Cronobacter sakazakii induces necrotizing enterocolitis by regulating NLRP3 inflammasome expression via TLR4

J Med Microbiol. 2020 May;69(5):748-758. doi: 10.1099/jmm.0.001181. Epub 2020 Mar 25.


Introduction. Neonatal infection with Cronobacter sakazakii can cause severe intestinal damage and necrotizing enterocolitis (NEC). The inflammasome and Toll-like receptors mediate intestinal damage caused by other intestinal pathogens causing NEC, but the exact mechanism is unclear.Aim. We evaluated the molecular mechanisms underlying C. sakazakii-induced NEC.Methodology. The effects of C. sakazakii treatment on two cell lines and a Sprague-Dawley rat model of NEC were evaluated by a cell death assay, western blot and real-time PCR analyses of the NLRP3 inflammasome and downstream factors, and observation of cell and intestinal damage.Results. C. sakazakii caused cellular damage in vitro, as well as intestinal damage in an animal model. NLRP3, caspase-1, TLR4 and MyD88, as well as the downstream factor IL-1β, were upregulated in C. sakazakii-infected J774A.1 and HT-29 cells. Western blotting showed that C. sakazakii-infected J774A.1 and HT-29 cells and the NEC rat model had higher expression levels of N-terminal gasdermin D (GSDMD) compared with those in the control groups. C. sakazakii and its components promote NF-κB expression via the TLR4/MyD88 signalling pathway, thereby regulating the NLRP3 inflammasome and mediating GSDMD cleavage, resulting in pyroptosis-induced intestinal damage.Conclusion. We found that C. sakazakii upregulates NF-κB via TLR4/MyD88 to promote activation of the NLRP3 inflammasome, leading to the up-regulation of downstream caspase-1, release of IL-1β, GSDMD-mediated pyroptosis and development of NEC. These findings clarify the mechanisms by which C. sakazakii contributes to NEC.

Keywords: Cronobacter sakazakii; Gasdermin D; NLRP3; necrotizing enterocolitis.

MeSH terms

  • Animals
  • Animals, Newborn
  • Cell Cycle / drug effects
  • Cell Line
  • Cells, Cultured
  • Cronobacter sakazakii / physiology*
  • Disease Models, Animal
  • Enterocolitis, Necrotizing / genetics*
  • Enterocolitis, Necrotizing / metabolism*
  • Enterocolitis, Necrotizing / microbiology*
  • Enterocolitis, Necrotizing / pathology
  • Gene Expression Regulation*
  • Host-Pathogen Interactions
  • Humans
  • NF-kappa B / metabolism
  • NLR Family, Pyrin Domain-Containing 3 Protein / genetics*
  • Pyroptosis
  • Rats
  • Signal Transduction
  • Sulfonamides / pharmacology
  • Toll-Like Receptor 4 / antagonists & inhibitors
  • Toll-Like Receptor 4 / metabolism*


  • NF-kappa B
  • NLR Family, Pyrin Domain-Containing 3 Protein
  • Sulfonamides
  • Toll-Like Receptor 4
  • ethyl 6-(N-(2-chloro-4-fluorophenyl)sulfamoyl)cyclohex-1-ene-1-carboxylate