Microtubular Disruption Alters Intestinal Permeability in Murine Neonatal Anemic Intestine

Anemia is a frequent diagnosis in premature and critically ill infants and is caused by phlebotomy, which is essential for medical care. The severity of anemia often correlates with the acuity of illness. We recently showed that severe neonatal anemia causes a "leaky gut" in neonatal mouse pups by altering the structure and function of epithelial adherens junctions through decreased E-cadherin expression. We hypothesize that microtubules play a major role in maintaining adherens junction integrity. To test this hypothesis, C57BL/6 mice were subjected to timed phlebotomy between postnatal days (P) 2-10 to induce severe anemia (hematocrits 20%-24%). Microtubular assembly was evaluated by electron microscopy, quantitative real-time polymerase chain reaction (qRT-PCR), immunohistochemistry, and/or immunoblotting on intestinal tissues, Caco-2 intestinal epithelial-like cells, and colonic organoids. The electron micrographs showed loss of microtubules in the anemic intestine and disrupted intestinal epithelial microtubular assembly. Microtubular disruption was associated with decreased expression of the α-tubulin 4a (tuba4a) subunit. The observed ultrastructural changes that disrupt epithelial barrier functions result from hypoxia-induced microRNA let-7e destabilizing tuba4a expression in the anemic intestinal epithelium. In conclusion, phlebotomy-induced anemia in mouse neonates is associated with a "leaky gut" by the disruption of microtubular assembly.