IFN-γ production by CD4 CD8 double positive cells is essential for expansion of Proteobacteria in the neonatal murine intestine.

Abstract

Abstract Background Expansion of Proteobacteria is seen in the neonatal intestine in both humans and mice. Proteobacteria are also significantly over-represented in preterm infants that develop Necrotizing Enterocolitis (NEC). Neonatal mice are more susceptible to models of NEC. The mechanism of expansion of Proteobacteria in neonatal mice and susceptibility to NEC remain undefined. Objective To identify immune modulators that favor expansion of Proteobacteria in the neonatal murine intestine. Methods qRT-PCR of bacterial 16S rRNA gene sequence tags of colonic fecal contents from 1–3 week (wk) old wild type (WT), IFN-γ knock out and Rag deficient mice was performed to identify major bacterial groups. qRT-PCR and ELISA of whole colon were used to profile cytokine expression. Flow-cytometry of lamina propria cells was used to characterize cells in 1-,2- and 3-wk old mice. IFN-γ neutralizing Ab was given IP to 3 day old mice and then similarly examined at 2 wks. Results 2-wk-old WT mice showed increased expansion of Proteobacteria and increased production of IFN-γ. IFN-γ KO mice had minimal expansion of Proteobacteria in the colon, this was also seen in WT mice blocked with IFN-γ Ab. Flow-cytometry revealed a unique subset of CD4 CD8 double positive cells, only present in neonatal mice, that produce IFN-γ in WT mice. Rag deficient mice did not have this subset of cells and also had minimal colonization by Proteobacteria at 2-wks. Conclusions IFN-γ promotes the expansion of Proteobacteria in the neonatal intestine. A unique and transient subset of CD4 CD8 cells, that are absent in Rag deficient mice, appear to be responsible for the increased IFN-γ in neonatal mice. This is a potential target for intervention in the prevention of NEC in preterm infants.