Effect of EGFL7 on GVHD severity in B6→B6D2F1 mice. (A) Weights of animals that had undergone transplantation were measured daily and averaged for the group (green circles: syngeneic T cells [no GVHD]; red triangles: allogeneic treated with PBS; blue squares: allogeneic treated with rEGFL7). Daily EGFL7 treatment was initiated at day +21 post-BMT. Data were pooled from 3 experiments with 6 to 13 mice per group. (B) Survival curve of transplanted mice (dotted green line: syngeneic control [SYN], blue line: allogeneic treated with rEGFL7, and thin dotted red line: allogeneic treated with PBS). Data were pooled from 3 experiments with 6 to 13 mice per group. (C) Clinical scores of GVHD + PBS, GVHD + rEGFL7, and syngeneic mice at day +28 post-BMT. (D) Histopathology of the gut 28 days after allo-HSCT. Left: magnification ×200 and right: magnification ×400. (E) Gastrointestinal (GI) histopathology score of the gut of GVHD + rEGFL7 and GVHD + PBS-treated mice. Results show mean ± standard error of the mean (SEM). (F) Immunofluorescence analysis of the intestine from PBS or rEGFL7-treated mice transplanted with allogenic splenocytes that were stained for immunofluorescence. Cells were stained with CD31 (secondary antibody: donkey anti-goat alexa fluor 488), CD45 (secondary antibody: donkey anti-rabbit alexa fluor 647), and Ki-67 antibodies (secondary antibody donkey anti-rat alexa fluor 594). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole (DAPI). Magnification ×400. (G) Cells were stained with CD3 (secondary antibody donkey anti-goat alexa fluor 488), CD4 (secondary antibody donkey anti-rabbit alexa fluor 647), and CD8 antibodies (secondary antibody donkey anti-rat alexa fluor 594). Nuclei were counterstained with 4′,6-diamidino-2-phenylindole. Magnification ×400. Data show 1 representative sample from each experimental group, which consists of 3 to 4 mice per group.