Ruilin Huang

Effects of dietary supplementation with an expressed fusion peptide bovine lactoferricin-lactoferrampin on performance, immune function and intestinal mucosal morphology in piglets weaned at age 21 d

Lactoferrin has antimicrobial activity associated with peptide fragments lactoferricin (LFC) and lactoferrampin (LFA) released on digestion. These two fragments have been expressed in Photorhabdus luminescens as a fusion peptide linked to protein cipB. The construct cipB-LFC-LFA was tested as an alternative to antimicrobial growth promoters in pig production. Sixty piglets with an average live body weight of 5.42 (sem 0.59) kg were challenged with enterotoxigenic Escherichia coli and randomly assigned to four treatment groups fed a maize-soyabean meal diet containing either no addition (C), cipB at 100 mg/kg (C+B), cipB-LFC-LFA at 100 mg/kg (C+L) or colistin sulfate at 100 mg/kg (C+CS) for 3 weeks. Compared with C, dietary supplementation with C+L for 3 weeks increased daily weight gain by 21 %, increased recovery from diarrhoea, enhanced serum glutathione peroxidase (GPx), peroxidase (POD) and total antioxidant content (T-AOC), liver GPx, POD, superoxide dismutase and T-AOC, Fe, total Fe-binding capacity, IgA, IgG and IgM levels (P < 0.05), decreased the concentration of E. coli in the ileum, caecum and colon (P < 0.05), increased the concentration of lactobacilli and bifidobacteria in the ileum, caecum and colon (P < 0.05), and promoted development of the villus-crypt architecture of the small intestine. Growth performance was similar between C+L- and C+CS-supplemented pigs. The present results indicate that LFC-LFA is an effective alternative to the feed antibiotic CS for enhancing growth performance in piglets weaned at age 21 d.

Dietary l -arginine supplementation enhances intestinal development and expression of vascular endothelial growth factor in weanling piglets

Oral administration of L-arginine has been reported to prevent gut disease in human infants. However, little is known about the effects of dietary arginine supplementation on intestinal development of weaned piglets. In the present study, twenty 21-d-old castrated piglets with 5·3 (SEM 0·13) kg body weight (BW) were weaned from sows, individually housed and randomly assigned to one of the two maize- and soyabean meal-based diets supplemented with 0 or 1% L-arginine. After consuming the diets for 7 d, six pigs were randomly selected from each group to obtain various tissues. Compared with control pigs, dietary supplementation with 1% L-arginine did not affect feed intake but enhanced (P<0·05) the relative weight of the small intestine (+33 %), daily BW gain (+38 %) and feed efficiency (+28 %). The villus height of the duodenum, jejunum and ileum in arginine-supplemented piglets was 21, 28 and 25% greater (P<0·05) than in the nonsupplemented control group. Arginine supplementation increased (P<0·05) protein levels for vascular endothelial growth factor(VEGF) in duodenal, jejunal and ileal mucosae by 14, 39 and 35 %, respectively. Compared with the control group, dietary supplementation with 1% L-arginine increased (P<0·05) plasma concentrations of arginine and insulin (+36 %), and decreased (P<0·05) plasma concentrations of cortisol (233 %), NH3 (221 %) and urea (219 %). These results indicate that arginine supplementation enhances intestinal growth, development and expression of VEGF in early-weaned pigs fed a maize- and soyabean meal-based diet. The findings may have important implications for neonatal pigs under stressful or diseased conditions.

Reduced expression of intestinal N-acetylglutamate synthase in suckling piglets: a novel molecular mechanism for arginine as a nutritionally essential amino acid for neonates

The objective of this study was to determine developmental changes in mRNA and protein levels for N-acetylglutamate synthase (NAGS; a key enzyme in synthesis of citrulline and arginine from glutamine/glutamate and proline) in the small intestine of suckling piglets. The porcine NAGS gene was cloned using the real-time polymerase-chain reaction (RT-PCR) method. The porcine NAGS gene encoded 368 amino acid residues and had a high degree of sequence similarity to the “conserved domain” of human and mouse NAGS genes. The porcine NAGS gene was expressed in E. coli BL21 and a polyclonal antibody against the porcine NAGS protein was developed. Real-time RT-PCR and western-blot analyses were performed to quantify NAGS mRNA and protein, respectively, in the jejunum and ileum of 1- to 28-day-old pigs. Results indicated that intestinal NAGS mRNA levels were lower in 7- to 28-day-old than in 1-day-old pigs. Immunochemical analysis revealed that NAGS protein was localized in enterocytes of the gut. Notably, intestinal NAGS protein abundance declined progressively during the 28-day suckling period. The postnatal decrease in NAGS protein levels was consistent with the previous report of reduced NAGS enzymatic activity as well as reduced synthesis of citrulline and arginine in the small intestine of 7- to 28-day-old pigs. Collectively, these results suggest that intestinal NAGS expression is regulated primarily at the post-transcriptional level. The findings also provide a new molecular basis to explain that endogenous synthesis of arginine is impaired in sow-reared piglets and arginine is a nutritionally essential amino acid for the neonates.

Molecular cloning, tissue distribution and ontogenetic expression of the amino acid transporter b0,+ cDNA in the small intestine of Tibetan suckling piglets

The small intestine is the main absorption place of peptides and free amino acids in mammals. The amino acid transporter system b(0,+) mediates apical uptake of basic amino acids, especially lysine, arginine and cysteine. The aim of the current study was to clone Tibetan porcine amino acid transporter b(0,+)AT (SLC7A9) for comparing the sequences of Tibetan and common (Sus scrofa) pigs, and investigating the tissue distribution and ontogenetic expression in the small intestine of Tibetan suckling piglets. The Tibetan porcine SLC7A9 gene was first cloned from the porcine small intestine and found to encode the amino acid transporter b(0,+)AT. The entire open reading frame (ORF) of the SLC7A9 is 1464 bp and codes for 487 amino acid residues, with a higher degree of sequence similarity with common pig (99.59%) and horse counterparts (91.2%) than with monkey (89.5%) or human (88.7%). The deduced protein has 12 putative transmembrane domains. In this study, SLC7A9 mRNA was detected in brain, kidney, duodenum, jejunum, ileum, heart, liver, lung and muscle from Tibetan pigs at 7 and 21 days by PCR. We also investigated the age-dependent expression of b(0,+)AT in Tibetan suckling piglets in duodenum, anterior jejunum, posterior jejunum, ileum and kidney from day 1 to 35. The abundance of SLC7A9 mRNA in duodenum and jejunum was highest and lowest, respectively. Expression patterns were similar in duodenum and anterior jejunum, where the mRNA level was decreased before the suckling period and increased until day 35. Posterior jejunum expression was increasing steadily with age, except on day 7. The ileum has the highest expression at day 14 and became steady after day 28. The mRNA abundance in the kidney is opposite to duodenum, increasing until day 14 and reducing thereafter. Our results showed the pattern of b(0,+)AT expressed in small intestine of Tibetan pig and lay the foundation for in depth investigations of the regulation of b(0,+)AT in vivo.

Effects of Dietary Arginine and Glutamine on Alleviating the Impairment Induced by Deoxynivalenol Stress and Immune Relevant Cytokines in Growing Pigs

Deoxynivalenol (DON) is a mycotoxin that reduces feed intake and animal performance, especially in swine. Arginine and glutamine play important roles in swine nutrition. The objective of this study was to determine the effects of dietary supplementation with arginine and glutamine on both the impairment induced by DON stress and immune relevant cytokines in growing pigs. A total of forty 60-d-old healthy growing pigs with a mean body weight of 16.28±1.54 kg were randomly divided into 5 groups, and assigned to 3 amino acid treatments fed 1.0% arginine (Arg), 1.0% glutamine (Gln) and 0.5% Arg+0.5% Gln, respectively, plus a toxin control and a non-toxin control. Pigs in the 3 amino acid treatments were fed the corresponding amino acids, and those in non-toxin control and toxin control were fed commercial diet with 1.64% Alanine as isonitrogenous control for 7 days. The toxin control and amino acid treatments were then challenged by feeding DON-contaminated diet with a final DON concentration of 6 mg/kg of diet for 21 days. No significant differences were observed between toxin control and the amino acid groups with regard to the average daily gain (ADG), although the values for average daily feed intake (ADFI) in the amino acid groups were significantly higher than that in toxin control (P<0.01). The relative liver weight in toxin control was significantly greater than those in non-toxin control, arginine and Arg+Glu groups (P<0.01), but there were no significant differences in other organs. With regard to serum biochemistry, the values of BUN, ALP, ALT and AST in the amino acid groups were lower than those in toxin control. IGF1, GH and SOD in the amino acid groups were significantly higher than those in toxin control (P<0.01). The IL-2 and TNFα values in the amino acid groups were similar to those in non-toxin control, and significantly lower than those in toxin control (P<0.01). These results showed the effects of dietary supplementation with arginine and glutamine on alleviating the impairment induced by DON stress and immune relevant cytokines in growing pigs.

Chemerin regulates proliferation and differentiation of myoblast cells via ERK1/2 and mTOR signaling pathways.

Obesity in human is an alarming major public health crisis worldwide and insulin resistance is a hallmark of it. The negative cross-talk between skeletal muscle and adipose tissue through adipokines is now accepted as one of the leading cause of insulin resistance. Chemerin is a novel adipokine previously reported to induce insulin resistance in primary human skeletal muscle cells. To investigate the role of chemerin in myogenesis, C2C12 cells were used and treated with chemerin in proliferation and differentiation stages. Our results showed that chemerin promoted proliferation and suppressed differentiation of C2C12 cells through extracellular-signal regulated kinase-1/2 (ERK1/2) and mammalian target of rapamycin (mTOR) signaling pathways, and these two pathways were interacted with each other in C2C12 cells treated with chemerin. It is concluded from this in vitro study that chemerin which expression is increased during myoblast differentiation appears to be able, likely in an autocrine/paracrine manner, to increase myoblast proliferation and decrease myoblast differentiation.

Physicochemical Properties and In Vitro Starch Digestibility of Cooked Rice from Commercially Available Cultivars in Canada

ABSTRACTThe influence of amylose content, cooking, and storage on starch structure, thermal behaviors, pasting properties, and rapidly digestible starch (RDS), slowly digestible starch (SDS), and resistant starch (RS) in different commercial rice cultivars was investigated. Long grain rice with high-amylose content had a higher gelatinization temperature and a lower gelatinization enthalpy than the other rice cultivars with intermediate amylose content (Arborio and Calrose) and waxy type (glutinous). The intensity ratio of 1047/1022 cm–1 determined by Fourier Transform Infrared (FT-IR), which indicated the ordered structure in starch granules, was the highest in glutinous and the lowest in long grain. Results from Rapid ViscoAnalyser (RVA) showed that the rice cultivar with higher amylose content had lower peak viscosity and breakdown, but higher pasting temperature, setback, and final viscosity. The RDS content was 28.1, 38.6, 41.5, and 57.5% in long grain, Arborio, Calrose, and glutinous rice, respectiv...

Chitosan Oligosaccharide Reduces Intestinal Inflammation That Involves Calcium-Sensing Receptor (CaSR) Activation in Lipopolysaccharide (LPS)-Challenged Piglets.

Chitosan oligosaccharide (COS) is a degradation product of chitosan with antioxidative, anti-inflammatory, and antibacterial effects. This study was conducted to investigate the effects of dietary COS on the intestinal inflammatory response and the calcium-sensing receptor (CaSR) and nuclear transcription factor kappa B (NF-κB) signaling pathways that may be involved using a lipopolysaccharide (LPS)-challenged piglet model. A total of 40 weaned piglets were used in a 2 × 2 factorial design; the main factors were dietary treatment (basal or 300 μg/kg COS) and inflammatory challenge (LPS or saline). On the morning of days 14 and 21 after the initiation of treatment, the piglets were injected intraperitoneally with Escherichia coli LPS at 60 and 80 μg/kg body weight or the same amount of sterilized saline, respectively. Blood and small intestine samples were collected on day 14 or 21, respectively. The results showed that piglets challenged with LPS have a significant decrease in average daily gain and gain:feed and histopathological injury in the jejunum and ileum, whereas dietary supplementation with COS significantly alleviated intestinal injury induced by LPS. Piglets fed the COS diet had lower serum concentrations of tumor necrosis factor alpha (TNF-α), interleukin (IL) 6, and IL-8 as well as lower intestinal abundances of pro-inflammatory cytokine mRNA but higher anti-inflammatory cytokine mRNA compared with piglets fed the basal diet among LPS-challenged piglets (p < 0.05). Dietary COS increased intestinal CaSR and PLCβ2 protein expressions in both saline- and LPS-treated piglets, but decreased p-NF-κB p65, IKKα/β, and IκB protein expressions in LPS-challenged piglets (p < 0.05). These findings indicate that COS has the potential to reduce the intestinal inflammatory response, which is concomitant with the activation of CaSR and the inhibition of NF-κB signaling pathways under an inflammatory stimulus.

Dietary supplementation with proline confers a positive effect in both porcine circovirus-infected pregnant and non-pregnant mice

Porcine circovirus type 2 (PCV2) is associated with various diseases that impose a significant economic burden on the swine industry. We hypothesised that nutritional supplementation with proline to enhance the immune response might be a useful prophylactic measure against PCV2 infection. To test this hypothesis, in the present study, we measured clinical data, including blood parameters, serum cytokine profile, PCV2 virus load in organs and serum, and microscopic lesions in the lung, liver and spleen, in both PCV2-infected pregnant and non-pregnant mice. Dietary supplementation with proline had no effect (P>0·05) on abortion rates in PCV2-infected pregnant mice, although a numerically lower abortion rate (22·2 v. 44·4%) was observed compared with the control. Dietary supplementation with proline significantly increased serum C-reactive protein levels (P= 0·03) in PCV2-infected pregnant mice, and increased serum TNF-α levels (P= 0·01), leucocytes (P< 0·05), lymphocytes (P< 0·05) and neutrophilic granulocytes (P< 0·05) in PCV2-infected non-pregnant mice. Meanwhile, dietary proline significantly (P< 0·05) decreased the PCV2 virus load in the lung. Furthermore, mice in the dietary proline group showed a significant (P< 0·01) decrease in microscopic lesion scores in the lung, liver and spleen compared with those in the alanine group. Collectively, dietary proline supplementation confers a functional role in PCV2-infected mice.

Acute and sub-acute oral toxicological evaluations and mutagenicity of N-carbamylglutamate (NCG).

N-carbamylglutamate (NCG) is a metabolically stable analog of N-acetylglutamate that activates carbamyl phosphate synthase-1, a key arginine synthesis enzyme in enterocytes. It is a promising feed additive in swine in China. In this study, we assessed the acute and sub-acute toxicity of NCG in Sprague-Dawley (SD) rats. All rats survived until they were killed at a scheduled time point. No adverse effects or mortality was observed following acute oral administration of 5000 mg/kg NCG to SD rats. No biologically significant or test substance-related differences were observed in body weights, feed consumption, clinical signs, a functional observational battery, organ weights, histopathology, ophthalmology, hematology, coagulation, and clinical chemistry parameters in any of the treatment groups in sub-acute doses of NCG at target concentrations corresponding to 500, 2000, and 3000 mg/kg/day for 28 days neither. In addition, no evidence of mutagenicity or genotoxicity was found, either in vitro in bacterial reverse mutation assay or in vivo in mice bone marrow micronucleus assay and sperm shape abnormality assay. On the basis of our findings, we conclude that NCG is a non-toxic substance with no genotoxicity.