Diarrhoea caused by pathogens such as enterotoxigenic E. coli (ETEC) is a serious threat to the health of young animals and human infants. Here, we investigated the protective effect of fructo-oligosaccharides (FOS) on the intestinal epithelium with ETEC challenge in a weaned piglet model. Twenty-four weaned piglets were randomly divided into three groups: (1) non-ETEC-challenged control (CON); (2) ETEC-challenged control (ECON); and (3) ETEC challenge + 2·5 g/kg FOS (EFOS). On day 19, the CON pigs were orally infused with sterile culture, while the ECON and EFOS pigs were orally infused with active ETEC (2·5 × 109 colony-forming units). On day 21, pigs were slaughtered to collect venous blood and small intestine. Result showed that the pre-treatment of FOS improved the antioxidant capacity and the integrity of intestinal barrier in the ETEC-challenged pigs without affecting their growth performance. Specifically, compared with ECON pigs, the level of GSH peroxidase and catalase in the plasma and intestinal mucosa of EFOS pigs was increased (P < 0·05), and the intestinal barrier marked by zonula occluden-1 and plasmatic diamine oxidase was also improved in EFOS pigs. A lower level (P < 0·05) of inflammatory cytokines in the intestinal mucosa of EFOS pigs might be involved in the inhibition of TLR4/MYD88/NF-κB pathway. The apoptosis of jejunal cells in EFOS pigs was also lower than that in ECON pigs (P < 0·05). Our findings provide convincing evidence of possible prebiotic and protective effect of FOS on the maintenance of intestinal epithelial function under the attack of pathogens.

To explore the effect of manno-oligosaccharide (MOS) on intestinal health in weaned pigs upon enterotoxigenic Escherichia coli K88 (ETEC) challenge, thirty-two male weaned pigs were randomly assigned into four groups. Pigs fed with a basal diet or basal diet containing MOS (0·6 g/kg) were orally infused with ETEC or culture medium. Results showed that MOS significantly elevated the digestibility of crude protein and gross energy in both ETEC-challenged and non-challenged pigs (P < 0·05). MOS also elevated serum concentrations of IgA and IgM (P < 0·05), but decreased serum concentrations of TNF-α, IL-1β and IL-6 (P < 0·05) in ETEC-challenged pigs. Interestingly, MOS increased villus height and the ratio of villus height:crypt depth in duodenum and ileum (P < 0·05). MOS also increased duodenal sucrase and ileal lactase activity in ETEC-challenged pigs (P < 0·05). MOS decreased the abundance of E. coli, but increased the abundance of Lactobacillus, Bifidobacterium and Bacillus in caecum (P < 0·05). Importantly, MOS not only elevated the expression levels of zonula occludens-1 (ZO-1), claudin-1 and GLUT-2 in duodenum (P < 0·05) but also elevated the expression levels of ZO-1, GLUT-2 and L-type amino acid transporter-1 in ileum (P < 0·05) upon ETEC challenge. These results suggested that MOS can alleviate inflammation and intestinal injury in weaned pigs upon ETEC challenge, which was associated with suppressed secretion of inflammatory cytokines and elevated serum Ig, as well as improved intestinal epithelium functions and microbiota.

Here, we explored the influences of dietary inulin (INU) supplementation on growth performance and intestinal health in a porcine model. Thirty-two male weaned pigs (with an average body weight of 7·10 (sd 0·20) kg) were randomly assigned to four treatments and fed with a basal diet (BD) or BD containing 2·5, 5·0 and 10·0 g/kg INU. After a 21-d trial, pigs were killed for collection of serum and intestinal tissues. We show that INU supplementation had no significant influence on the growth performance in weaned pigs. INU significantly elevated serum insulin-like growth factor-1 concentration but decreased diamine oxidase concentration (P < 0·05). Interestingly, 2·5 and 5·0 g/kg INU supplementation significantly elevated the villus height in jejunum and ileum (P < 0·05). Moreover, 2·5 and 5·0 g/kg INU supplementation also elevated the villus height to crypt depth (V:C) in the duodenum and ileum and improved the distribution and abundance of tight-junction protein zonula occludens-1 in duodenum and ileum epithelium. INU supplementation at 10·0 g/kg significantly elevated the sucrase activity in the ileum mucosa (P < 0·05). INU supplementation decreased the expression level of TNF-α but elevated the expression level of GLUT 2 and divalent metal transporter 1 in the intestinal mucosa (P < 0·05). Moreover, INU increased acetic and butyric acid concentrations in caecum (P < 0·05). Importantly, INU elevated the Lactobacillus population but decreased the Escherichia coli population in the caecum (P < 0·05). These results not only indicate a beneficial effect of INU on growth performance and intestinal barrier functions but also offer potential mechanisms behind the dietary fibre-regulated intestinal health.

The present study aimed to investigate whether arginine (Arg) promotes porcine type I muscle fibres formation via improving mitochondrial biogenesis. In the in vivo study, a total of sixty Duroc × Landrace × Yorkshire weaning piglets with an average body weight of 6·55 (sd 0·36) kg were randomly divided into four treatments and fed with a basal diet or a basal diet supplemented with 0·5, 1·0 and 1·5 % l-Arg, respectively, in a 4-week trial. Results showed that dietary supplementation of 1·0 % Arg significantly enhanced the activity of succinate dehydrogenase, up-regulated the protein expression of myosin heavy chain I (MyHC I) and increased the mRNA levels of MyHC I, troponin I1, C1 and T1 (Tnni1, Tnnc1 and Tnnt1) in longissimus dorsi muscle compared with the control group. In addition, ATPase staining analysis indicated that 1·0 % Arg supplementation significantly increased the number of type I muscle fibres and significantly decreased the number of type II muscle fibres. Furthermore, 1·0 % Arg supplementation significantly up-regulated PPAR-γ coactivator-1α (PGC-1α), sirtuin 1 and cytochrome c (Cytc) protein expressions, increased PGC-1α, nuclear respiratory factor 1 (NRF1), mitochondria transcription factor B1 (TFB1M), Cytc and ATP synthase subunit C1 (ATP5G) mRNA levels and increased mitochondrial DNA content. In the in vitro study, mitochondrial complex I inhibitor rotenone (Rot) was used. We found that Rot annulled Arg-induced type I muscle fibres formation. Together, our results provide for the first time the evidence that Arg promotes porcine type I muscle fibres formation through improvement of mitochondrial biogenesis.

Muscle fibre types can transform from slow-twitch (slow myosin heavy chain (MyHC)) to fast-twitch (fast MyHC) or vice versa. Leucine plays a vital effect in the development of skeletal muscle. However, the role of leucine in porcine myofibre type transformation and its mechanism are still unclear. In this study, effects of leucine and microRNA-27a (miR-27a) on the transformation of porcine myofibre type were investigated in vitro. We found that leucine increased slow MyHC protein level and decreased fast MyHC protein level, increased the levels of phospho-protein kinase B (Akt)/Akt and phospho-forkhead box 1 (FoxO1)/FoxO1 and decreased the FoxO1 protein level. However, blocking the Akt/FoxO1 signalling pathway by wortmannin attenuated the role of leucine in porcine myofibre type transformation. Over-expression of miR-27a decreased slow MyHC protein level and increased fast MyHC protein level, whereas inhibition of miR-27a had an opposite effect. We also found that expression of miR-27a was down-regulated following leucine treatment. Moreover, over-expression of miR-27a repressed transformation from fast MyHC to slow MyHC caused by leucine, suggesting that miR-27a is interdicted by leucine and then contributes to porcine muscle fibre type transformation. Our finding provided the first evidence that leucine promotes porcine myofibre type transformation from fast MyHC to slow MyHC via the Akt/FoxO1 signalling pathway and miR-27a.

Leucine, one of the branched-chain amino acids, is the only amino acid to regulate protein turnover in skeletal muscle. Leucine not only increases muscle protein synthesis, but also decreases muscle protein degradation. It is well documented that leucine plays a positive role in differentiation of murine muscle cells. However, the role of leucine on porcine myoblast differentiation and its mechanism remains unclear. In this study, porcine myoblasts were induced to differentiate with differentiation medium containing different concentrations of leucine, and wortmannin was used to interdict the activity of protein kinase B (Akt). We found that leucine increased the number of myosin heavy chain-positive cells and creatine kinase activity. Moreover, leucine increased the mRNA and protein levels of myogenin and myogenic determining factor (MyoD). In addition, leucine increased the levels of phosphorylated Akt/Akt and phosphorylated Forkhead box O1 (P-FoxO1)/FoxO1, as well as decreased the protein level of FoxO1. However, wortmannin, a specific repressor of PI3K/Akt signalling pathway, attenuated the positive role of leucine on porcine myoblast differentiation. Our results suggest that leucine promotes porcine myoblast differentiation through the Akt/FoxO1 signalling pathway.

The intestine plays key roles in maintaining body arginine (Arg) homoeostasis. Meanwhile, the intestine is very susceptible to reactive oxygen species. In light of this, the study aimed to explore the effects of Arg supplementation on intestinal morphology, Arg transporters and metabolism, and the potential protective mechanism of Arg supplementation in piglets under oxidative stress. A total of thirty-six weaned piglets were randomly allocated to six groups with six replicates and fed a base diet (0·95 % Arg,) or base diet supplemented with 0·8 % and 1·6 % l-Arg for 1 week, respectively. Subsequently, a challenge test was conducted by intraperitoneal injection of diquat, an initiator of radical production, or sterile saline. The whole trial lasted 11 d. The diquat challenge significantly decreased plasma Arg concentration at 6 h after injection (P<0·05), lowered villus height in the jejunum and ileum (P<0·05) as well as villus width and crypt depth in the duodenum, jejunum and ileum (P<0·05). Oxidative stress significantly increased cationic amino acid transporter (CAT)-1, CAT-2 and CAT-3, mRNA levels (P<0·05), decreased arginase II (ARGII) and inducible nitric oxide synthase mRNA levels, and increased TNF-α mRNA level in the jejunum (P<0·05). Supplementation with Arg significantly decreased crypt depth (P<0·05), suppressed CAT-1 mRNA expression induced by diquat (P<0·05), increased ARGII and endothelial nitric oxide synthase mRNA levels (P<0·05), and effectively relieved the TNF-α mRNA expression induced by diquat in the jejunum (P<0·05). It is concluded that oxidative stress decreased Arg bioavailability and increased expression of inflammatory cytokines in the jejunum, and that Arg supplementation has beneficial effects in the jejunum through regulation of the metabolism of Arg and suppression of inflammatory cytokine expression in piglets.

In the present study, twenty-four Duroc × Landrance × Yorkshire (initial body weight (BW) of 21·82 (sem 2·06) kg) cross-bred pigs were used to determine whether dietary vitamin D supplementation could confer protection against viral infections through the retinoic acid-inducible gene I (RIG-I) signalling pathway in pigs. Experimental treatments were arranged in a 2 × 2 factorial manner with the main effects of immune challenge (control v. porcine rotavirus (PRV) challenge) and dietary concentrations of vitamin D (200 and 5000 IU; where 1 IU of vitamin D is defined as the biological activity of 0.025 mg of cholecalciferol). The pigs were fed a diet containing 200 or 5000 IU vitamin D in the first week of the study period. On day 8, the pigs were orally dosed with 4 ml of Dulbecco's modified Eagle's medium/Ham's F-12 medium containing PRV or essential medium (control). Serum samples were collected on day 8 (pre-challenge), and 6 d after the PRV challenge, the pigs were killed to evaluate intestinal morphology and tissue gene expression following the last blood collection. Pigs challenged with PRV had decreased BW gain (P< 0·01), feed intake (P< 0·01), villus height (P< 0·01), faecal consistency (P< 0·05), and serum 1,25-dihydroxyvitamin D concentration (P< 0·01) and increased (P< 0·01) serum IL-2, IL-6 and interferon (IFN)-β concentrations. Vitamin D supplementation mitigated these effects. The mRNA expression of RIG-I (P< 0·01), IFN-β promoter stimulator 1 (P< 0·01), IFN-β (P< 0·01) and interferon-stimulated gene 15 (ISG15) (P< 0·01) was up-regulated by the PRV challenge and vitamin D supplementation in the intestine. In conclusion, vitamin D supplementation could activate the RIG-I signalling pathway and thus alleviate the negative effects caused by PRV challenge.

Low birth weight (LBW) exerts persistent effects on the growth and development of offspring. The present study was conducted to test the hypothesis that LBW alters the response of pigs to high-fat (HF) diet-induced changes in meat quality and skeletal muscle proteome. Normal-birth weight (NBW) and LBW piglets were fed a control diet or a HF diet from weaning to slaughter at 110 kg body weight. Most of the meat quality traits were influenced by LBW. Meat quality analysis revealed that LBW piglets had a greater ability to deposit intramuscular lipids than their heavier littermates when fed a HF diet. Increased shear force, lower pH45min and drip loss were observed in the skeletal muscle of LBW piglets compared with NBW piglets. Proteomic analysis revealed forty-six differentially expressed proteins in the skeletal muscle of LBW and NBW piglets fed the control diet or HF diet. These proteins play a central role in cell structure and motility, glucose and energy metabolism, lipid metabolism, and cellular apoptosis, as well as stress response. Of particular interest is the finding that LBW altered the response to HF diet-induced changes in the expression of proteins related to stress response (heat shock protein) and glucose and energy metabolism (pyruvate kinase, phosphoglycerate mutase, enolase and triosephosphate isomerase). Taken together, our findings revealed that the HF diet-induced changes in the expression of glucose and energy metabolism-related proteins varied between NBW and LBW piglets, which provides a possible mechanism to explain higher intramuscular fat store in LBW pigs when fed a HF diet.

The objective of the present study was to evaluate the effects of fibre source on intestinal mucosal barrier function in weaning piglets. A total of 125 piglets were randomly allotted on the basis of their body weight and litters to one of five experimental diets, i.e. a control diet without fibre source (CT), and diets in which expanded maize was replaced by 10 % maize fibre (MF), 10 % soyabean fibre (SF), 10 % wheat bran fibre (WBF) or 10 % pea fibre (PF). The diets and water were fed ad libitum for 30 d. Piglets on the WBF and PF diets had lower diarrhoea incidence compared with the MF- and SF-fed animals. A higher ratio of villous height:crypt depth in the ileum of WBF-fed piglets and higher colonic goblet cells in WBF- and PF-fed piglets were observed compared with CT-, MF- and SF-fed piglets. In the intestinal digesta, feeding WBF and PF resulted in increased Lactobacillus counts in the ileum and Bifidobacterium counts in the colon. Lower Escherichia coli counts occurred in the ileum and colon of WBF-fed piglets than in SF-fed piglets. Tight junction protein (zonula occludens 1; ZO-1) and Toll-like receptor 2 (TLR2) gene mRNA levels were up-regulated in the ileum and colon of pigs fed WBF; however, feeding MF and SF raised IL-1α and TNF-α mRNA levels. Furthermore, higher diamine oxidase activities, transforming growth factor-α, trefoil factor family and MHC-II concentration occurred when feeding WBF and PF. In conclusion, the various fibre sources had different effects on the ileal and colonic barrier function. Clearly, WBF and PF improved the intestinal barrier function, probably mediated by changes in microbiota composition and concomitant changes in TLR2 gene expression.