Fig. 4

solTNF neutralization with XPro1595 reduces colonic LCN2 and reverses tight junction protein alterations associated with HFHC diet consumption. a-c HFHC diet promoted and decrease in colon length (n = 12–13, diet effect P < 0.0001) and an inflammatory colonic profile demonstrated by elevated LCN2 in colon (n = 6–9, diet effect P = 0.0091) and feces (n = 8–10, diet effect P = 0.0252). d Colonic Il-1b mRNA expression (n = 6, diet effect P < 0.0001). XPro 1595 reverses colonic LCN2 (P = 0.0460). e solTNF neutralization and HFHC diet-associated alterations in proteins involved in barrier function in the colon (CLDN2 n = 6, diet effect P = 0.0091), solTNF inhibition decreased the ratio of claudin-2 high/low in the HFHC diet/XPro1595 group (CLDN2 n = 6, XPro effect P = 0.0425). f HFHC diet decreases small intestine length (n = 12–13, diet effect P < 0.0001). g-i Small intestinal barrier alterations in HFHC mice (Tjp1 mRNA expression, n = 6, diet effect P = 0.0012; TJP1 protein expression, n = 6, diet effect P = 0.05; Ocln mRNA, n = 6, diet effect P < 0.0001). j, l Muc (n = 6, diet effect, P < 0.0001) and Il-6 (n = 6, diet effect, P = 0.0004) mRNA expression in the small intestine. Tissues were analyzed by qPCR; for each animal, the Ct values were normalized to the Ct values for Gapdh and Ppia. The relative expression level of the target gene ratio of high molecular weight to low molecular weight forms of CLDN2 and TJP1 protein expression was assessed by immunoblot. Immunoblots are representative of two independent experiments. Band intensity was calculated using Image Studio Lite, and values were normalized to intensity of β-actin. Blot images were cropped for comparison. Data were analyzed by two-way ANOVA followed by Tukey’s multiple comparisons in GraphPad Prism 6. Bar height indicates mean of samples; error bars indicate standard error of the mean (SEM). Lowercase letters indicate post hoc analysis. Means with different lowercase letters are significantly different from each other, P < 0.05