Dietary isobutyric acid supplementation improves intestinal mucosal barrier function and meat quality by regulating cecal microbiota and serum metabolites in weaned piglets

Abstract

This study aimed to provide evidence for the effects of isobutyric acid on the intestinal mucosal barrier and gut microbiota in weaned piglets. In this study, 30 piglets were divided into two groups: one group was fed a standard diet (CON group), and the other group was fed a diet supplemented with 0.5% isobutyric acid (IB group) for 21 days. The results showed that isobutyric acid significantly increased (p < 0.05) serum immunity and antioxidant capacity in weaned piglets. In small intestine of piglets, the ratio of villus height to crypt depth was significantly increased (p < 0.05). Administration of isobutyric acid also increased (p < 0.05) the expression of genes related to intestinal mucosal barrier function. Cecal microbiota analysis revealed that isobutyric acid significantly increased (p < 0.05) the abundance of the Eubacterium coprostanoligenes group. Untargeted serum metabolomics analysis indicated that the top three categories of metabolites were lipids and lipid-like molecules, organic acids and derivatives, and organic heterocyclic compounds. Additionally, in longissimus thoracis muscle, isobutyric acid significantly increased (p < 0 0.05) intramuscular fat and triglyceride content compared with the CON group. Overall, isobutyric acid can improve small intestinal mucosal barrier function, and may influence the fat deposition through the regulation of serum metabolites in weaned piglets.

Keywords: gut microbiota; intestinal mucosal barrier function; intramuscular fat; isobutyric acid; serum metabolome; weaned piglets.

Figure 1

Effect of isobutyric acid on immunoglobulin level and antioxidant capacity of weaned piglets. (A) Immunoglobulin A (Ig A); (B) Immunoglobulin G (Ig G); (C) Immunoglobulin M (Ig M); (D) Total antioxidant capacity (T-AOC); (E) Superoxide dismutase (SOD); (F) Glutathione S-transferase (GST); (G) Malondialdehyde (MDA).* p < 0.05 and ** p < 0.01.

Figure 2

Effect of isobutyric acid on small intestinal mucosal barrier function. (A) HE staining shows the morphology of various segments of the intestine; (B) Villus height of the duodenum, jejunum and ileum; (C) Crypt depth of the duodenum, jejunum and ileum; (D) Ratio of villus height to crypt depth of the duodenum, jejunum and ileum; (E) Number of goblet cells of the duodenum, jejunum and ileum; (F) The relative mRNA expression levels of Claudin-1, Occludin and ZO-1 in duodenum between the CON and IB groups; (G) Jejunum; (H) Ileum; (I) The relative mRNA expression levels of MUC2 in duodenum, jejunum and ileum. * p < 0.05 and ** p < 0.01.

Figure 3

Effect of isobutyric acid on the expression of genes and proteins involved in lipid metabolism. (A) Effect of isobutyric acid on the relative mRNA expression of lipid synthesis; (B) Effect of isobutyric acid on the relative mRNA expression of lipid breakdown and transport; (C) Western blot showing the protein level; (D) Bar plots showing the densitometric values. * p < 0.05 and ** p < 0.01.

Figure 4

Diversity analysis between CON and IB group. (A) Principal component analysis (PCA); (B) Venn at the genus level; (C) Venn at the species level; (D) The proportion of bacteria at the phylum level; (E) Linear discriminant analysis Effect Size (LEfSe); (F) The proportion of the top 20 bacteria at the genus level; (G) The top 10 bacterial abundance at the genus level in boxplot.

Figure 5

Correlation analysis on cecal microbiota. (A) Heat map of the correlation between the differential bacteria and intestinal morphology in duodenum; (B) Heat map of the correlation between the differential bacteria and intestinal morphology in jejunum; (C) Heat map of the correlation between the differential bacteria and intestinal morphology in ileum; (D) Heat map of the correlation between the differential bacteria and lipid traits in LT.

Figure 6

Effect of isobutyric acid on serum metabolites. (A) OPLS-DA; (B) Volcano plot of serum metabolites; (C) Donut plot of metabolite classification and proportion; (D) Heatmap of the top 20 serum metabolites; (E) The top six serum metabolites in boxplot. (F) KEGG enrichment bubble for differential metabolites; (G) Differential abundance score for differential metabolites; (H) Network analysis for differential metabolites.

Figure 7

Weighted gene co-expression network analysis (WGCNA). (A) Scale independence (left) and mean connectivity (right) analysis for choosing the soft threshold in establishing the WGCNA network; (B) Gene clustering and identification of gene modules using WGCNA; (C) Heatmap of the relationship between gene modules and lipid traits. The relationships were assessed by calculating the Pearson correlation coefficients between the module eigengenes and lipid traits (IMF and TG); (D) Four core modules that are highly correlated with lipid traits; (E) Heatmap of the relationship between serum metabolites and lipid traits; (F) Venn of modules and association analysis.

Figure 8

Effect of isobutyric acid on intestinal mucosal barrier function and meat quality of weaned piglets.