Zinc alleviates thermal stress-induced damage to the integrity and barrier function of cultured chicken embryonic primary jejunal epithelial cells via the MAPK and PI3K/AKT/mTOR signaling pathways.

Zinc (Zn) could alleviate the adverse effect of high temperature (HT) on intestinal integrity and barrier function of broilers, but the underlying mechanisms remain unclear. We aimed to investigate the possible protective mechanisms of Zn on primary cultured broiler jejunal epithelial cells exposed to thermal stress (TS). In Exp.1, jejunal epithelial cells were exposed to 40℃ (normal temperature, NT) and 44℃ (HT) for 1, 2, 4, 6, or 8 h. Cells incubated for 8 h had the lowest transepithelial resistance (TEER) and the highest phenol red permeability under HT. In Exp.2, the cells were preincubated with different Zn sources (Zn sulfate as iZn and Zn proteinate with the moderate chelation strength as oZn) and Zn supplemental levels (50 and 100 µmol/L) under NT for 24 h, and then continuously incubated under HT for another 8 h. TS increased phenol red permeability, lactate dehydrogenase (LDH) activity and p-PKC/PKC level, and decreased TEER, cell proliferation, mRNA levels of claudin-1, occludin, zona occludens-1 (ZO-1), PI3K, AKT and mTOR, protein levels of claudin-1, ZO-1 and junctional adhesion molecule-A (JAM-A), and the levels of p-ERK/ERK, p-PI3K/PI3K and p-AKT/AKT. Under HT, oZn was more effective than iZn in increasing TEER, occludin, ZO-1, PI3K, and AKT mRNA levels, ZO-1 protein level, and p-AKT/AKT level; supplementation with 50 µmol Zn/L was more effective than 100 µmol Zn/L in increasing cell proliferation, JAM-A, PI3K, AKT, and PKC mRNA levels, JAM-A protein level, and the levels of p-ERK/ERK and p-PI3K/PI3K; furthermore, supplementation with 50 µmol Zn/L as oZn had the lowest LDH activity, and the highest ERK, JNK-1, and mTOR mRNA levels. Therefore, supplemental Zn, especially 50 µmol Zn/L as oZn, could alleviate the TS-induced integrity and barrier function damage of broiler jejunal epithelial cells possibly by promoting cell proliferation and tight junction protein expression via the MAPK and PI3K/AKT/mTOR signaling pathways.

Intestinal Carnitine Status and Fatty Acid Oxidation in Response to Clofibrate and Medium-Chain Triglyceride Supplementation in Newborn Pigs.

To investigate the role of peroxisome proliferator-activated receptor alpha (PPARα) in carnitine status and intestinal fatty acid oxidation in neonates, a total of 72 suckled newborn piglets were assigned into 8 dietary treatments following a 2 (±0.35% clofibrate) × 4 (diets with: succinate+glycerol (Succ), tri-valerate (TC5), tri-hexanoate (TC6), or tri-2-methylpentanoate (TMPA)) factorial design. All pigs received experimental milk diets with isocaloric energy for 5 days. Carnitine statuses were evaluated, and fatty acid oxidation was measured in vitro using [1-14C]-palmitic acid (1 mM) as a substrate in absence or presence of L659699 (1.6 µM), iodoacetamide (50 µM), and carnitine (1 mM). Clofibrate increased concentrations of free (41%) and/or acyl-carnitine (44% and 15%) in liver and plasma but had no effects in the intestine. The effects on carnitine status were associated with the expression of genes involved in carnitine biosynthesis, absorption, and transportation. TC5 and TMPA stimulated the increased fatty acid oxidation rate induced by clofibrate, while TC6 had no effect on the increased fatty acid oxidation induced by clofibrate (p > 0.05). These results suggest that dietary clofibrate improved carnitine status and increased fatty acid oxidation. Propionyl-CoA, generated from TC5 and TMPA, could stimulate the increased fatty acid oxidation rate induced by clofibrate as anaplerotic carbon sources.

Effects of medium chain triglycerides on hepatic fatty acid oxidation in clofibrate-fed newborn piglets.

To investigate whether increasing tricarboxylic acid (TCA) cycle activity and ketogenic capacity would augment fatty acid (FA) oxidation induced by the peroxisome proliferator-activated receptor-alpha (PPARα) agonist clofibrate, suckling newborn piglets (n = 54) were assigned to 8 groups following a 2 ( ± clofibrate) × 4 (glycerol succinate [SUC], triglycerides of 2-methylpentanoic acid [T2M], valeric acid [TC5] and hexanoic acid [TC6]) factorial design. Each group was fed an isocaloric milk formula containing either 0% or 0.35% clofibrate (wt/wt, dry matter basis) with 5% SUC, T2M, TC5 or TC6 for 5 d. Another 6 pigs served as newborn controls. Fatty acid oxidation was examined in fresh homogenates of liver collected on d 6 using [1-14C] palmitic acid (1 mM) as a substrate (0.265 µCi/µmol). Measurements were performed in the absence or presence of L-carnitine (1 mM) or inhibitors of 3-hydroxy-3-methylglutaryl-CoA synthase (L659699, 1.6 µM) or acetoacetate-CoA deacylase (iodoacetamide, 50 µM). Without clofibrate stimulation, 14C accumulation in CO2 was higher from piglets fed diets containing T2M and TC5 than SUC, but similar to those fed TC6. Under clofibrate stimulation, accumulation also was higher in homogenates from piglets fed TC5 than all other dietary treatments. Interactions between clofibrate and carnitine or the inhibitors were observed (P = 0.0004) for acid soluble products (ASP). In vitro addition of carnitine increased 14C-ASP (P < 0.0001) above all other treatments, regardless of clofibrate treatment. The percentage of 14C in CO2 was higher (P = 0.0023) in TC5 than in the control group. From these results we suggest that dietary supplementation of anaplerotic and ketogenic FA could impact FA oxidation and modify the metabolism of acetyl-CoA (product of ß-oxidation) via alteration of TCA cycle activity, but the modification has no significant impact on the hepatic FA oxidative capacity induced by PPARα. In addition, the availability of carnitine is a critical element to maintain FA oxidation during the neonatal period.

The health benefits of selenium in food animals: a review.

Selenium is an essential trace mineral important for the maintenance of homeostasis in animals and humans. It evinces a strong antioxidant, anti-inflammatory and potential antimicrobial capacity. Selenium biological function is primarily achieved by its presence in selenoproteins as a form of selenocysteine. Selenium deficiency may result in an array of health disorders, affecting many organs and systems; to prevent this, dietary supplementation, mainly in the forms of organic (i.e., selenomethionine and selenocysteine) inorganic (i.e., selenate and selenite) sources is used. In pigs as well as other food animals, dietary selenium supplementation has been used for improving growth performance, immune function, and meat quality. A substantial body of knowledge demonstrates that dietary selenium supplementation is positively associated with overall animal health especially due to its immunomodulatory activity and protection from oxidative damage. Selenium also possesses potential antiviral activity and this is achieved by protecting immune cells against oxidative damage and decreasing viral replication. In this review we endeavor to combine established and novel knowledge on the beneficial effects of dietary selenium supplementation, its antioxidant and immunomodulatory actions, and the putative antimicrobial effect thereof. Furthermore, our review demonstrates the gaps in knowledge pertaining to the use of selenium as an antiviral, underscoring the need for further in vivo and in vitro studies, particularly in pigs.

Modulation of intestinal stem cell homeostasis by nutrients: a novel therapeutic option for intestinal diseases.

Intestinal stem cells, which are capable of both self-renewal and differentiation to mature cell types, are responsible for maintaining intestinal epithelial homeostasis. Recent evidence indicates that these processes are mediated, in part, through nutritional status in response to diet. Diverse dietary patterns including caloric restriction, fasting, high-fat diets, ketogenic diets and high-carbohydrate diets as well as other nutrients control intestinal stem cell self-renewal and differentiation through nutrient-sensing pathways such as mammalian target of rapamycin and AMP-activated kinase. Herein, we summarise the current understanding of how intestinal stem cells contribute to intestinal epithelial homeostasis and diseases. We also discuss the effects of diet and nutrient-sensing pathways on intestinal stem cell self-renewal and differentiation, as well as their potential application in the prevention and treatment of intestinal diseases.

Supplementation of Maternal Diets with Docosahexaenoic Acid and Methylating Vitamins Impacts Growth and Development of Fetuses from Malnourished Gilts.

BACKGROUND: Like many species, pregnant swine mobilize and repartition body nutrient stores during extreme malnutrition to support fetal development. OBJECTIVE: The objective of this study was to model chronic human maternal malnutrition and measure effects of methylating-vitamins (MVs, containing choline, folate, B-6, B-12, and riboflavin) and docosahexaenoic acid (DHA) supplementation on fetal growth and development. METHODS: Pregnant gilts (n = 24) were either fully nourished (2.0 kg/d) with a corn-plus-isolated-soy-protein basal diet (control) supplemented with MVs and DHA or nourishment was restricted throughout gestation. Basal diet fed to malnourished gilts was reduced progressively from 50% to 70% restriction (1.0 to 0.6 kg/d) and was supplemented following a 2 (±MVs) x 2 (±DHA) factorial design. Full-term c-sections were performed to assess impacts on low and normal birth weight (LBW/NBW) fetuses (n = 238). RESULTS: Body weight gain of malnourished gilts was 10% of full-fed control dams (P < 0.05), but offspring birth weight, length, girth, and percentage of LBW fetuses were not different between treatments. The number of pigs per litter was reduced by 30% in malnourished control dams. Fetal brain weights were reduced by 7% compared to positive controls (P < 0.05). Micronutrient supplementation to malnourished dams increased fetal brain weights back to full-fed control levels. Dams with DHA produced offspring with higher DHA concentrations in brain and liver (P < 0.05). Plasma choline concentration was 4-fold higher in fetuses from unsupplemented malnourished dams (P < 0.0001). Global DNA methylation status of fetuses from restricted dams was higher than in control fetuses, including brain, liver, heart, muscle, and placenta tissues (P < 0.05). Addition of DHA increased methylation in LBW fetal brains (P < 0.05). CONCLUSIONS: Despite the mobilization of maternal stores, malnourished litters displayed reduced brain development that was fully mitigated by micronutrient supplementation. Severe maternal malnutrition increased global DNA methylation in several fetal tissues that was unaltered by choline and B-vitamin supplementation.

Maternal dietary manganese protects chick embryos against maternal heat stress via epigenetic-activated antioxidant and anti-apoptotic abilities.

Maternal heat stress induced the aberrant epigenetic patterns resulting in the abnormal development of offspring embryos. It is unclear whether maternal dietary manganese supplementation as an epigenetic modifier could protect the chick embryonic development against maternal heat stress via epigenetic mechanisms. To test this hypothesis using an avian model, a completely randomized design with a 2 (maternal normal and high environmental temperatures of 21 and 32°C, respectively) × 3 (maternal dietary manganese sources, the control diet without manganese supplementation and the control diet + 120 mg/kg as either inorganic or organic manganese) factorial arrangement was adopted. Maternal environmental hyperthermia increased mRNA expressions of heat shock proteins 90 and 70, cyclin-dependent kinase 6 and B-cell CLL/lymphoma 2-associated X protein displaying oxidative damage and apoptosis in the embryonic heart. Maternal environmental hyperthermia impaired the embryonic development associated with the alteration of epigenetic status, as evidenced by global DNA hypomethylation and histone 3 lysine 9 hypoacetylation in the embryonic heart. Maternal dietary manganese supplementation increased the heart anti-apoptotic gene B-cell CLL/lymphoma 2 expressions under maternal environmental hyperthermia and manganese superoxide dismutase enzyme activity in the embryonic heart. Maternal dietary organic Mn supplementation effectively eliminated the impairment of maternal environmental hyperthermia on the embryonic development. Maternal dietary manganese supplementation up-regulated manganese superoxide dismutase mRNA expression by reducing DNA methylation and increasing histone 3 lysine 9 acetylation of its promoter. It is suggested that maternal dietary manganese addition could protect the chick embryonic development against maternal heat stress via enhancing epigenetic-activated antioxidant and anti-apoptotic abilities.

Maternal dietary zinc supplementation enhances the epigenetic-activated antioxidant ability of chick embryos from maternal normal and high temperatures.

The role of maternal dietary zinc supplementation in protecting the embryos from maternal hyperthermia-induced negative effects via epigenetic mechanisms was examined using an avian model (Gallus gallus). Broiler breeder hens were exposed to two maternal temperatures (21°C and 32°C) × three maternal dietary zinc treatments (zinc-unsupplemented control diet, the control diet + 110 mg zinc/kg inorganic or organic zinc) for 8 weeks. Maternal hyperthermia increased the embryonic mortality and induced oxidative damage evidenced by the elevated mRNA expressions of heat shock protein genes. Maternal dietary zinc deficiency damaged the embryonic development associated with the global DNA hypomethylation and histone 3 lysine 9 hyperacetylation in the embryonic liver. Supplementation of zinc in maternal diets effectively eliminated the embryonic mortality induced by maternal hyperthermia and enhanced antioxidant ability with the increased mRNA and protein expressions of metallothionein IV in the embryonic liver. The increased metallothionein IV mRNA expression was due to the reduced DNA methylation and increased histone 3 lysine 9 acetylation of the metallothionein IV promoter regardless of zinc source. These data demonstrate that maternal dietary zinc addition as an epigenetic modifier could protect the offspring embryonic development against maternal heat stress via enhancing the epigenetic-activated antioxidant ability.

Effect of dietary manganese on antioxidant status and expressions of heat shock proteins and factors in tissues of laying broiler breeders under normal and high environmental temperatures.

To investigate the effect of Mn on antioxidant status and on the expressions of heat shock proteins/factors in tissues of laying broiler breeders subjected to heat challenge, we used a completely randomised design (n 6) with a factorial arrangement of 2 environmental temperatures (normal, 21±1°C, and high, 32±1°C)×3 dietary Mn treatments (a Mn-unsupplemented basal diet (CON), or a basal diet supplemented with 120 mg Mn/kg diet, either as inorganic Mn sulphate (iMn) or as organic Mn proteinate (oMn)). There were no interactions (P>0·10) between environmental temperature and dietary Mn in any of the measured indices. High temperature decreased (P<0·003) Mn content, and also tended (P=0·07) to decrease Cu Zn superoxide dismutase (CuZnSOD) activity in the liver and heart. However, an increased Mn superoxide dismutase (MnSOD) activity (P<0·05) and a slight increase in malondialdehyde level (P=0·06) were detected in breast muscle. Up-regulated (P<0·05) expressions of heat shock factor 1 (HSF1) and HSF3 mRNA and heat shock protein 70 (HSP70) mRNA and protein were found in all three tissues. Broiler breeders fed either iMn or oMn had higher tissue Mn content (P<0·0001), heart MnSOD and CuZnSOD activities (P<0·01) and breast muscle MnSOD protein levels (P<0·05), and lower (P<0·05) breast muscle HSP70 mRNA and protein levels compared with those fed CON. Broiler breeders fed oMn had higher (P<0·03) bone Mn content than those fed iMn. These results indicate that high temperature decreases Mn retention and increases HSP70, HSF1 and HSF3 expressions in the tissues of laying broiler breeders. Furthermore, dietary supplementation with Mn in either source may enhance the heart's antioxidant ability and inhibit the expression of HSP70 in breast muscle. Finally, the organic Mn appears to be more available than inorganic Mn for bone in laying broiler breeders regardless of environmental temperatures.

Essential fatty acid supplementation during lactation is required to maximize the subsequent reproductive performance of the modern sow.

This study was conducted to investigate the effects of supplemental essential fatty acids (EFA) on sow reproductive efficiency and to estimate the concentrations of EFA required by the lactating sow for maximum subsequent reproduction. Data were collected on 480 sows (PIC Camborough) balanced by parity, with 241 and 239 sows representing Parity 1, and 3-5 (P3+), respectively. Sows were assigned randomly, within parity, to a 3 × 3 factorial arrangement plus a control diet without added lipids. Factors included linoleic (2.1%, 2.7%, and 3.3%) and α-linolenic acid (0.15%, 0.30%, and 0.45%), obtained by adding 4% of different mixtures of canola, corn and flaxseed oils to diets. Diets were corn-soybean meal based with 12% wheat middlings. The benefits of supplemental EFA were more evident for the subsequent reproduction of mature P3+ sows. For these sows, supplemental α-linolenic acid improved the proportion of sows that farrowed relative to sows weaned (linear P=0.080; 82.8, 80.5, and 92.8% for sows fed 0.15%, 0.30%, and 0.45% α-linolenic acid, respectively). In addition, supplemental linoleic acid, fed to Parity 1 and P3+ sows, tended to increase subsequent litter size (linear P=0.074; 13.2, 13.8 and 14.0 total pigs born for 2.1%, 2.7% and 3.3% linoleic acid, respectively). These results demonstrate that a minimum dietary intake of both α-linolenic and linoleic acid is required for the modern lactating sow to achieve a maximum reproductive outcome through multiple mechanisms that include rapid return to estrus, increased maintenance of pregnancy and improved subsequent litter size.

Dietary Isomers of Sialyllactose Increase Ganglioside Sialic Acid Concentrations in the Corpus Callosum and Cerebellum and Modulate the Colonic Microbiota of Formula-Fed Piglets.

BACKGROUND: Sialyllactose is a key human milk oligosaccharide and consists of sialic acid (SA) bound to a lactose molecule. Breastfed infants have increased accumulation of ganglioside-bound SA compared with formula-fed infants. OBJECTIVE: This study aimed to determine whether different isomers of sialyllactose enrich brain SA and modulate the microbiome of developing neonatal piglets. METHODS: Day-old pigs were randomly allocated to 6 diets (control, 2 or 4 g 3'-sialyllactose/L, 2 or 4 g 6'-sialyllactose/L, or 2 g polydextrose/L + 2 g galacto-oligosaccharides/L; n = 9) and fed 3 times/d for 21 d. Pigs were killed, and the left hemisphere of the brain was dissected into cerebrum, cerebellum, corpus callosum, and hippocampus regions. SA was determined by using a modified periodic acid-resorcinol reaction. Microbial composition of the intestinal digesta was analyzed with the use of 16S ribosomal DNA Illumina sequencing. RESULTS: Dietary sialyllactose did not affect feed intake, growth, or fecal consistency. Ganglioside-bound SA in the corpus callosum of pigs fed 2 g 3'-sialyllactose or 6'-sialyllactose/L increased by 15% in comparison with control pigs. Similarly, ganglioside-bound SA in the cerebellum of pigs fed 4 g 3'-sialyllactose/L increased by 10% in comparison with control pigs. Significant (P < 0.05, Adonis Test) microbiome differences were observed in the proximal and distal colons of piglets fed control compared with 4-g 6'-sialyllactose/L formulas. Differences were attributed to an increase in bacterial taxa belonging to species Collinsella aerofaciens (phylum Actinobacteria), genera Ruminococcus and Faecalibacterium (phylum Firmicutes), and genus Prevotella (phylum Bacteroidetes) (Wald test, P < 0.05, DeSeq2) compared with piglets fed the control diet. Taxa belonging to families Enterobacteriaceae and Enterococcaceae (phylum Proteobacteria), as well as taxa belonging to family Lachnospiraceae and order Lactobacillales (phylum Firmicutes), were 2.3- and 4-fold lower, respectively, in 6'-sialyllactose-fed piglets than in controls. CONCLUSIONS: Supplementation of formula with 3'- or 6'-sialyllactose can enrich ganglioside SA in the brain and modulate gut-associated microbiota in neonatal pigs. We propose 2 potential routes by which sialyllactose may positively affect the neonate: serving as a source of SA for neurologic development and promoting beneficial microbiota.

Effect of dietary manganese on antioxidant status and expression levels of heat-shock proteins and factors in tissues of laying broiler breeders under normal and high environmental temperatures.

To investigate the effect of Mn on antioxidant status and expression levels of heat-shock proteins/factors in tissues of laying broiler breeders subjected to heat challenge, we used a completely randomised design (n 6) with a factorial arrangement of 2 environmental temperatures (normal, 21 (sem 1)°C and high, 32 (sem 1)°C)×3 dietary Mn treatments (an Mn-unsupplemented basal diet (CON), or a basal diet supplemented with 120 mg Mn/kg diet as inorganic Mn sulphate (iMn) or organic Mn proteinate (oMn)). There were no interactions (P>0·10) between environmental temperature and dietary Mn in all of the measured indices. High temperature decreased (P<0·003) Mn content, and also tended (P=0·07) to decrease copper zinc superoxide dismutase (CuZnSOD) activity in the liver and heart. However, an increased manganese superoxide dismutase (MnSOD) activity (P<0·05) and a slight increase of malondialdehyde level (P=0·06) were detected in breast muscle. Up-regulated (P<0·05) expression levels of heat-shock factor 1 (HSF1) and HSF3 mRNA and heat-shock protein 70 (HSP70) mRNA and protein were found in all three tissues. Broiler breeders fed either iMn or oMn had higher tissue Mn content (P<0·0001), heart MnSOD and CuZnSOD activities (P<0·01) and breast muscle MnSOD protein levels (P<0·05), and lower (P<0·05) breast muscle HSP70 mRNA and protein levels than those fed CON. Broiler breeders fed oMn had higher (P<0·03) bone Mn content than those fed iMn. These results indicate that high temperature decreases Mn retention and increases HSP70 and HSF1, HSF3 expression levels in tissues of laying broiler breeders. Furthermore, dietary supplementation with Mn in either source may enhance heart antioxidant ability and inhibit the expression of HSP70 in breast muscle. Finally, the organic Mn appears to be more available than inorganic Mn for bone in laying broiler breeders regardless of environmental temperatures.

Peroxidised dietary lipids impair intestinal function and morphology of the small intestine villi of nursery pigs in a dose-dependent manner.

The objective of this study was to investigate the effect of increasing degrees of lipid peroxidation on structure and function of the small intestine of nursery pigs. A total of 216 pigs (mean body weight was 6·5 kg) were randomly allotted within weight blocks and sex and fed one of five experimental diets for 35 d (eleven pens per treatment with three to four pigs per pen). Treatments included a control diet without added lipid, and diets supplemented with 6 % soyabean oil that was exposed to heat (80°C) and constant oxygen flow (1 litre/min) for 0, 6, 9 and 12 d. Increasing lipid peroxidation linearly reduced feed intake (P<0·001) and weight gain (P=0·024). Apparent faecal digestibility of gross energy (P=0·001) and fat (P<0·001) decreased linearly as the degree of peroxidation increased. Absorption of mannitol (linear, P=0·097) and d-xylose (linear, P=0·089), measured in serum 2 h post gavage with a solution containing 0·2 g/ml of d-xylose and 0·3 g/ml of mannitol, tended to decrease progressively as the peroxidation level increased. Increasing peroxidation also resulted in increased villi height (linear, P<0·001) and crypt depth (quadratic, P=0·005) in the jejunum. Increasing peroxidation increased malondialdehyde concentrations (quadratic, P=0·035) and reduced the total antioxidant capacity (linear, P=0·044) in the jejunal mucosa. In conclusion, lipid peroxidation progressively diminished animal performance and modified the function and morphology of the small intestine of nursery pigs. Detrimental effects were related with the disruption of redox environment of the intestinal mucosa.

Clofibrate increases long-chain fatty acid oxidation by neonatal pigs.

BACKGROUND: Utilization of energy-dense lipid fuels is critical to the rapid development and growth of neonates. OBJECTIVE: To increase efficiency of milk fat utilization by newborn pigs, the effect of clofibrate on in vivo and in vitro long-chain fatty acid (LCFA) oxidation was evaluated. METHODS: Newborn male pigs were administered 5 mL of vehicle (2% Tween 80) with or without clofibrate (75 mg/kg body weight) once daily via i.g. gavage for 4 d. Total LCFA oxidative capacity was measured in respiration chambers after gastric infusion (n = 5 per treatment) with isoenergetic amounts of [1-(14)C]triglycerides (TGs), either oleic acid (18:1n-9) TG [3.02 mmol/kg body weight (BW)(0.75)] or erucic acid (22:1n-9) TG (2.46 mmol/kg BW(0.75)). Total expired (14)CO2 was collected and quantified at 20-min intervals over 24 h. Hepatic in vitro LCFA oxidation was determined simultaneously using [1-(14)C]oleic acid and erucic acid substrates. RESULTS: The in vivo 24-h accumulative [1-(14)C]TG oxidation (percentage of energy intake/kg BW(0.75)) tended to increase with clofibrate supplementation (P = 0.10), although there was no difference in the peak or mean utilization rate. The maximal extent of oleic acid TG oxidation was 1.6-fold that of erucic acid TG (P < 0.006). Hepatic in vitro LCFA oxidation increased 61% with clofibrate (P < 0.0008). The increase in mitochondria was 4-fold greater than in peroxisomes. The relative abundance of mRNA increased 2- to 3-fold for hepatic peroxisome proliferator-activated receptor α and its target genes (fatty acyl-coenzyme A oxidase and carnitine palmitoyltransferase) in the pigs that were administered clofibrate (P < 0.04). CONCLUSION: Clofibrate may improve in vivo LCFA oxidative utilization in neonatal pigs.

Dietary supplementation of aspartate enhances intestinal integrity and energy status in weanling piglets after lipopolysaccharide challenge.

The intestine has a high requirement for ATP to support its integrity, function and health, and thus, energy deficits in the intestinal mucosa may play a critical role in intestinal injury. Aspartate (Asp) is one of the major sources of ATP in mammalian enterocytes via mitochondrial oxidation. We hypothesized that dietary supplementation of Asp could attenuate lipopolysaccharide (LPS)-induced intestinal damage via modulation of intestinal energy status. Twenty-four weanling piglets were allotted to one of four treatments: (1) nonchallenged control, (2) LPS-challenged control, (3) LPS+0.5% Asp treatment, and (4) LPS+1.0% Asp treatment. On day 19, pigs were injected with saline or LPS. At 24 h postinjection, pigs were killed and intestinal samples were obtained. Asp attenuated LPS-induced intestinal damage indicated by greater villus height and villus height/crypt depth ratio as well as higher RNA/DNA and protein/DNA ratios. Asp improved intestinal function indicated by increased intestinal mucosal disaccharidase activities. Asp also improved intestinal energy status indicated by increased ATP, ADP and total adenine nucleotide contents, adenylate energy charge and decreased AMP/ATP ratio. In addition, Asp increased the activities of tricarboxylic acid cycle key enzymes including citrate synthase, isocitrate dehydrogenase and alpha-oxoglutarate dehydrogenase complex. Moreover, Asp down-regulated mRNA expression of intestinal AMP-activated protein kinase α1 (AMPKα1), AMPKα2, silent information regulator 1 (SIRT1) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC1α) and decreased intestinal AMPKα phosphorylation. These results indicate that Asp may alleviate LPS-induced intestinal damage and improve intestinal energy status.

Fish oil alleviates activation of the hypothalamic-pituitary-adrenal axis associated with inhibition of TLR4 and NOD signaling pathways in weaned piglets after a lipopolysaccharide challenge.

Long-chain n-3 (ω-3) polyunsaturated fatty acids exert beneficial effects in neuroendocrine dysfunctions in animal models and clinical trials. However, the mechanism(s) underlying the beneficial effects remains to be elucidated. We hypothesized that dietary treatment with fish oil (FO) could mitigate LPS-induced activation of the hypothalamic-pituitary-adrenal (HPA) axis through inhibition of Toll-like receptor 4 and nucleotide-binding oligomerization domain protein signaling pathways. Twenty-four weaned pigs were used in a 2 × 2 factorial design, and the main factors consisted of diet (5% corn oil vs. 5% FO) and immunological challenge (saline vs. LPS). After 21 d of dietary treatment with 5% corn oil or FO diets, pigs were treated with saline or LPS. Blood samples were collected at 0 (preinjection), 2, and 4 h postinjection, and then pigs were humanely killed by intravenous injection of 40 mg/kg body weight sodium pentobarbital for tissue sample collection. FO led to enrichment of eicosapentaenoic acid and docosahexaenoic acid and total n-3 polyunsaturated fatty acids in hypothalamus, pituitary gland, adrenal gland, spleen, and thymus. FO decreased plasma adrenocorticotrophin and cortisol concentrations as well as mRNA expressions of hypothalamic corticotropin releasing hormone and pituitary proopiomelanocortin. FO also reduced mRNA expression of tumor necrosis factor-α in hypothalamus, adrenal gland, spleen, and thymus, and of cyclooxygenase 2 in hypothalamus. Moreover, FO downregulated the mRNA expressions of Toll-like receptor 4 (TLR4) and its downstream molecules, including cluster differentiation factor 14, myeloid differentiation factor 2, myeloid differentiation factor 88, interleukin-1 receptor-associated kinase 1, tumor necrosis factor-α receptor-associated factor 6, and nuclear factor kappa-light-chain-enhancer of activated B cells p65, and also decreased the mRNA expressions of nucleotide-binding oligomerization domain 1, nucleotide-binding oligomerization domain 2, and their adaptor molecule receptor-interacting serine/threonine-protein kinase 2. These results suggested that FO attenuates the activation of the HPA axis induced by LPS challenge. The beneficial effects of FO on the HPA axis may be associated with decreasing the production of brain or peripheral proinflammatory cytokines through inhibition of TLR4 and nucleotide-binding oligomerization domain protein signaling pathways.

Effects of creep feeding and supplemental glutamine or glutamine plus glutamate (Aminogut) on pre- and post-weaning growth performance and intestinal health of piglets.

BACKGROUND: Creep feeding is used to stimulate piglet post-weaning feed consumption. L-Glutamine (GLN) is an important source of fuel for intestinal epithelial cells. The objective of this study was to determine the impact of creep feeding and adding GLN or AminoGut (AG; containing glutamine + glutamate) to pre- and post-weaning diets on pig performance and intestinal health. Litters (N = 120) were allotted to four treatments during 14-21 d of lactation: 1) No creep feed (NC, n = 45); 2) creep fed control diet (CFCD, n = 45); 3) creep fed 1% GLN (CFGLN, n = 15); 4) creep fed .88% AG (CFAG, n = 15). After weaning, the NC and CFCD groups were sub-divided into three groups (n = 15 each), receiving either a control nursery diet (NC-CD, CFCD-CD) or a diet supplemented with either GLN (NC-GLN, CFCD-GLN) or with AG (NC-AG, CFCD-AG). Litters that were creep fed with diets containing GLN or AG also were supplemented with those amino acids in the nursery diets (CFGLN-GLN, CFAG-AG). Glutamine was added at 1% in all three post-weaning diet phases and AG was added at .88% in phase 1 and 2 and at .66% in phase 3. RESULTS: Feed conversion (feed/gain) showed means among treatment means close to significance (P = 0.056) and Tukey's test for pairwise mean comparisons showed that Pigs in the CFGLN-GLN group had the best feed conversion (feed/gain) in the first three-week period post-weaning, exceeding (P = 0.044) controls (CFCD-CD) by 34%. The NC-AG group had (P = 0.02) the greatest feed intake in the last three week of the study, exceeding controls (CFCD-CD) by 12%. CFGLN-GLN, CFCD-GLN and sow reared (SR) pigs had the greatest (P = 0.049) villi height exceeding the CFCD-AG group by 18%, 20% and 19% respectively. The CFAG-AG group had the deepest (P = 0.001) crypts among all treatments. CFGLN-GLN, CFCD-GLN and SR groups had the greatest (P = 0.001) number of cells proliferating (PCNA) exceeding those in the NC-CD group by 43%, 54% and 63% respectively. Sow reared pigs showed the greatest (P = 0.001) intestinal absorption capacity for xylose and mannitol. CONCLUSION: Supplementation of creep feed and nursery diets with GLN and/or AminoGut in the first three week improved feed conversion possibly due to improved intestinal health.

Dietary supplementation of Bifidobacterium longum strain AH1206 increases its cecal abundance and elevates intestinal interleukin-10 expression in the neonatal piglet.

Intestinal microbiota of infants differ in response to gestational age, delivery mode and feeding regimen. Dietary supplementation of probiotic bacteria is one method of promoting healthy populations. We examined the impact of a novel probiotic strain of Bifidobacterium longum (AH1206) on the health, growth and development of neonatal pigs as a model for infants. Day-old pigs were fed milk-based formula containing AH1206 at 0, 109, or 10¹¹ CFU/d for 18 d (n=10/treatment). Differences were not detected in growth, organ weights or body temperatures (P>0.1); however pigs fed the high dose showed a small (2%) reduction in feed intake. Bacterial translocation was not affected as indicated by total anaerobic and aerobic counts (CFU) in samples of spleen, liver and mesenteric lymph nodes (P>0.1). Feeding AH1206 had no effects on fecal consistency, but increased the density of B. longum in the cecum. Ileal TNF expression tended to increase (P=0.08) while IL-10 expression increased linearly (P=0.01) with supplementation. Based upon findings in the suckling piglet model, we suggest that dietary supplementation with B. longum (AH1206) may be safe for human infants based on a lack of growth, development or deleterious immune-related effects observed in piglets.

Fish oil increases muscle protein mass and modulates Akt/FOXO, TLR4, and NOD signaling in weanling piglets after lipopolysaccharide challenge.

Proinflammatory cytokines play a key role in the pathophysiology of muscle atrophy. In addition, n3 polyunsaturated fatty acids (PUFAs) exert an inhibitory effect on proinflammatory cytokines affecting many inflammatory diseases. We hypothesized that dietary supplementation of fish oil could attenuate lipopolysaccharide (LPS)-induced muscle atrophy. Weanling pigs were used in a 2 × 2 factorial design and the main factors included diet (5% corn oil or 5% fish oil) and immunological challenge (LPS or saline). After 21 d of treatment with either fish oil or corn oil, pigs received an i.p. injection of either saline or LPS. At 4 h postinjection, blood and muscle samples were obtained. Fish oil led to enrichment of eicosapentaenoic acid, docosahexaenoic acid, and total n3 PUFAs in muscles. Fish oil increased muscle protein mass, indicated by a higher protein:DNA ratio in gastrocnemius and longissimus dorsi (LD) muscles. In addition, fish oil increased Akt1 mRNA abundance and decreased Forkhead Box O (FOXO) 1 and FOXO4 mRNA abundance. Fish oil also increased phosphorylation of Akt and FOXO1 in gastrocnemius and LD muscles. Fish oil decreased the mRNA abundance of muscle atrophy F-box (MAFbx) and muscle RING finger 1 in gastrocnemius and LD muscles. Moreover, fish oil reduced the plasma tumor necrosis factor (TNF) α, muscle TNFα, and prostaglandin E2 concentrations, and muscle TNFα and cyclooxygenase 2 (COX2) mRNA abundance. Finally, fish oil downregulated the mRNA abundance of muscle toll-like receptor (TLR4) and its downstream signaling molecules [myeloid differentiation factor 88 (MyD88), TNFα receptor-associated factor 6 (TRAF6), and NF-κB p65], and nucleotide-binding oligomerization domain protein (NOD1), NOD2, and their adaptor molecule [receptor-interacting serine/threonine-protein kinase 2 (RIPK2)]. These results indicate fish oil may suppress muscle proinflammatory cytokine production via regulation of TLR and NOD signaling pathways and therefore improve muscle protein mass, possibly through maintenance of Akt/FOXO signaling.

Fish oil enhances intestinal integrity and inhibits TLR4 and NOD2 signaling pathways in weaned pigs after LPS challenge.

Long-chain (n-3) PUFA exert beneficial effects on inflammatory bowel diseases in animal models and clinical trials. In addition, pattern recognition receptors such as toll-like receptors (TLR) and nucleotide-binding oligomerization domain proteins (NOD) play a critical role in intestinal inflammation. We hypothesized that fish oil could alleviate Escherichia coli LPS-induced intestinal injury via modulation of TLR4 and NOD signaling pathways. Twenty-four weaned piglets were used in a 2 × 2 factorial design and the main factors included a dietary treatment (5% corn oil or 5% fish oil) and immunological challenge (LPS or saline). After feeding fish oil or corn oil diets for 21 d, pigs were injected with LPS or saline. At 4 h postinjection, blood samples were collected and pigs were killed. EPA, DHA, and total (n-3) PUFA were enriched in intestinal mucosa through fish supplementation. Fish oil improved intestinal morphology, indicated by greater villus height and villus height:crypt depth ratio, and intestinal barrier function, indicated by decreased plasma diamine oxidase (DAO) activity and increased mucosal DAO activity as well as enhanced protein expression of intestinal tight junction proteins including occludin and claudin-1. Moreover, fish oil decreased intestinal TNFα and PGE(2) concentrations and caspase-3 and heat shock protein 70 protein expression. Finally, fish oil downregulated the mRNA expression of intestinal TLR4 and its downstream signals myeloid differentiation factor 88, IL-1 receptor-associated kinase 1, TNFα receptor-associated factor 6, and NOD2, and its adaptor molecule, receptor-interacting serine/threonine-protein kinase 2. Fish oil decreased the protein expression of intestinal NFκB p65. These results indicate that fish oil supplementation is associated with inhibition of TLR4 and NOD2 signaling pathways and concomitant improvement of intestinal integrity under an inflammatory condition.