Neonatal Intestinal Macrophages Promote Microvascular Development through an IGF‐1‐dependent Mechanism and Lack of Macrophage‐derived IGF‐1 Predisposes Newborn Mice to Necrotizing Enterocolitis

Abstract

Defective intestinal microvascular development has been previously shown to play a role in necrotizing enterocolitis (NEC) pathogenesis. Decreased serum levels of IGF-1 in premature infants are associated with a higher risk of developing NEC and we recently found that serum and intestinal tissue IGF-1 level are decreased in a neonatal mouse NEC model. In the embryo, macrophages have been reported to promote angiogenesis. Here we hypothesize that neonatal intestinal macrophages promote endothelial cell proliferation and angiogenesis by producing IGF-1, thus protecting against NEC development. For this purpose, we first examined the role of IGF-1 on intestinal endothelial cell sprouting in vitro using IGF-1 and IGF-1R inhibitor and on intestinal endothelial cell proliferation in vivo using IGF-1R inhibitor. Secondly, using mice with IGF-1-deficient-macrophages and IGF-1-sufficient littermates, we assessed whether macrophage-derived IGF-1 plays a role in perinatal microvascular development by examining endothelial cell proliferation in vivo and endothelial cell sprouting in vitro in Matrigel co-cultures. Finally, we examined whether macrophage-derived IGF-1 protects against NEC. We found that, neonatal intestinal CX3CR1+ macrophages were essential for small intestinal endothelial cells to sprout in Matrigel, which process was further enhanced by exogenous IGF-1 and inhibited by the selective IGF-1R inhibitor picropodophyllin (PPP). Intestinal endothelial cell proliferation was also inhibited by PPP in vivo, to a similar extent as observed during NEC development. Furthermore, neonatal IGF-1-deficient macrophages significantly decreased pro-angiogenic activity in vitro. Mice with IGF-1-deficient-macrophages had less robust intestinal endothelial proliferation and decreased intestinal VEGF protein expression in vivo. Lastly, pups with IGF-1-deficient macrophages had increased incidence of severe NEC-like intestinal injury and mortality. In conclusion, neonatal intestinal macrophages are critical for promoting intestinal microvascular growth and for protecting pups from NEC development via IGF-1 production.