Experimental necrotizing enterocolitis induces neuroinflammation in the neonatal brain

George Biouss,Shogo Seo,Joshua S O’Connell,Elke Zani-Ruttenstock,Bo Li,Nataliia Svergun,Agostino Pierro,Abidur Rahman,Lina Antounians,Jennifer Guadagno,Vincenzo D Catania,Augusto Zani

doi:10.1186/s12974-019-1481-9

George Biouss, Shogo Seo + Show 10 more

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https://doi.org/10.1186/s12974-019-1481-9

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Abstract

BackgroundNecrotizing enterocolitis (NEC) is an inflammatory gastrointestinal disease primarily affecting preterm neonates. Neonates with NEC suffer from a degree of neurodevelopmental delay that is not explained by prematurity alone. There is a need to understand the pathogenesis of neurodevelopmental delay in NEC. In this study, we assessed the macroscopic and microscopic changes that occur to brain cell populations in specific brain regions in a neonatal mouse model of NEC. Moreover, we investigated the role of intestinal inflammation as part of the mechanism responsible for the changes observed in the brain of pups with NEC.MethodsBrains of mice were assessed for gross morphology and cerebral cortex thickness (using histology). Markers for mature neurons, oligodendrocytes, neural progenitor cells, microglia, and astrocytes were used to quantify their cell populations in different regions of the brain. Levels of cell apoptosis in the brain were measured by Western blotting and immunohistochemistry. Endoplasmic reticulum (ER) stress markers and levels of pro-inflammatory cytokines (in the ileum and brain) were measured by RT-qPCR and Western blotting. A Pearson test was used to correlate the levels of cytokines (ELISA) in the brain and ileum and to correlate activated microglia and astrocyte populations to the severity of NEC.ResultsNEC pups had smaller brain weights, higher brain-to-body weight ratios, and thinner cortices compared to control pups. NEC pups had increased levels of apoptosis and ER stress. In addition, NEC was associated with a reduction in the number of neurons, oligodendrocytes, and neural progenitors in specific regions of the brain. Levels of pro-inflammatory cytokines and the density of activated microglia and astrocytes were increased in the brain and positively correlated with the increase in the levels pro-inflammatory cytokines in the gut and the severity of NEC damage respectively.ConclusionsNEC is associated with severe changes in brain morphology, a pro-inflammatory response in the brain that alters cell homeostasis and density of brain cell populations in specific cerebral regions. We show that the severity of neuroinflammation is associated with the severity of NEC. Our findings suggest that early intervention during NEC may reduce the chance of acute neuroinflammation and cerebral damage.

Highlights

Necrotizing enterocolitis (NEC) is an inflammatory gastrointestinal disease primarily affecting preterm neonates
The density of cleaved caspase 3 (CC3)+ cells increased in the brain of pups exposed to hypoxia only in comparison with that of breastfed control (p < 0.0001) in the hippocampus [hypoxia group 130 (67–258)], basal ganglia/thalamus [hypoxia group 146 (82–247)], and the cerebral cortex regions [hypoxia group 82 (33–184)]; CC3+ cell density was less in the brain of pups with NEC (p = 0.002, p < 0.0001, p = 0.01, in the hippocampus, basal ganglia/thalamus, and cerebral cortex, respectively Additional file 2: Figure S2C, D)
As we observed a decrease in the number of neurons and oligodendrocytes, we investigated the density of Sox2+ neural progenitor cells that can differentiate into these cell types [50, 51]

Summary

Introduction

Necrotizing enterocolitis (NEC) is an inflammatory gastrointestinal disease primarily affecting preterm neonates. Neonates with NEC suffer from a degree of neurodevelopmental delay that is not explained by prematurity alone. There is a need to understand the pathogenesis of neurodevelopmental delay in NEC. Necrotizing enterocolitis (NEC) is the most severe gastrointestinal neonatal emergency that primarily affects preterm infants with very low and extremely low birth weight [1, 2]. The morbidity rates for NEC survivors are high and include not just poor long-term gastrointestinal outcomes, and neurodevelopmental delay [4]. A systematic review of the literature showed that more than 50% of infants with NEC have neurodevelopmental delay, with significantly worse neurodevelopmental outcome than prematurity alone [5]. Among patients diagnosed with NEC, those that are surgically managed have been reported to have worse mental and psychomotor developmental index scores compared with age-matched control [5, 6]

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