Preventive effects of bovine colostrum supplementation in TNBS-induced colitis in mice

Abstract

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder for which the current medical therapy is not completely effective. Bovine colostrum (BC) is a biological fluid rich in bioactive molecules that may have beneficial effects on several gastrointestinal disorders. The objectives of this study were to assess the preventive effects of BC supplementation in a mouse model of 2,4,6 trinitrobenzene sulfonic acid (TNBS)-induced colitis using a multidisciplinary approach. Specifically, the following parameters were evaluated: (i) disease activity index (DAI), (ii) histological score, (iii) expression levels of TLR4, anti- and pro-inflammatory cytokines, and (iv) count of some bacterial species of the intestinal microbiota. Mice received a daily suspension of BC (BC group, n = 12) or saline solution (control, CN group, n = 12) for 21 days before the intrarectal inoculation with 1% of TNBS solution. BC was well tolerated and did not induce any histological damage or clinical symptoms. After TNBS treatment, BC group showed a reduction of body weight (BW) loss (P<0.01) and histological score (P<0.05) compared to CN. Moreover, the expression levels of TLR4 (P<0.05), IL-1β (P<0.001), IL-8 (P<0.001), and IL-10 (P<0.001) were lower in mice administered with BC, while the concentrations of TNF-α did not show any differences between groups. Finally, the supplementation with BC resulted in a differential response to TNBS treatment in the bacterial count. In CN group, E. coli and Enterococci increased (P<0.001), while Anaerobes (P<0.01), Lactobacilli, and Bifidobacteria (P<0.001) reduced. Conversely, no significant changes in bacterial load were found after the inoculation of TNBS in BC pre-treated mice. This study confirms that TNBS-induced colitis model in mice is useful for studying the mechanisms involved in IBD pathogenesis and shows that pre-treatment with BC reduces the intestinal damages and clinical signs of the colitis. Molecular mechanisms and intestinal microflora could be involved in the protective effect of colostrum.

Fig 1. Colon histology before TNBS administration.

Comparative assessment of histological aspects from mice treated for 21 days with saline solution (A. General view; B. Detailed aspect; Histological score 2) and Bovine Colostrum (C. General view; D. Detailed aspect; Histological score 1) before the TNBS administration. Both sections revealed a normal structure of the colon with very mild inflammation of the mucosa. H&E stained, magnification of 4x (A-C) and 10x (B-D).

Fig 2. Colon histology after TNBS administration.

Comparative assessment of histological aspects of mice treated for 21 days with saline solution (E. General view; F. Detailed aspect; Histological score 4) and Bovine Colostrum (G. General view; H. Detailed aspect; Histological score 3) after the treatment with TNBS. The saline solution group (E-F) showed a severe architectural modification of the mucosa, with focal destruction of the epithelium and crypt loss associated with an inflammatory reaction characterized by severe infiltration of lymphocytes and polymorphonuclear cells (neutrophils and eosinophils granulocytes) in epithelium, lamina propria and submucosal layers. Inflammation also involves partially the muscular layer. Colon of Bovine Colostrum group (G-H) showed evidence of a mild to moderate architectural modification of the mucosa, ulceration of the epithelium, crypt loss accompanied by a mild inflammatory reaction with infiltration of lymphocytes, a few neutrophils and eosinophils granulocytes in lamina propria and submucosa layers. H&E stained, magnification of 4x (E-G) and 10x (F-H).

Fig 3. Impact of TNBS treatment on mRNA expression of TLR4 (2^^-ΔCt) in Control and Bovine Colostrum (BC) groups.

*P <0.05. Control vs BC group before (Pre) and after (Post) TNBS treatment.

Fig 4

Impact of TNBS treatment on mRNA expression of IL-1β, IL-8, IL-10, and TNF-α (2^^-ΔCt; Panel A-D) in control and Bovine Colostrum (BC) groups. ***P <0.001. Control vs BC group before (Pre) and after (Post) TNBS treatment.

Fig 5

Impact of TNBS treatment on E. coli, Enterococci, Anaerobes, Lactobacilli, and Bifidobacteria count (Panel A-E) in control and Bovine Colostrum groups. Values are mean±SE. *P<0.05, ***P <0.001. Control vs Bovine Colostrum group before (Pre) and after (Post) TNBS treatment.