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Calcium/calmodulin-dependent protein kinase IV signaling pathway is upregulated in experimental necrotizing enterocolitis

  • Mashriq Alganabi
  • Haitao Zhu
  • Joshua S. O’Connell
  • George Biouss
  • Andrea Zito
  • Bo Li
  • Edoardo Bindi
  • Agostino PierroEmail author
Original Article

Abstract

Purpose

Activation of calcium/calmodulin-dependent protein kinase IV (CaMKIV) has been shown to increase intestinal injury and inhibit epithelial cell proliferation in dextran sulfate sodium (DSS)-induced colitis mice. However, the role of CaMKIV in necrotizing enterocolitis (NEC) is unknown. We aimed to study the expression and activation of CaMKIV in experimental NEC.

Methods

Following ethical approval, NEC (n = 10) was induced in C57BL/6 mouse pups by hypoxia, gavage hyperosmolar formula feeding and lipopolysaccharide from postnatal days P5 to 9. Breastfed pups served as control (n = 10). Mouse pups were sacrificed on P9 and the terminal ileum was harvested. Gene NEC injury was scored blindly by three independent investigators. CaMKIV, CREM and IL17 gene expression, and CaMKIV and pCaMKIV protein expression were assessed. The data were compared using Mann–Whitney U test. P < 0.05 was considered significant.

Results

Intestinal injury was induced in the NEC mice and confirmed by histological scoring and inflammatory cytokine IL6. CaMKIV and its downstream target genes of CREM and IL17 were significantly elevated in NEC mice relative to control. Similarly, phosphorylated-CaMKIV (pCaMKIV), the active form of CaMKIV, was more notably expressed in the NEC ileal tissue relative to control ileal tissue. Elevated pCaMKIV protein expression was also confirmed by western blot.

Conclusion

CaMKIV expression and activation are upregulated in experimental NEC suggesting a potential contributing factor in the pathogenesis of NEC.

Keywords

Necrotizing enterocolitis (NEC) Calcium/calmodulin-dependent protein kinase IV (CaMKIV) pCaMKIV IL17 

Notes

Acknowledgements

AP is supported by a Canadian Institutes of Health Research (CIHR) Foundation Grant 353857 and The Hospital for Sick Children. The funding had no impact on study design, data collection, analysis, interpretation, the writing of the report, or the decision to submit the paper for publication. We would also like to thank Maarten Janssen Lok for his contribution to histological scoring and manuscript revision.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

Animal experiments received ethical approval by the Animal Care Committee of The Hospital for Sick Children (No. 44032) and were performed in accordance with its guidelines and regulations.

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2020

Authors and Affiliations

  1. 1.Division of General and Thoracic Surgery, Translational Medicine Program, The Hospital for Sick ChildrenUniversity of TorontoTorontoCanada

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