A Diet Fortified with Anthocyanin-Rich Extract (RED) Reduces Ileal Inflammation in a Senescence-Prone Mice Model of Crohn's-Disease-like Ileitis

Abstract

SAMP mice develop progressive Crohn's disease (CD)-like ileitis without spontaneous colitis that worsens over time without chemical, genetic, or immunological manipulation. Even growing in an identical vivarium and fed with the same diet, SAMP mice reveal a distinct fecal microbiome, metabolome, and lipidome profile compared to AKR mice, their non-inflamed parental control strain. Differences are already present in 5-week-old mice, with a tendency to increase in 15-week-old mice. SAMP and AKR mice metabolome and lipidome profiles were substantially different, belonging to two clusters in line with the progression of intestinal disease. Similarly, the 16S analysis confirmed differences between 15-week-old AKR and SAMP mice. The protective role of dietary polyphenols has been documented in inflammatory bowel diseases (IBD); thus, we supplemented the chow diet with an anthocyanin-rich extract (RED) to evaluate disease reduction in SAMP mice and changes in fecal microbiota/metabolome. Our data reveal that 10-week supplementation with anthocyanin-rich extract ameliorated disease severity in SAMP mice despite limited fecal microbiota/metabolome differences.

Keywords: Crohn’s disease; adjuvant therapy; metabolome; microbiota; polyphenols.

Figure 1

Shannon index and PCA plot for 5-week-old SAMP and AKR mice (A). Stacked bar plots for representative genera in AKR and SAMP mice (B). Dot plots for the significant genera found in 5-week-old AKR and SAMP mice (C,D) (* p-value < 0.05, ** p-value < 0.005, *** p-value < 0.001, and **** p-value < 0.0001). Data expressed as mean ± SEM (n = 16).

Figure 2

Shannon index and PCA plot for 15-week-old SAMP and AKR mice (A). Stacked bar plots for representative genera in the two strains (B). Dot plots for the significant genera found in 15-week-old SAMP and AKR mice (C,D) (* p-value < 0.05, *** p-value < 0.001, and **** p-value < 0.0001). Data expressed as mean ± SEM (n = 3 for AKR and 8 for SAMP mice).

Figure 3

COMDIM score plots for the metabolome and lipidome of 5-week-old SAMP and AKR mice (A,B) and respective heatmaps showing the top 50 significantly modulated metabolites and lipids (p-value < 0.05) (C,D).

Figure 4

Quantitative enrichment analysis (QEA) overview presenting the top 25 related metabolic pathways ranked according to the fold enrichment p-value.

Figure 5

In vivo treatment of SAMP and AKR mice with RED. Five-week-old SAMP mice were administered with RED powder (53 mg/kg) or vehicle for 70 days; AKR mice were used as parental controls for each condition. Weight was measured each week for all the treatment (10 weeks) in AKR (A) and SAMP (B) mice. Representative H&E-stained ilea from AKR (left panel) and SAMP (right panel) mice treated with vehicle (top row) or RED (top row) (C). Inflammatory score of ileal inflammation (D) and colon inflammation (E) was calculated for all the experimental groups, as well as colon length/BW (F) and colon weight/BW (G) ratio percentage measured at the time of sacrifice, and TLOs count (H). Representative TLO stereomicroscopy of RED-treated SAMP mice and its respective H&E (I). Magnification: 10×. Abbreviation: BW: body weight. (* p-value < 0.05 and ** p-value < 0.005). Data expressed as mean ± SEM (n = 3 for AKR vehicle, 4 SAMP vehicle, 8 AKR RED, and 8 SAMP RED).

Figure 6

Shannon index and PCA plot for 15-week-old AKR and SAMP mice after RED treatment (A). Stacked bar plots for representative genera in the two strains (B). Dot plots for the significant genera modulated by RED treatment in 15-week-old SAMP and AKR mice (C,D) (* p-value < 0.05 and ** p-value < 0.005). Data expressed as mean ± SEM (n = 3 for AKR vehicle, 4 SAMP vehicle, 8 AKR RED, and 8 SAMP RED).