Abstract
Intestinal ischemic injury results sloughing of the mucosal epithelium leading to host sepsis and death unless the mucosal barrier is rapidly restored. Volvulus and neonatal necrotizing enterocolitis (NEC) in infants have been associated with intestinal ischemia, sepsis and high mortality rates. We have characterized intestinal ischemia/repair using a highly translatable porcine model in which juvenile (6-8-week-old) pigs completely and efficiently restore barrier function by way of rapid epithelial restitution and tight junction re-assembly. In contrast, separate studies showed that younger neonatal (2-week-old) pigs exhibited less robust recovery of barrier function, which may model an important cause of high mortality rates in human infants with ischemic intestinal disease. Therefore, we aimed to further refine our repair model and characterize defects in neonatal barrier repair. Here we examine the defect in neonatal mucosal repair that we hypothesize is associated with hypomaturity of the epithelial and subepithelial compartments. Following jejunal ischemia in neonatal and juvenile pigs, injured mucosa was stripped from seromuscular layers and recovered ex vivo while monitoring transepithelial electrical resistance (TEER) and 3H-mannitol flux as measures of barrier function. While ischemia-injured juvenile mucosa restored TEER above control levels, reduced flux over the recovery period and showed 93±4.7% wound closure, neonates exhibited no change in TEER, increased flux, and a 11±23.3% increase in epithelial wound size. Scanning electron microscopy revealed enterocytes at the wound margins of neonates failed to assume the restituting phenotype seen in restituting enterocytes of juveniles. To attempt rescue of injured neonatal mucosa, neonatal experiments were repeated with the addition of exogenous prostaglandins during ex vivo recovery, ex vivo recovery with full thickness intestine, in vivo recovery and direct application of injured mucosal homogenate from neonates or juveniles. Neither exogenous prostaglandins, intact seromuscular intestinal layers, nor in vivo recovery enhanced TEER or restitution in ischemia-injured neonatal mucosa. However, ex vivo exogenous application of injured juvenile mucosal homogenate produced a significant increase in TEER and enhanced histological restitution to 80±4.4% epithelial coverage in injured neonatal mucosa. Thus, neonatal mucosal repair can be rescued through direct contact with the cellular and non-cellular milieu of ischemia-injured mucosa from juvenile pigs. These findings support the hypothesis that a defect in mucosal repair in neonates is due to immature repair mechanisms within the mucosal compartment. Future studies to identify and rescue specific defects in neonatal intestinal repair mechanisms will drive development of novel clinical interventions to reduce mortality in infants affected by intestinal ischemic injury.
Highlights
The intestinal mucosa is lined by a single layer of epithelial cells, which form the principal barrier against luminal bacteria and their toxins and simultaneously facilitate selective absorption of electrolytes, water and nutrients
Volvulus in infants most commonly occurs due to anatomical abnormalities and has poorer outcomes associated with younger age, delay of intervention, and presence of intestinal necrosis[6,7,8]
In order to determine the effect of postnatal age on level of injury following intestinal ischemia, neonatal and juvenile aged animals were subjected to jejunal ischemia for varying durations and intestinal barrier integrity was studied
Summary
The intestinal mucosa is lined by a single layer of epithelial cells, which form the principal barrier against luminal bacteria and their toxins and simultaneously facilitate selective absorption of electrolytes, water and nutrients. Increasing durations of ischemia will induce progressively increased epithelial loss down the villus-crypt axis until the crypts are sloughed and the intestinal architecture is distorted[4] This breakdown of the intestinal epithelial barrier leads to sepsis, intestinal necrosis and host death unless the barrier completely and rapidly repairs following an ischemic event. Novel treatments have focused on enhancing de novo formation of new epithelial cells, but this requires support of the patient for days following the initial injury until newly produced epithelial cells can restore the mucosal barrier [11, 12] In this subacute repair phase, remaining epithelium must immediately restitute the damaged barrier to curtail sepsis early and prevent host mortality until the regenerative phase can fully restore intestinal architecture [1]. Our lab has focused on understanding the mechanisms of the subacute phase of repair, as interventions enhancing this phase will improve patient survival and hopefully reduce intestinal necrosis and the need for resection in order to improve long-term quality of life
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