Severe intestinal inflammation associated necrotizing enterocolitis does not trigger brain injury in neonates

Necrotizing enterocolitis (NEC) is a severe inflammatory gastrointestinal condition that predominantly affects preterm infants. Despite of its acute intestinal impact, clinical studies show that survivors of severe NEC are at increased risk for neurodevelopmental impairment later in their life. Emerging evidence suggests that NEC-associated pathophysiological events may contribute to these long-term effects. Previous research shows that brain injury occurs in mouse pups during NEC development in response to NEC-associated stressors, suggesting the role of the gut-brain axis in the pathogenesis of NEC-associated neurodevelopmental impairment. However, the role of intestinal injury on NEC-associated brain injury is not clear. Here, we thought to examine whether NEC-associated intestinal inflammation can trigger neonatal brain injury using our novel mouse model of NEC induced by crypt intestinal epithelial cell (IEC) apoptosis. To this end, seven days old pups of 3xTg-iAP cIEC mice (3 Transgenes [Tg; Fasl, rtTA, and Vil-Cre] for inducible Apoptosis in crypt-IEC) were administered single dose of doxycycline (Dox; 20 µg/gm) through subcutaneous route to induce NEC as described in Cell Mol Gastroenterol Hepatol. 2024;18(3):101364. The neonatal pups with the absence of Fasl Tg were considered as littermate controls and received the same treatment as the control group. The growth of mouse pups was monitored on a daily basis. After three days of Dox treatment, pups were sacrificed, and intestinal and brain tissues were collected for H.E. staining followed by histopathological examinations. Furthermore, brain tissues were processed for characterization of cell apoptosis and microglial activation with immunofluorescence and TUNEL staining, respectively. We confirmed that 3xTg-iAP cIEC mouse pups developed NEC-like intestinal injury within 72 h after Dox treatment. Clinical observation showed a slight retardation in the animal’s growth and body weight of Dox-treated 3xTg- iAP cIEC pups during NEC development. Histological examination did not reveal any abnormality in the brain of 3xTg-iAP cIEC mouse pups with NEC compared to their littermate controls at 72 h. Using TUNEL staining, we observed scattered apoptosis mainly in microglial cells of littermate controls. However, 3xTg-iAP cIEC mouse pups with NEC did not exhibit an increase in apoptosis in their brain. Taken together, our study suggests that the pathophysiological events associated with NEC, induced by intestinal crypt IEC apoptosis, do not result in brain injury in neonates simultaneously. The mechanism underlying the increased risk of neurodevelopmental impairment in NEC survivors remains to be studied in the future. This work is supported by grants from NIH and VA. This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.