenesis, and that high ER stress within the premature intestine predisposes to NEC development. Using transgenic mice and cultured enteroids, we now demonstrate that TLR4 induces ER stress within Lgr5 (leucine-rich repeat-containing G-protein-coupled receptor 5)-positive ISCs, resulting in crypt apoptosis. TLR4 signaling within crypts was required, because crypt ER stress and apoptosis occurred in TLR4 ΔIEC-OVER mice expressing TLR4 only within intestinal crypts and epithelium, but not TLR4ΔIEC mice lacking intestinal TLR4. TLR4-mediated ER stress and apoptosis of ISCs required PERK (protein kinaserelated PKR-like ER kinase), CHOP (C/EBP homologous protein), and MyD88 (myeloid differentiation primary response gene 88), but not ATF6 (activating transcription factor 6) or XBP1 (X-box-binding protein 1). Human and mouse NEC showed high crypt ER stress and apoptosis, whereas genetic inhibition of PERK or CHOP attenuated ER stress, crypt apoptosis, and NEC severity. Strikingly, using intragastric delivery into fetal mouse intestine, prevention of ER stress reduced TLR4-mediated ISC apoptosis and mucosal disruption. These findings identify a novel link between TLR4-induced ER stress and ISC apoptosis in NEC pathogenesis and suggest that increased ER stress within the premature bowel predisposes to NEC development.

Original languageEnglish
Pages (from-to)9584-9599
Number of pages16
JournalJournal of Biological Chemistry
Issue number14
StatePublished - 4 Apr 2014
Externally publishedYes


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