Glycerol monolaurate attenuated immunological stress and intestinal mucosal injury by regulating the gut microbiota and activating AMPK/Nrf2 signaling pathway in lipopolysaccharide-challenged broilers

Abstract

This study was conducted to investigate the effects of glycerol monolaurate (GML) on lipopolysaccharide (LPS)-induced immunological stress and intestinal mucosal injury in broilers and its underlying mechanisms. A total of 144 one-d-old Arbor Acres broilers were allocated to a 2 × 2 factorial arrangement involving dietary treatment (0 or 1,200 mg/kg dietary GML) and LPS challenge (injected with saline or Escherichia coli LPS on d 16, 18, and 20). Samples were collected on d 21. The results revealed that dietary GML augmented serum immunoglobulin A (P = 0.009) and immunoglobulin G (P < 0.001) levels in challenged birds. Dietary GML normalized LPS-induced variations in serum interleukin-6, interferon-gamma, and LPS levels (P < 0.05), jejunal villus height (P = 0.030), and gene expression of interleukin-6, macrophage inflammatory protein-3 alpha, Toll-like receptor 4, nuclear factor kappa-B, caspase-1, tight junction proteins, adenosine monophosphate-activated protein kinase alpha 1 (AMPKα1), nuclear factor-erythroid 2-related factor 2 (Nrf2), and superoxide dismutase-1 (P < 0.05). GML supplementation ameliorated LPS-induced peroxidation by reducing malondialdehyde content and increasing antioxidant enzyme activity (P < 0.05). Dietary GML enhanced the abundances of Anaerostipes, Pseudoflavonifractor, and Gordonibacter and reduced the proportion of Phascolarctobacterium in challenged birds. Dietary GML was positively correlated with alterations in antioxidant enzyme activities and AMPKα1, Nrf2, and zonula occludens-1 expressions. The genera Anaerostipes, Lachnospira, Gordonibacter, Lachnospira, Marvinbryantia, Peptococcus, and Pseudoflavonifractor were linked to attenuated inflammation and improved antioxidant capacity of challenged birds. In conclusion, dietary GML alleviated LPS-induced immunological stress and intestinal injury of broilers by suppressing inflammation and oxidative stress. Dietary GML regulated cecal microbiota and activated the AMPK/Nrf2 pathway in LPS-challenged broilers.

Keywords: Adenosine monophosphate-activated protein kinase; Antioxidation; Glycerol monolaurate; Gut microbiota; Inflammation; Lipopolysaccharide challenge.

Fig. 1

Effects of dietary glycerol monolaurate on cecal microbiota diversity of lipopolysaccharide-challenged broilers. Veen diagram (A); PCoA based on unweighted Unifrac distance (B); ANOSIM based on unweighted Unifrac distance (C). ^a-b Means with no common superscripts differ significantly (P < 0.05). Means are based on 6 replicates per treatment with 1 bird per replicate. CON = control, birds fed a basal diet; GML = glycerol monolaurate, birds fed a basal diet containing 1,200 mg/kg GML; LPS = lipopolysaccharide, birds fed a basal diet and challenged with LPS; LPS + GML = birds fed a basal diet containing 1,200 mg/kg GML and challenged with LPS; ANOSIM = analysis of similarities.

Fig. 2

Effects of dietary glycerol monolaurate on cecal microbiota composition of lipopolysaccharide-challenged broilers. Microbial composition at the phylum and family levels (A, B); relative abundances analysis (C). ^a-c Means with no common superscripts differ significantly (P < 0.05). Means are based on 6 replicates per treatment with 1 bird per replicate. CON = control, birds fed a basal diet; GML = glycerol monolaurate, birds fed a basal diet containing 1,200 mg/kg GML; LPS = lipopolysaccharide, birds fed a basal diet and challenged with LPS; LPS + GML = birds fed a basal diet containing 1,200 mg/kg GML and challenged with LPS.

Fig. 3

Alteration of the cecal microbiota at the genus level. Relative abundances of the top 20 bacterial genera (A); LEfSe of the cecal microbiota (B); relative abundances analysis (C). ^a-c Means with no common superscripts differ significantly (P < 0.05). Means are based on 6 replicates per treatment with 1 bird per replicate. CON = control, birds fed a basal diet; GML = glycerol monolaurate, birds fed a basal diet containing 1,200 mg/kg GML; LPS = lipopolysaccharide, birds fed a basal diet and challenged with LPS; LPS + GML = birds fed a basal diet containing 1,200 mg/kg GML and challenged with LPS.

Fig. 4

Correlation analysis. The correlation between factors and samples distribution in serum (A) and jejunum (B) by redundancy analysis; heatmap of correlations determined by Spearman analysis among the cecal microbiota, inflammatory factors, tight junctions, antioxidant enzymes, and intestinal barrier parameters (C). ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001. LPS = lipopolysaccharide; MDA = malondialdehyde; IL-6 = interleukin 6; T-AOC = total antioxidant capacity; CAT = catalase; IFN-γ = interferon gamma; IgG = immunoglobulin G; GSH-px = glutathione peroxidase; IgA = immunoglobulin A; CON = control, birds fed a basal diet; GML = glycerol monolaurate, birds fed a basal diet containing 1,200 mg/kg GML; LPS = birds fed a basal diet and challenged with LPS; LPS + GML = birds fed a basal diet containing 1,200 mg/kg GML and challenged with LPS NF-κB = nuclear factor kappa-B; TLR4 = Toll-like receptor; OCLN = occludin; SOD1 = superoxide dismutase 1; CLDN2 = claudin-2; ZO-1 = zonula occludens-1; AMPKα1 = adenosine monophosphate-activated protein kinase alpha 1; Nrf2 = nuclear factor-erythroid 2-related factor 2; J T-AOC = total antioxidant capacity in jejunum; S T-AOC = total antioxidant capacity in serum.