Iron Transporter Regulation in Necrotizing Enterocolitis

Authors:
Christopher Luschen, Heather Grubbs, Luciana Previato de Almeida, Bhawana Luitel, Catherine Hunter

Body of Abstract:
Introduction:

Necrotizing Enterocolitis (NEC) is a devastating disease affecting preterm infants associated with hyperinflammation, increased intestinal permeability, and cell death. Ferroptosis has been identified as an important pathway of cell death in NEC pathogenesis. Labile intracellular iron is the critical driver of ferroptosis through the Fenton Reaction. The increase in intracellular iron is not well delineated. The transport of iron predominantly occurs through FPN, DMT1 and TFTR. In other scenarios with elevated iron, FPN often is found to be upregulated. We hypothesize that FPN, DMT1, and TFTR show no difference in control vs NEC tissue leading to the accumulation of iron.

Methods:

Following IRB approval, human intestinal tissue segments from infants undergoing bowel resection for benign reasons such as atresia or ostomy takedowns as well as from infants undergoing bowel resection for NEC were obtained. The tissue then underwent inductively coupled mass spectrometry and iron calorimetric assay processing for iron concentration. Additionally, RNA was extracted from control tissue and from NEC tissue. Total RNA isolated from NEC and control tissue was then converted to cDNA for RT-qPCR analysis of FPN, DMT1, and TFTR.

Results:

Using inductively coupled plasma mass spectrometry and iron calorimetric assay, total iron was found to be significantly elevated in necrotizing enterocolitis tissue compared to control (p<0.05). FPN expression was found to be nonsignificant in human NEC tissue compared to control (p<0.5764). DMT1 expression was found to be nonsignificant in comparing control to NEC tissue (p<0.3515). Additionally, TFTR expression was found to be nonsignificant in control vs NEC tissue (p<0.7028). Conclusion: Iron levels are significantly elevated emphasizing the role of ferroptotic cell death in necrotizing enterocolitis. FPN, DMT, and TFTR display no difference even in the presence of elevated iron level as seen in NEC. FPN, when exposed to elevated iron levels, often is upregulated. However, this is not displayed in our experiment therefore showing a dysregulated response. This provides the foundation for further investigation into iron regulation and the ferroptosis pathway.