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. 2018 May 15;23(7):1962–1976. doi: 10.1016/j.celrep.2018.04.053

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© 2018 The Author(s)

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

PMC Copyright notice

Decreased NO Levels in ASL CKO Immune Cells

(A) Nitrite (NO₂⁻) levels were measured by dedicated high-performance liquid chromatography (HPLC) in blood samples before and after colitis induction. Results show a significant elevation in NO levels after colitis induction in RBCs of the control group but not in Vav1^Cre: Asl^f/f mice (n ≥ 3 in each group).

(B) A significant elevation in NO₂⁻ levels is seen in peritoneal macrophages growth medium 18 hr following LPS administration in the control group compared with the Vav1^Cre: Asl^f/f group (n = 6 in each group).

(C and D) A significant reduction in glucose levels (C) and elevation in lactate levels (D) in response to LPS administration is demonstrated in control but not in Vav1^Cre: Asl^f/f macrophages. Measurements were performed using a NOVA measuring instrument, (n ≥ 4 in each group).

(E) Immunohistochemistry staining for the total number of macrophages (using the F4-80 marker, top) as well as for activated macrophages (using the MAC2 marker of activation, bottom), in distal colon sections after colitis induction. The ratio between F4-80- and MAC2-positive cells shows a significant reduction in the number of activated macrophages in Vav1^Cre: Asl^f/f mice compared with controls. Cells were counted from at least 8 microscopic fields taken from 3 different animals. Right: quantification analysis.

(F and G) Colitis was induced by DSS in CD11c^Cre:Asl^f/f and in control Asl^f/f mice.

(F) Right: a representative graph showing that CD11c^Cre:Asl^f/f mice have decreased colitis severity compared with control mice, as demonstrated by endoscopic evaluation on day 12 after colitis induction. The experiment was performed on a total of n ≥ 13 mice in each group. Left: a representative image of the colon taken by colonoscopy.

(G) A representative graph showing a significant increase in colon length in CD11c^Cre:Asl^f/f mice compared with the control group. The experiment was performed on a total of n ≥ 10 mice in each group.

(H) ELISA measurement of colon tissues extracted from mice on day12 following colitis induction, showing significant elevation of pro-inflammatory cytokine levels in the control group compared with CD11c^Cre:Asl^f/f mice.

(I) Human data taken from a GEO dataset analysis (GEO: GSE57945; ID:200057945), showing significant upregulation of ASL and iNOS RNA expression in patients suffering from Crohn’s disease (CD) and ulcerative colitis (UC) compared with controls, with no significant change in ornithine decarboxylase (ODC) expression.

Error bars represent SEM.