Fig. 3. Toll-like receptor 4 dependent loss of enteric glia loss is required for the development of NEC in mice.

(A to D) representative H&E-stained photomicrographs of wild-type and Sox10-Tlr4^ko mice that are either breast fed and allowed to stay with the dam (“Ctrl”) or induced to develop NEC (“NEC”). (E to H) Representative confocal images of terminal ileum of wild-type and Sox10-Tlr4^ko mice that were either control or induced to develop NEC; sections were stained for DAPI (blue) and enteric glia (Sox10, red). (I) quantification of Sox10 immunofluorescence in wild type and Sox10-TLR4^ko intestinal sections corresponding to E to H, reflecting the abundance of enteric glia (n≥10 sections/group). (J) qRT-PCR expression of Tnf-α in the intestinal mucosa of the indicated group (n≥6 mice/group) (K) NEC severity scores corresponding to E to H (n=6 histological sections scored/group). (L) Intestinal motility in wild-type and Sox10-TLR4^ko mice with and without NEC as indicated (n≥7 mice/group); Each dot represents data from individual mice. Statistical significance was determined by one-way ANOVA, followed by Tukey’s multiple comparisons tests using GraphPad prism 9 software. ***p<0·001. Scale bars: 100μm on panels A to D, and 50μm on panels E to H. Mouse experiments were repeated three times with at least 3–4 mice per group in each experiment.