Complex Regulation of Intestinal Relaxin Receptor (LGR7) Levels Is Time-Dependent and Altered By Necrotizing Enterocolitis in Rat Pups
Purpose: Necrotizing enterocolitis (NEC), an often fatal disorder of neonates, is at least partially mediated by intestinal vasoconstriction with subsequent hypoperfusion, inflammation and organ failure. The pathophysiology of NEC-associated vasoconstriction is complex and includes deranged balance of microvascular tone with enhanced endothelin vasoactivity and limited compensatory nitric oxide vasodilation. Relaxin, a potent vasodilatory hormone that regulates pregnancy and is found in maternal milk, has also been implicated in the pathogenesis of NEC and oral relaxin might provide a new treatment modality. Relaxin-mediated vasodilation is controlled by ubiquitous G-protein coupled relaxin receptors (LGR7) found on endothelial cells including in the gastrointestinal tract. The pattern of LGR7 protein expression in neonatal rat pups is not known. We hypothesized that LGR7 receptors might be dysregulated during NEC in response to varying exposure to vasoactive relaxin.
Methods: Sprague-Dawley rat pups were separated by litter. CONTROL pups were vaginally delivered and dam-fed. NEC was induced in premature pups (C-section) by intermittent exposure to cold, hypoxia, lipopolysaccharide and formula feeding. At 12-, 24-, 48-, 72- and 96-hours post-birth, serum, ileum and enteral samples were obtained from pups. Serum relaxin, enteral relaxin levels and LGR-7 levels in ileum samples (western blot) were measured. Levels were compared between groups and time points by 2-way analysis of variance (ANOVA) and Tukey test with p<0.05 a priori.
Results: The pattern of ileal LGR7 protein expression in CONTROL pups was time-dependent and bi-phasic with peaks in expression observed at 48- and 96-hours post-birth in spite of stable enteral levels in maternal milk. This pattern was altered in the groups undergoing the NEC protocol such that a single lesser peak was observed at 72-hours post-birth, with no relaxin detected in formula. LGR-7 levels increased over time in NEC animals in response to decreasing serum levels of relaxin, while this decrease in serum levels did not prompt a similar response in the CONTROL group.
Conclusion: Our data demonstrate that the expression of vasoactive relaxin receptors in the neonatal period vary, probably due to feedback regulation in response to diminishing relaxin levels over time in the CONTROLS. However, in this model of NEC, ileal LGR7 expression is dampened and delayed compared to the CONTROL pattern. In NEC, altered relaxin levels could be attributed to both lack of relaxin in formula feeds and loss of the maternal source of relaxin after delivery. Since relaxin is thought to cross the intestinal barrier intact in neonates, these observations suggest that oral relaxin supplementation might provide a therapeutic benefit in NEC since LGR7 receptors, while diminished, are still present.