Marine algal polysaccharides alleviate aflatoxin B1-induced bursa of Fabricius injury by regulating redox and apoptotic signaling pathway in broilers

Abstract

Aflatoxin B1 (AFB1) causes toxic effect and leads to organ damage in broilers. Marine algal polysaccharides (MAP) of Enteromorpha prolifera exert multiple biological activities, maybe have a potential detoxification effect on AFB1, but the related research in broilers is extremely rare. Therefore, the purpose of this study was to investigate whether MAPs can alleviate AFB1-induced oxidative damage and apoptosis of bursa of Fabricius in broilers. A total of 216 five-week-old male indigenous yellow-feathered broilers (with average initial body weight 397.35 ± 6.32 g) were randomly allocated to one of three treatments (6 replicates with 12 broilers per replicate), and the trial lasted 4 wk. Experimental groups were followed as basal diet (control group); basal diet mixed with 100 μg/kg AFB1 (AFB1 group, the AFB1 is purified form); basal diet with 100 μg/kg AFB1 + 2,500 mg/kg MAPs (AFB1 + MAPs group). The results showed that the diet with AFB1 significantly decreased the relative weight of bursa of Fabricius (P < 0.05), antioxidant enzymes activities of total superoxide dismutase (T-SOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutathione S-transferase (GST), and total antioxidation capacity (T-AOC), while increased malondialdehyde (MDA) content (P < 0.05). Besides, compared with AFB1 group, dietary MAPs improved the relative weight of bursa of Fabricius and activities of antioxidant enzymes (T-SOD, GSH-Px, CAT, GST) with decreased MDA contents (P < 0.05). Moreover, the consumption of AFB1 downregulated the mRNA expression of SOD1, SOD2, GSTA3, CAT1, GPX1, GPx3, GSTT1, Nrf2, HO-1, and p38MAPK (P < 0.05). Dietary MAPs upregulated the mRNA expression of SOD2, GSTA3, CAT1, GPX1, GSTT1, p38MAPK, Nrf2, and HO-1 in comparison with AFB1 group (P < 0.05). The histological analysis confirmed restoration of apoptotic cells of bursa of Fabricius (P < 0.01), which seen with MAPs supplemented broilers. Besides, dietary MAPs down-regulated the mRNA expression of caspase-3 and Bax (P < 0.05), while up-regulated the mRNA expression of Bcl-2 (P < 0.05) compared with AFB1 group. In addition, according to protein expression results, dietary MAPs up-regulated the protein expression level of antioxidant and apoptosis-associated proteins (Nrf2, HO-1, p38MAPK, Bcl-2) (P < 0.01), but down-regulated the protein expression level of caspase-3 and Bax (P < 0.01). In conclusion, dietary MAPs alleviated AFB1-induced bursa of Fabricius injury through regulating Nrf2-mediated redox and mitochondrial apoptotic signaling pathway in broilers.

Keywords: aflatoxin B1; broilers; bursa of Fabricius; marine algal polysaccharides.

Figure 1

Effect of dietary aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on relative weight of bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs or AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg AFB1 and 2,500 mg/kg MAPs. All data were expressed as means ± SE (n = 6). ∗ Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05).

Figure 2

Effects of dietary aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on histological changes of the bursa of Fabricius in broilers. (A) Control group, basal diet without addition of MAPs and AFB1; (B) AFB1 group, basal diet with addition of 100 μg/kg AFB1; (C) AFB1 + MAPs group, basal diet with addition of 100 μg/kg AFB1 and 2,500 mg/kg MAPs. The bursa of Fabricius sections were stained with hematoxylin and eosin. The scar bar is 50 μm of 200 × and 25 μm of 400×.

Figure 3

Effect of aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on antioxidant capacity of the bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs and AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg AFB1 and 2,500 mg/kg MAPs. (A) Total antioxidation capacity (T-AOC); (B) total superoxide dismutase (T-SOD) activities; (C) glutathione peroxidase (GSH-Px) activities; (D) catalase (CAT) activities in bursa of Fabricius; (E) glutathione S-transferase (GST) activities; (F) malondialdehyde (MDA) contents. All data were expressed as means ± SE (n = 6). ∗Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05; ∗∗P < 0.01).

Figure 4

Effect of marine algal polysaccharides (MAPs) on aflatoxin B1 (AFB1)-induced apoptosis of the bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs and AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg and 2,500 mg/kg MAPs. (A) TUNEL assay findings with immunofluorescence staining of apoptosis in bursa of Fabricius of broilers, all images were magnified 200 × ; (B) apoptosis rate of bursa of Fabricius in broilers. All data were expressed as means ± SE (n = 6). ∗Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05; ∗∗P < 0.01).

Figure 5

Effect of dietary aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on relative mRNA expression of antioxidant-related genes of the bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs and AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg and 2,500 mg/kg MAPs. (A) Relative mRNA expression of superoxide dismutase 1 (SOD1); (B) relative mRNA expression of superoxide dismutase 2 (SOD2); (C) relative mRNA expression of glutathione peroxidase 1 (GPx1); (D) relative mRNA expression of glutathione peroxidase 3 (GPx3); (E) relative mRNA expression of catalase 1 (CAT1); (F) relative mRNA expression of glutathione S-transferase theta 1 (GSTT1); (G) relative mRNA expression of glutathione S-transferase omega 1 (GSTO1); (H) relative mRNA expression of glutathione S-transferase alpha 3 (GSTA3); (I) relative mRNA expression of NF-E2-related factor 2 (Nrf2); (J) relative mRNA expression of heme oxygenase-1 (HO-1); (K) relative mRNA expression of p38 mitogen-activated protein (p38MAPK). All data were expressed as means ± SE (n = 6). ∗Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05; ∗∗P < 0.01).

Figure 6

Effect of dietary aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on relative mRNA expression of apoptosis-related genes of the bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs and AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg and 2,500 mg/kg MAPs. (A) Relative mRNA expression of Bcl-2-associated X (Bax); (B) relative mRNA expression of B-cell lymphoma-2 (Bcl-2); (C) relative mRNA expression of caspase-3; (D) relative mRNA expression of caspase-9. All data were expressed as means ± SE (n = 6). ∗Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05; ∗∗P < 0.01).

Figure 7

Effect of dietary aflatoxin B1 (AFB1) and marine algal polysaccharides (MAPs) on protein expression levels of antioxidant and apoptosis-associated molecules of bursa of Fabricius in broilers. CON, control group, basal diet without addition of MAPs and AFB1; AFB1, basal diet with addition of 100 μg/kg AFB1; AFB1 + MAPs, basal diet with addition of 100 μg/kg and 2,500 mg/kg MAPs. (A) Protein expression levels of NF-E2-related factor 2 (Nrf2); (B) protein expression levels of heme oxygenase-1 (HO-1); (C) protein expression levels of p38 mitogen-activated protein (p38MAPK); (D) protein expression levels of Bcl-2-associated X (Bax); (E) protein expression levels of B-cell lymphoma-2 (Bcl-2); (F) protein expression levels of Caspase-3. All data were expressed as means ± SE (n = 6). ∗Means with an asterisk indicates a significant difference from control by Tukey's test (∗P < 0.05; ∗∗P < 0.01).

Figure 8

Proposed mechanism of MAPs alleviates AFB1-induced bursa of Fabricius injury by regulating redox and apoptotic signaling pathway in broilers.