Effects of microencapsulated blend of organic acids and botanicals on growth performance, intestinal barrier function, inflammatory cytokines, and endocannabinoid system gene expression in broiler chickens

Abstract

With restricted usage of growth-promoting antibiotics, identifying alternative feed additives that both improve intestinal barrier function and reduce inflammation is the center to improve chickens' health. This study examined the effects of a microencapsulated feed additive containing citric acid, sorbic acids, thymol, and vanillin on intestinal barrier function and inflammation status. A total of 240 birds were assigned to either a commercial control diet or control diet supplemented with 500 g/MT of the microencapsulated additive product. Birds were raised by feeding a 2-phase diet (starter, d 1 to d 21; and grower, d 15 to d 42). Growth performance was recorded weekly. At d 21 and d 42, total gastrointestinal tract permeability was evaluated by FITC-dextran (FD4) oral gavage. Jejunum-specific barrier functions were evaluated by Ussing chamber. Intestinal gene expression of selected epithelial cell markers, tight junction (TJ) proteins, inflammatory cytokines, and endocannabinoid system (ECS) markers were determined by RT-PCR. Statistical analysis was performed using Student t test. Results showed significant improvement of feed efficiency in the birds supplemented with the blend of organic acids and botanicals. At d 21, both oral and jejunal FD4 permeability were lower in the supplemented group. Jejunal transepithelial resistance was higher in the supplemented birds. At d 21, expression of TJs mRNA (CLDN1 and ZO2) was both upregulated in the jejunum and ileum of supplemented birds, while CLDN2 was downregulated in cecum. Proliferating cell marker SOX9 was higher expressed in jejunum and ceca. Goblet cell marker (MUC2) was upregulated, while Paneth cell marker (LYZ) was downregulated in the ileum. Proinflammatory cytokine expressions of IL1B, TNFA, and IFNG were downregulated in jejunum, while anti-inflammatory IL10 expression was higher in jejunum, ileum, cecum, and cecal tonsil. The ECS markers expressions were upregulated in most intestinal regions. Together, these results demonstrated that the blend of organic acids and botanical supplementation reduced inflammation, improved the TJs expression and intestinal barrier function, and thus improved chicken feed efficiency. The activated ECS may play a role in reducing intestinal tissue inflammation.

Keywords: botanical; endocannabinoid; gut barrier; organic acid; tight junction.

Figure 1

Intestinal barrier integrity of birds fed with control or supplemented diet. Serum FD4 concentration measured after FD4 oral gavage in birds at d 21 and d 42 of age (A). Transepithelial resistance (TER) of the jejunum mucosa of birds at d 21 and d 42 of age on Ussing chambers (B). Tissue FD4 flux rate of the jejunum mucosa of birds at d 21 and d 42 of age on Ussing chambers (C). Difference between 2 groups at each growth period was labeled. * indicates significant difference (P ≤ 0.05). Dietary treatments were as follows: Con, basal diet; OA+B, basal diet + 500 ppm encapsulated organic acids and botanicals.

Figure 2

Intestinal tight junction proteins mRNA expression of birds fed with control or supplemented diet. At d 21 of age, gene expressions in jejunum (A), ileum (B), and cecum (C). Difference between 2 groups at each growth period was labeled. * indicates significant difference (P ≤ 0.05). ^# indicates trend difference (0.05 < P ≤ 0.1). Dietary treatments were as follows: Con, basal diet; OA+B, basal diet + 500 ppm encapsulated organic acids and botanicals. Abbreviations: CLDN1, claudin 1; CLDN2, claudin 2; OCLN, occludin; TJP1, tight junction protein 1; ZO2, Zonula occludins 2.

Figure 3

Intestinal epithelial cell markers mRNA expression of birds fed with control or supplemented diet. At d 21 of age, gene expressions in jejunum (A), ileum (B), and cecum (C). Difference between 2 groups at each growth period was labeled. * indicates significant difference (P ≤ 0.05). ^# indicates trend difference (0.05 < P ≤ 0.1). Dietary treatments were as follows: Con, basal diet; OA+B, basal diet + 500 ppm encapsulated organic acids and botanicals. Abbreviations: CHGA, chromogranin A; LGR5, leucine-rich repeat-containing G-protein coupled receptor 5; LYZ, lysozyme; MUC2, mucin 2; SOX9, SRY-box transcription factor 9.

Figure 4

Intestinal cytokines mRNA expression of birds fed with control or supplemented diet. At d 21 of age, gene expressions in jejunum (A), ileum (B), cecum (C), and cecal tonsil (D). Difference between 2 groups at each growth period was labeled. * indicates significant difference (P ≤ 0.05). ^# indicates trend difference (0.05 < P ≤ 0.1). Dietary treatments were as follows: Con, basal diet; OA+B, basal diet + 500 ppm encapsulated organic acids and botanicals. Abbreviations: IFNG, interferon gamma; IL, interleukin; TNFA, tumor necrosis factor alpha.

Figure 5

Intestinal endocannabinoids system markers mRNA expression of birds fed with control or supplemented diet. At d 21 of age, gene expressions in jejunum (A), ileum (B), and cecum (C). Difference between 2 groups at each growth period was labeled. * indicates significant difference (P ≤ 0.05). ^# indicates trend difference (0.05 < P ≤ 0.1). Dietary treatments were as follows: Con, basal diet; OA+B, basal diet + 500 ppm encapsulated organic acids and botanicals. Abbreviations: CNR1, cannabinoid receptor 1; FAAH, fatty acid amide hydrolase; NAPEPLD, N-acylphosphatidylethanolamine-specific phospholipase D.