O2160: A Titratable Model of Dextran Sodium Sulfate-induced Necrotizing Enterocolitis (NEC) Demonstrates Neuroinflammation and Activation of Microglia in Newborn Mice

Background: Necrotizing enterocolitis (NEC) is a devastating condition of preterm infants with a mortality rate of 10 – 30%. NEC leads to an intense gastrointestinal and systemic inflammatory response that can result in multi-organ damage, including brain injury. The biological pathways underlying the progression of enterocolitis to systemic and cerebral inflammation and associated organ injury are, however, poorly understood. We therefore developed a titratable model of NEC based upon the dextran sodium sulfate which will allow us to investigate the various severity stages of NEC on the immature newborn brain.

Methods: Postnatal day 3 (PND3) C57BL/6 mice were fed formula (Esbilac) supplemented with increasing concentration of dextran sodium sulfate (DSS). Data from DSS fed mice were compared to mice fed formula alone (0% DSS) or those fed by their dams. Animal weight and clinical sickness score (Zani et al., 2008, Eur J Pediatr Surgery) were collected during NEC induction . After 72 hours, animals were euthanized and organ tissues (intestine, liver, brain) were harvested for histological evaluation, cytokine analysis, immunohistochemistry and quantitative image analysis. ANOVA, student’s t-test were used to determine significance.

Results: Mice fed higher concentrations of DSS exhibited more rapid weight loss and clinical deterioration as measured by clinical sickness score when compared to formula fed or dam fed mice. Macroscopic gut assessment demonstrated significantly worse scores in mice fed higher DSS concentrations. In contrast, histological evaluation of small intestines demonstrated enterocolitis in all mice that received DSS supplementation (Figure 1). Pro-inflammatory cytokine concentrations in hepatic tissue increased with rising DSS concentrations (Figure 2). Both formula feeding and DSS supplementation resulted in proinflammatory cytokine expression in the brain compared to dam-fed controls. Although the density of microglia in the brain did not change, morphological analysis demonstrated microglial activation within the brains of all DSS fed mice.

Conclusion: Utilizing a novel titratable model of NEC, we demonstrate that even mild NEC can induce microglia activation and neuroinflammation, which become more pronounced with increasing NEC severity. This model will allow us to study the early stages of NEC and may help to identify brain protective treatment strategies in the future.