Microbial-Driven Butyrate Regulates Jejunal Homeostasis in Piglets During the Weaning Stage - PubMed (original) (raw)

Microbial-Driven Butyrate Regulates Jejunal Homeostasis in Piglets During the Weaning Stage

Xi Zhong et al. Front Microbiol. 2019.

Abstract

Microbe-derived butyrate plays an important role in the gut health of young mammals during the weaning stage. A greater understanding of how butyrate regulates intestinal development is necessary for overcoming post-weaning diarrheal diseases. We aimed to investigate whether jejunal microbial metabolite butyrate modulates the apoptosis/proliferation balance and immune response in piglets during the post-weaning period of the first 3 weeks of life. On the one hand, during the first week post-weaning, the relative abundances of the dominant bacterial families Erysipelotrichaceae (P < 0.01) and Lachnospiraceae (P < 0.01) were increased, which induced decreases in both butyrate production (P < 0.05) and its receptor (G-protein coupled receptor 43) expression (P < 0.01). The resulting intestinal inflammation (inferred from increased TNF-α and IFN-γ expression) contributed to the onset of cell apoptosis and the inhibition of cell-proliferation along the crypt-villus axis, which were followed by impaired jejunal morphology (i.e., increased crypt-depth) (P < 0.05) and intestinal dysfunction (i.e., decreased creatine kinase, and lactate dehydrogenase) (P < 0.05). On the other hand, during the second week post-weaning, the relative abundances of Lactobacillaceae (P < 0.01) and Ruminococcaceae (P < 0.05) were increased. The increases were accompanied by increased butyrate production (P < 0.05) and its receptor expression (P < 0.01), leading to the inhibition of cell apoptosis and the stimulation of cell proliferation via decreased pro-inflammatory cytokines and thereby the improvement of intestinal development and function. Herein, this study demonstrates that microbial-driven butyrate might be a key modulator in the maintenance of intestinal homeostasis after weaning. The findings suggest that strategies to promote butyrate production can maintain the apoptosis/proliferation balance via minimizing intestinal inflammation, and thereby improving post-weaning jejunal adaptation toward gut health.

Keywords: Sus scrofa; apoptosis; butyrate; inflammatory response; jejunum; microbiota; proliferation; weaned.

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Figures

FIGURE 1

FIGURE 1

Jejunal microbial diversities and bacterial compositions of weaned piglets. Venn diagram showing the unique and shared operational taxonomic units (OTUs) (3% distance level) (A), core OTUs (>1.00% of the community) (B), estimated Shannon index (C), estimated Simpson index (D), and bacterial compositions at the family level (E) of jejunal samples from weaned piglets. OTUs, operational taxonomic units; PW, post-weaning. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 2

FIGURE 2

The core bacteria (at the family level) in the jejunum of weaned piglets. PW, post-weaning. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 3

FIGURE 3

The SCFAs and its receptors in the jejunum of weaned piglets. (A) The ratios of acetate, propionate, and butyrate to total SCFAs in the jejunum of piglets during the first 3 weeks post-weaning. (B) The mRNA expression of SCFAs receptors (e.g., GPR41 and GPR43) in the jejunum of weaned piglets. (C) Representative western blot and quantification of SCFAs receptors (e.g., GPR41 and GPR43) in the jejunum of weaned piglets. B-Actin was used as the loading control. SCFAs, short-chain fatty acids; PW, post-weaning; D0, post-weaning day 0; D7, post-weaning day 7; D14 = post-weaning day 14; D21, post-weaning day 21. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 4

FIGURE 4

Pro-inflammatory cytokines in the jejunum of weaned piglets. The mRNA expression (A) and protein levels (B) of pro-inflammatory cytokines (i.e., TNF-α, IFN-γ, IL-8, and IL-1β). PW, post-weaning. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 5

FIGURE 5

The proliferating cell nuclear antigen (PCNA) in the jejunum of weaned piglets. (A) Representative images used for the immunohistochemical detection of PCNA in the jejunum segment of piglets at post-weaning days 0, 7, 14, and 21. (B) The density of PCNA-positive cells in the jejunum of piglets at post-weaning days 0, 7, 14, and 21. PWD, post-weaning day; D0, post-weaning day 0; D7, post-weaning day 7; D14, post-weaning day 14; D21, post-weaning day 21. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 6

FIGURE 6

The signaling markers of cell proliferation along the jejunal crypt-villus axis of weaned piglets. (A) The mRNA expression of cell-proliferation makers (e.g., GLP-2R, EGF-R, and EGF). (B) Representative western blot and quantification of cell-proliferation markers (e.g., GLP-2R, EGF-R, EGF, and ErbB). B-Actin was used as the loading control. PWD, post-weaning day; D0, post-weaning day 0; D7, post-weaning day 7; D14, post-weaning day 14; D21, post-weaning day 21. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 7

FIGURE 7

The signaling markers of cell apoptosis along the jejunal crypt-villus axis of weaned piglets. (A) The mRNA expression of cell-apoptosis makers (e.g., Bcl-2 and Bax). (B) Representative western blot and quantification of cell-apoptosis makers (e.g., Bcl-2, Bax, active-Caspase 3, and active-Caspase 9). B-Actin was used as the loading control. PWD, post-weaning day; D0, post-weaning day 0; D7, post-weaning day 7; D14, post-weaning day 14; D21, post-weaning day 21. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 8

FIGURE 8

The jejunal morphology of weaned piglets. (A) Representative light micrographs of a cross-section of the jejunum of piglets at post-weaning days 0, 7, 14, and 21. Images of the whole slide were scanned by Mito More Than Microscopy, and images were then taken at 40× magnification using a light microscope. (B) The quantification of villus height, crypt-depth and the ratio of villus height/crypt-depth (IVR). PWD, post-weaning day; D0, post-weaning day 0; D7, post-weaning day 7; D14, post-weaning day 14; D21, post-weaning day 21. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 9

FIGURE 9

Digestive enzymes in the jejunum of weaned piglets during the first 3 weeks post-weaning. The mRNA expression (A) and activities (B) of digestive enzymes (e.g., AIP, CK, and LDH). PW, post-weaning; AIP, alkaline phosphatase; CK, creatine kinase; LDH, lactate dehydrogenase. Bars (mean ± SEM, n = 5) with different letters are considered significantly different (P < 0.05).

FIGURE 10

FIGURE 10

Schematic representation of microbial metabolite butyrate-induced intestinal adaptation via the immune response to weaning stress. (A) Weaning stress-induced intestinal dysfunction: at the first week post-weaning (transition from lactation to weaning), weaning stress induces significant alterations of core OTUs and alpha diversity, especially increases in two dominant taxonomic groups (i.e., Erysipelotrichaceae and Lachnospiraceae). These changes result in decreased butyrate production and decreased expression of its receptor (GPR-43), which contributes to the onset of apoptosis and the inhibition of proliferation along the crypt-villus axis via inducing increased pro-inflammatory cytokines. As a result, pronounced shifts of the intestinal structure and function toward post-weaning intestinal dysfunction (increased crypt-depth) occur, which significantly decrease digestive-enzyme activities (i.e., CK and LDH) and growth check (i.e., low feed intake and body weight). (B) Self-repairing mechanism-induced jejunal adaptation toward health status: at the second week post-weaning (transition from weaning to post-weaning), increasing the relative abundances of Lactobacillaceae and Ruminococcaceae are accompanied by decreased in core OTUs and the alpha diversity of bacteria community. These changes stimulate butyrate production and expression of its receptor, which lead to the inhibition of apoptosis and the stimulation of proliferation via decreased pro-inflammatory cytokines, thereby promoting the development of post-weaning intestinal environment toward health status. OTUs, operational taxonomic units; CK, creatine kinase; LDH, lactate dehydrogenase.

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