Jejunal Tissue Research Articles

Influence of postruminal casein infusion and exogenous glucagon-like peptide 2 administration on the jejunal mucosal transcriptome in cattle.

We previously demonstrated that postruminal casein infusion and exogenous glucagon-like peptide 2 (GLP-2) administration independently stimulated growth and carbohydrase activity of the pancreas and jejunal mucosa in cattle. The objective of the current study was to profile the jejunal mucosal transcriptome of cattle using next-generation RNA sequencing in response to postruminal casein infusion and exogenous GLP-2. Twenty-four Holstein steers [250 ± 23.1 kg body weight (BW)] received a continuous abomasal infusion of 3.94 g raw corn starch/kg of BW combined with either 0 or 1.30 g casein/kg of BW for 7 d. Steers received subcutaneous injections at 0800 and 2000 h to provide either 0 or 100 μg GLP-2/kg of BW per day. At the end of the 7-d treatment period, steers were slaughtered for collection of the jejunal mucosa. Total RNA was extracted from jejunal mucosal tissue, strand-specific cDNA libraries were prepared, and RNA sequencing was conducted to generate 150-bp paired-end reads at a depth of 40 M reads per sample. Differentially expressed genes (DEG), KEGG pathway enrichment, and gene ontology enrichment were determined based on the FDR-corrected P-value (padj). Exogenous GLP-2 administration upregulated (padj < 0.05) 667 genes and downregulated 1,101 genes of the jejunal mucosa. Sphingolipid metabolism, bile secretion, adherens junction, and galactose metabolism were among the top KEGG pathways enriched with upregulated DEG (padj < 0.05) in response to exogenous GLP-2 administration. The top gene ontologies enriched with upregulated DEG (padj < 0.05) in response to exogenous GLP-2 administration included nutrient metabolic processes, brush border and bicellular tight junction assembly, and enzyme and transporter activities. Exogenous GLP-2 administration increased or tended to increase (padj < 0.10) brush border carbohydrase (MGAM, LCT, TREH), hexose transporter (SLC5A1, SLC2A2), and associated transcription factor (HNF1, GATA4, KAT2B) mRNA expression of the jejunal mucosa. Gene ontologies and KEGG pathways that were downregulated (padj < 0.05) in response to exogenous GLP-2 were related to genetic information processing. Postruminal casein infusion downregulated (padj < 0.05) 7 jejunal mucosal genes that collectively did not result in enriched KEGG pathways or gene ontologies. This study highlights some of the transcriptional mechanisms associated with increased growth, starch assimilation capacity, and barrier function of the jejunal mucosa in response to exogenous GLP-2 administration.

Developing an adult stem cell derived microphysiological intestinal system for predicting oral prodrug bioconversion and permeability in humans.

Microphysiological systems (MPS) incorporating human intestinal organoids have shown the potential to faithfully model intestinal biology with the promise to accelerate development of oral prodrugs. We hypothesized that an MPS model incorporating flow, shear stress, and vasculature could provide more reliable measures of prodrug bioconversion and permeability. Following construction of jejunal and duodenal organoid MPS derived from 3 donors, we determined the area under the concentration-time (AUC) curve for the active drug in the vascular channel and characterized the enzymology of prodrug bioconversion. Fosamprenavir underwent phosphatase mediated hydrolysis to amprenavir while dabigatran etexilate (DABE) exhibited proper CES2- and, as anticipated, not CES1-mediated de-esterification, followed by permeation of amprenavir to the vascular channel. When experiments were conducted in the presence of bio-converting enzyme inhibitors (orthovanadate for alkaline phosphatase; bis(p-nitrophenyl)phosphate for carboxylesterase), the AUC of the active drug decreased accordingly in the vascular channel. In addition to functional analysis, the MPS was characterized through imaging and proteomic analysis. Imaging revealed proper expression and localization of epithelial, endothelial, tight junction and catalytic enzyme markers. Global proteomic analysis was used to analyze the MPS model and 3 comparator sources: an organoid-based transwell model (which was also evaluated for function), Matrigel embedded organoids and finally jejunal and duodenal cadaver tissues collected from 3 donors. Hierarchical clustering analysis (HCA) and principal component analysis (PCA) of global proteomic data demonstrated that all organoid-based models exhibited strong similarity and were distinct from tissues. Intestinal organoids in the MPS model exhibited strong similarity to human tissue for key epithelial markers via HCA. Quantitative proteomic analysis showed higher expression of key prodrug converting and drug metabolizing enzymes in MPS-derived organoids compared to tissues, organoids in Matrigel, and organoids on transwells. When comparing organoids from MPS and transwells, expression of intestinal alkaline phosphatase (ALPI), carboxylesterase (CES)2, cytochrome P450 3A4 (CYP3A4) and sucrase isomaltase (SI) was 2.97-, 1.2-, 11.3-, and 27.7-fold higher for duodenum and 7.7-, 4.6-, 18.1-, and 112.2-fold higher for jejunum organoids in MPS, respectively. The MPS approach can provide a more physiological system than enzymes, organoids, and organoids on transwells for pharmacokinetic analysis of prodrugs that account for 10% of all commercial medicines.

Krameria lappacea root extract's anticoccidial properties and coordinated control of CD4 T cells for IL-10 production and antioxidant monitoring.

Recently, the use of botanicals as an alternative to coccidiostats has been an appealing approach for controlling coccidiosis. Therefore, this study was conducted to evaluate the potential role of aqueous methanolic extract (200 mg/kg) of Krameria lappacea (roots) (KLRE) against infection induced by Eimeria papillata. A total of 25 male C57BL/6 mice were divided into five groups (I, II, III, IV, and V). On 1st day of the experiment, all groups except groups I (control) and II (non-infected-treated group with KLRE), were inoculated orally with 103 sporulated E. papillata oocysts. On the day of infection, group IV was treated with KLRE. Group V served as an infected-treated group and was treated with amprolium (coccidiostat). Treatment with extract and coccidiostat was continued for five consecutive days. While not reaching the efficacy level of the reference drug (amprolium), KLRE exhibited notable anticoccidial activity as assessed by key criteria, including oocyst suppression rate, total parasitic stages, and maintenance of nutrient homeostasis. The presence of phenolic and flavonoid compounds in KLRE is thought to be responsible for its positive effects. The Eimeria infection increased the oxidative damage in the jejunum. KLRE treatment significantly increased the activity of catalase and superoxide dismutase. On the contrary, KLRE decreased the level of malondialdehyde and nitric oxide. Moreover, KLRE treatment decreased macrophage infiltration in the mice jejunal tissue, as well as the extent of CD4 T cells and NFkB. E. papillata caused a state of systemic inflammatory response as revealed by the upregulation of inducible nitric oxide synthase (iNOs)-mRNA. Upon treatment with KLRE, the activity of iNOs was reduced from 3.63 to 1.46 fold. Moreover, KLRE was able to downregulate the expression of pro-inflammatory cytokines interferon-γ, nuclear factor kappa B, and interleukin-10 -mRNA by 1.63, 1.64, and 1.38 fold, respectively. Moreover, KLRE showed a significant reduction in the expression of IL-10 protein level from 104.27 ± 8.41 pg/ml to 62.18 ± 3.63 pg/ml. Collectively, K. lappacea is a promising herbal medicine that could ameliorate the oxidative stress and inflammation of jejunum, induced by E. papillata infection in mice.

Chronic Ouabain Targets Pore-Forming Claudin-2 and Ameliorates Radiation-Induced Damage to the Rat Intestinal Tissue Barrier.

Ionizing radiation (IR) causes disturbances in the functions of the gastrointestinal tract. Given the therapeutic potential of ouabain, a specific ligand of the Na,K-ATPase, we tested its ability to protect against IR-induced disturbances in the barrier and transport properties of the jejunum and colon of rats. Male Wistar rats were subjected to 6-day intraperitoneal injections of vehicle or ouabain (1 µg/kg/day). On the fourth day of injections, rats were exposed to total-body X-ray irradiation (10 Gy) or a sham irradiation. Isolated tissues were examined 72 h post-irradiation. Electrophysiological characteristics and paracellular permeability for sodium fluorescein were measured in an Ussing chamber. Histological analysis and Western blotting were also performed. In the jejunum tissue, ouabain exposure did not prevent disturbances in transepithelial resistance, paracellular permeability, histological characteristics, as well as changes in the expression of claudin-1, -3, -4, tricellulin, and caspase-3 induced by IR. However, ouabain prevented overexpression of occludin and the pore-forming claudin-2. In the colon tissue, ouabain prevented electrophysiological disturbances and claudin-2 overexpression. These observations may reveal a mechanism by which circulating ouabain maintains tight junction integrity under IR-induced intestinal dysfunction.

Ketogenic Diet High in Saturated Fat Promotes Colonic Claudin Expression without Changes in Intestinal Permeability to Iohexol in Healthy Mice.

Ketogenic diets (KDs) have been studied in preclinical models of intestinal diseases. However, little is known of how the fat source of these diets influences the intestinal barrier. Herein, we studied the impact of four-week feeding with KD high either in saturated fatty acids (SFA-KD) or polyunsaturated linoleic acid (LA-KD) on paracellular permeability of the intestine to iohexol in healthy male C57BL/6J mice. We investigated jejunal and colonic tight junction protein expression, histological changes, and inflammatory markers (Il1b, Il6, Tnf, and Lcn2), as well as the activity and expression of intestinal alkaline phosphatase (IAP) in feces and jejunal tissue, respectively, and plasma lipopolysaccharide. KDs did not change intestinal permeability to iohexol after two or twenty-six days of feeding regardless of fat quality. SFA-KD, but not LA-KD, upregulated the colonic expression of tight junction proteins claudin-1 and -4, as well as the activity of IAP. Both KDs resulted in increased epithelial vacuolation in jejunum, and this was pronounced in SFA-KD. Jejunal Il1β expression was lower and colonic Il6 expression higher in LA-KD compared to SFA-KD. In colon, Tnf mRNA was increased in LA-KD when compared to controls. Overall, the results suggest that KDs do not influence intestinal permeability to iohexol but elicit changes in colonic tight junction proteins and inflammatory markers in both jejunum and colon. Future research will show whether these changes become of importance upon proinflammatory insults.

Effects of spore-forming Bacillus probiotics on growth performance, intestinal morphology, and immune system of broilers housed on deep litter

Aim of the study: to evaluate the effects of feed supplementation of Bacillus subtilis KATMIRA1933, B. subtilis KB41, and Bacillus amyloliquefaciens KB54 probiotics on growth performance, biochemical blood parameters, intestinal morphology, and the immune system of Ross 308 broilers housed on deep litter. Methods and Results: a total of 160 newly hatched Ross 308 broilers were involved in the 42-day-long study with evaluation of growth performance at 7, 14, 21, 28, 35, and 42 days and biochemical blood analysis, histological investigation of jejunum tissues and analysis of IL-6 and IL-10 gene expression of broilers at 42 days. As a result, probiotic KB41 significantly improved growth performance on the 42nd day of the experiment (p<0.01), increased the number of Lactobacillus bacteria in the ceca (p=0.03), and promoted IL-6 and IL-10 genes expression in the spleen of the chickens (p<0.01). The results of this study generally correspond to our previous study, which included an evaluation of Bacillus-based KB41 and KB54 probiotics on production performance and the immune system of broilers housed in cages, although the effects are less prominent. Conclusions: less prominent effects are probably linked to the colonization of the broilers' gastrointestinal tract with unknown bacilli of apparent environmental origin, as the broilers were housed in standard conditions. Significance and Impact of Study: this study provides more evidence of B. subtilis KB41 and B. amyloliquefaciens KB54 effectiveness in poultry.

The probiotic Lacticaseibacillus rhamnosus HN001 influences the architecture and gene expression of small intestine tissue in a piglet model.

This study investigated the effects of Lacticaseibacillus rhamnosus HN001 supplementation on the architecture and gene expression in small intestinal tissues of piglets used as an animal model for infant humans. Twenty-four 10-d-old entire male piglets (4·3 (sd 0·59) kg body weight) were fed an infant formula (IF) (control) or IF supplemented with 1·3 × 105 (low dose) or 7·9 × 106 (high dose) colony-forming units HN001 per ml of reconstituted formula (n 8 piglets/treatment). After 24 d, piglets were euthanised. Samples were collected to analyse the histology and gene expression (RNAseq and qPCR) in the jejunal and ileal tissues, blood cytokine concentrations, and blood and faecal calprotectin concentrations. HN001 consumption altered (false discovery rate < 0·05) gene expression (RNAseq) in jejunal tissues but not in ileal tissues. The number of ileal goblet cells and crypt surface area increased quadratically (P < 0·05) as dietary HN001 levels increased, but no increase was observed in the jejunal tissues. Similarly, blood plasma concentrations of IL-10 and calprotectin increased linearly (P < 0·05) as dietary HN001 levels increased. In conclusion, supplementation of IF with HN001 affected the architecture and gene expression of small intestine tissue, blood cytokine concentration and frequencies, and blood calprotectin concentrations, indicating that HN001 modulated small intestinal tissue maturation and immunity in the piglet model.

Oral Vitamin C Intake Ameliorates Crude Oil-Polluted Water-Induced Jejunal Contractile Dysfunctions in Wistar rats

Inhabitants of rural Niger-Delta oil communities in Nigeria often, unintentionally, consume crude oil polluted water (COCW) due to crude oil spills in the region. The impact of vitamin C supplementation during COCW ingestion on the contractile mechanism of the jejunum is not fully known. Hence, this study investigates the outcomes of COCW water and vitamin C intake on the smooth muscle activity of the jejunum in Wistar rats using standard techniques. Data obtained showed that jejunal tissue SOD concentration was significantly reduced, while jejunal tissue MDA concentration was significantly increased in the COCW-only treated group. Contraction mediated by acetylcholine was significantly increased in the COCW-only treated rats. Calcium and potassium ion influx significantly increased jejunal contraction in the COCW-only rats. Incubation of the jejunal tissue in nifedipine, indomethacin, and L-NAME significantly increased acetylcholine-mediated contractions in the jejunal tissue of the COCW-only treated rats when compared to other groups. COCW causes free radical-induced jejunal damage that results in impaired jejunal contractile activity mediated by the M2 muscarinic receptor, nitric oxide synthase activity, voltage-gated large-conductance calcium channels, potassium channels, and prostaglandins. However, the oral intake of Vitamin C supplementation significantly ameliorated impairments by enhancing jejunal antioxidant activity.&#x0D;

Persea americana extract protects intestinal tissue from Eimeria papillata-induced murine Infection

Coccidiosis is the most prevalent disease-causing widespread economic loss among farm and domestic animals. Currently, several drugs are available for the control of this disease but resistance has been confirmed for all of them. There is an urgent need, therefore, for the identification of new sources as alternative treatments to control coccidiosis. The present work aimed to study the effect of the Persea americana extract (PAE) as an anti-coccidial, anti-oxidant, and anti-apoptotic modulator during murine intestinal Eimeria papillata infection. A total of 25 male mice were divided into five groups, as follows: Group1: Non-infected-non-treated (negative control), Group2: Non-infected-treated group with PAE (500 mg/kg b.w). Group3: Infected-non-treated (positive control), Group4: Infected-treated group with PAE (500 mg/kg b.w.), and Group5: Infected-treated group with Amprolium (120 mg/kg b.w.). Groups (3–5) were orally inoculated with 1 × 103 sporulated E. papillata oocysts. After 60 min of infection, groups (4 and 5) were treated for 5 consecutive days with the recommended doses of PAE and amprolium. The fact that PAE has an anti-coccidial efficacy against intestinal E. papillata infection in mice has been clarified by the reduction of fecal oocyst output on the 5th day post-infection by about 85.41%. Moreover, there is a significant reduction in the size of each parasite stage in the jejunal tissues of the infected-treated group with PAE. PAE counteracted the E. papillata-induced loss of glutathione peroxidase (GPx), superoxide dismutase (SOD), and total antioxidant capacity (TCA). E. papillata infection also induced an increase in the apoptotic cells expressed by caspase-3 which modulated after PAE treatment. Moreover, the mRNA expression of the goblet cell response gene, mucin (MUC2), was upregulated from 0.50 to 1.20-fold after treatment with PAE. Based on our results, PAE is a promising medicinal plant with anti-coccidial, anti-oxidant, and anti-apoptotic activities and could be used as a food additive.

Ileal mineral digestibility and expression of nutrient transporter genes of broiler chickens in response to variable dietary total Ca and phytase supplementation are influenced by time on experimental diet and age of the birds

Two experiments were conducted to determine the impact of Ca, phytase, sampling time, and age on the digestibility (AID) of Ca and P and the expression of their transporters. Cobb 500 male chicks (N = 600) were used in each experiment and allocated to cages with 10 (Exp 1, 8-11 d) or 5 (Exp 2, 21-24 d) birds/cage and 10 (Exp 1) or 20 (Exp 2) reps/treatment. Treatments were a 2 × 3 factorial arrangement, with low (LOW) or standard (STD) Ca level and 3 phytase (PHY) levels (0, 300, or 3,000 FYT/kg). Ileal digesta were collected at 8, 12, 24, 48, and 72 h, and jejunum tissues at 12, 48, and 72 h after the start of feeding experimental diets. In Exp 1, there was no effect of Ca or phytase on the AID of Ca at 8, 12, or 24 h. Phytase increased the AID of P (P < 0.05) at all time points, and the magnitude was influenced by Ca. At 12 h, the mRNA level of P (NaPi-IIb) and Ca (CaSR) transporters was greatest in the LOW diets without phytase (Ca × PHY, P ≤ 0.06). In Exp 2, the STD diet decreased the AID of Ca and P (P < 0.05) at 8, 24, 48, or 72 h. Phytase increased the AID of Ca (P < 0.05) at 8, 12, and 24 h, and decreased the AID of Ca (quadratic, P < 0.05) in the STD diet (48 h). The AID of P (P < 0.05) increased with phytase at all sampling times. At 48 h, 3,000 FYT/kg decreased (P < 0.05) mRNA expression of NaPi-IIb and Ca transporter ATP2B1 in the STD diet (Ca × PHY, P < 0.05). In conclusion, to avoid adaptation of broilers to Ca and P deficiencies, the optimal time on experimental diets is ≤ 48 h for young broilers and ≤ 24 h in older birds due to up- or down-regulation of Ca and P transporters in response to dietary Ca, P, and phytase.

Oral phages prophylaxis against mixed Escherichia coli O157:H7 and Salmonella Typhimurium infections in weaned piglets

Escherichia coli and Salmonella Typhimurium are the main pathogens of diarrhea in weaned piglets. The prevention of bacterial diarrhea in weaned piglets by phage is rarely reported. We conducted this study to evaluate the preventive effect of phages on mixed Escherichia coli and Salmonella Typhimurium infections in weaned piglets. A novel phage named NJ12 was isolated by using Salmonella Typhimurium SM022 as host bacteria and characterized by electron microscopy, genomic analysis and in vitro bacteriostatic activity. Phage NJ12 and a previously reported phage EP01 were microencapsulated with sodium alginate to make phage cocktail. Microencapsulated phage cocktail and PBS (Phosphate buffer solution) were used to piglets the phage and phage-free group through oral administration before bacterial infection 2 h, respectively. Piglets of the phage and phage-free group were consumed with feed contaminated with 6 mL (108CFU/mL) Escherichia coli O157:H7 GN07 (GXEC-N07) and 6 mL (108CFU/mL) SM022 every day for seven consecutive days. The results showed that piglets in the phage-free group had more severe diarrhea, larger decreased average weight gain and higher levels of neutrophils compared with piglets in phage group. Meanwhile, piglets in the phage-free group had higher load of SM022 and GN07 in jejunal tissue and more severe intestinal damage compared with piglets in group phage in vivo. In addition, oral administration phage can significant decreased the relative abundance of Enterobacteriaceae but hardly repaired the changes of diversity and composition of gut microbiota caused by the mixed infection of SM022 and GN07. This implies that phage used as a feed additive have a marvelous preventive effect on bacterial diarrhea during weaning of piglets.

A neutral polysaccharide from Persicaria hydropiper (L.) Spach ameliorates lipopolysaccharide-induced intestinal barrier injury via regulating the gut microbiota and modulating AKT/PI3K/mTOR and MAPK signaling pathways

Ethnopharmacological relevancePersicaria hydropiper (L.) Spach, a herb that is prevalent across Asia and Europe, finds utility as both a culinary ingredient and medicinal herb. In China, P. hydropiper decoction is commonly employed to alleviate dysentery, gastroenteritis, and diarrhea symptoms. Aim of the studyTo assess the effects of a neutral polysaccharide from P. hydropiper (PHP) on the intestinal barrier (IB) injury induced by lipopolysaccharide (LPS) in mice, and elucidate the molecular mechanisms involved. Materials and methodsPHP was extracted from dried P. hydropiper herb using hot water extraction, followed by ethanol precipitation. The extract underwent successive isolation and purification steps involving anion-exchange and gel filtration chromatography. The primary structure of PHP was determined using Fourier-transformed infrared spectroscopy, ion chromatography, gas chromatography-mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy. Male BALB/c mice were randomly assigned to control (CON), model (MOD), berberine hydrochloride (BBR), and PHP (20, 40 and 80 mg/kg) groups. Histopathological changes in jejunal tissues were assessed through hematoxylin and eosin (HE) staining. The expression levels of proteins and genes involved in AKT/PI3K/mTOR and MAPK signaling pathways were evaluated using qRT-PCR and Western blotting, respectively. The composition and abundance of the gut microbiota in mice were analyzed using high-throughput 16S rRNA gene sequencing. Additionally, the concentrations of short-chain fatty acids (SCFAs) were determined using GC-MS. ResultsThe main components of PHP included arabinose, galactose, and glucose (molar ratio = 1.00:5.52:11.39). The backbone of PHP consisted of →4)-Glcp-(1→, →4,6)-Glcp-(1→, →4)-Galp-(1→, →4,6)-Galp-(1→. The branched chains primarily consisted of 5)-Araf-(1→ residues, which were attached to the backbone through →6)-Glcp-(1→ and →6)-Galp-(1→ at the 6-position. Histological analysis demonstrated that PHP exhibited a mitigating effect on intestinal damage induced by LPS. PHP could markedly reduce the mRNA levels of PI3K, AKT, mTOR, p70 S6K, Ras, Raf1, MEK1/2, p38, ERK1/2, and JNK, while downregulating the protein levels of p-mTOR, p-PI3K, p-AKT, p-p38, p-ERK, and p-JNK. PHP also modulated the diversities and abundances of the gut microbiota, resulting in an increase in the abundances of Lactobacillaceae, Anaerovoracaceae, Lachnospiraceae, Eggerthellaceae, and Desulfovibrionaceae and a decrease in the abundances of Muribaculaceae, Prevotellaceae, and Rikenellaceae. Additionally, PHP significantly increased the content of various SCFAs. ConclusionPHP emerges as a pivotal factor in the repair of IB injury by virtue of its ability to regulate the gut microbiota, elevate SCFA levels, and inhibit the MAPK and AKT/PI3K/mTOR pathways. It is worth noting that the therapeutic effect of high-dose PHP was remarkably significant, surpassing even the positive control of berberine hydrochloride.

275 Effects of Increased Nsp with Or Without Nspase Supplementation on Growth Performance, Intestinal Health, and Intestinal Microbiome of Nursery Pigs and Implications on Subsequent Phases of Production

Abstract This study aimed to investigate the effects of increased NSP from alternative feedstuffs in feeds with or without NSPase supplementation on growth performance, nutrient digestibility, intestinal health, immune status, and modulation of the intestinal microbiome of nursery pigs and the carryover effect from dietary nursery intervention on the subsequent phases of production. One hundred and sixty newly weaned pigs at 7.0 ± 0.3 kg BW were allotted in a 2 x 2 factorial arrangement with NSP level and NSPase supplementation serving as factors and sex as block. Pigs were assigned to one of 4 dietary treatments with the control having corn DDGS as the main source of NSP and HNSP having soy hulls and wheat bran replacing corn DDGS as the source of NSP. Pigs were fed for 37 d in 2 phases (19 d and 18 d, respectively) and housed in a pen of 4. Growth performance was measured at the end of each phase. At d 37, a pig representing the median BW within each pen from each treatment was euthanized for sampling jejunal mucosa, jejunal tissue, and ileal digesta to analyze intestinal health variables, oxidative stress products, and digesta viscosity. The remaining pigs were then fed a common diet for the subsequent grower and finisher phases to evaluate the carryover effect. At the end of the finisher phase, all pigs were euthanized to obtain carcass data. Data were analyzed using Mixed procedure of SAS 9.4. Overall, NSP level and type nor NSPase supplementation had an effect on growth performance; however, HNSP tended to have improved (P = 0.087) feed efficiency (0.52 to 0.53) in the grower period immediately following the nursery phase. Additionally, HNSP increased (P &amp;lt; 0.05) IgG (4.84 to 5.83 µg/mg of protein) in the jejunal mucosa and tended to increase (P = 0.096) the villus height in the jejunum (482.8 to 506.6 μm). The body weight of HNSP pigs was more uniform (P &amp;lt; 0.05; 8.7 to 11.6%) than the control at the end of the 120-day trial. No differences were observed in carcass traits however, the hot carcass weight of pigs in HNSP tended to be more uniform (P = 0.089; 6.4 to 10.00%) than the control. In conclusion, increased amounts of NSP from alternative feedstuffs does not further impair or damage the intestinal health and subsequent growth performance of nursery pigs and may aid the pig through generated metabolites and alterations to the intestinal microbiota as they are transitioned to the ensuing phases of production as evidenced by the increased uniformity at the end of the trial.

273 Effects of Xylanase Supplementation in Diets with Reduced Metabolizable Energy and Supplemental Fat on Intestinal Health, Mucosa Associated Microbiota and Growth Performance of Nursery Pigs

Abstract This study aimed to investigate the effects of reducing metabolizable energy (ME) below the requirement by reducing supplemental fat on intestinal health and growth performance of nursery pigs. One hundred and ninety-two newly weaned pigs at 7.3 ± 0.6 kg BW were allotted in a RCBD with sex and initial BW as blocks. Pigs were assigned to one of 8 dietary treatments with varying ME levels (3,400, 3,375, 3,350, 3,325, and 3,300 kcal/kg ME) by reducing supplemental fat and increasing xylanase (X0: 0, X40: 1,200, X80: 2,400 and X120: 3,600 XU/kg of feed) in ME deficient diets by 100 kcals/kg. Pigs were fed diets for 35 d in 2 phases (14 d and 21 d, respectively) and growth performance was measured at the end of each phase. Phase 2 diet had titanium dioxide (0.4%) as indigestible external marker to measure ileal digestibility of nutrients. At d 35, a pig representing the median BW within each pen from 3 dietary treatments (ME at 3,400 and 3,300 kcal/kg ME with or without xylanase) were euthanized for sampling jejunal mucosa, jejunal tissue, and ileal digesta to analyze intestinal health parameters, mucosa associated microbiota, and apparent ileal digestibility (AID) of nutrients. Data were analyzed using Mixed procedure of SAS 9.4. Overall, reducing ME level had no effect on growth performance of nursery pigs. However, reducing ME by 100 kcal/kg ME below the requirement increased (P &amp;lt; 0.05) digesta viscosity (1.7 to 2.2 cP) and protein carbonyl (0.18 to 0.40 nmol/mg), tended to change the relative abundance of Prevotellaceae (P = 0.062, 49 to 23%) and Lactobacillaceae (P = 0.086, 7 to 19%), reduced (P &amp;lt; 0.05) villus height (467 to 433 µm) and villus height to crypt depth ratio (1.80 to 1.64) in the jejunum, and reduced (P &amp;lt; 0.05) AID of DM (59.5 to 47.3%), gross energy (59.1 to 43.0%), crude protein (63.4 to 44.6%), and ether extract (67.7 to 55.2%). Xylanase supplementation decreased (P &amp;lt; 0.05) digesta viscosity (2.2 to 1.82 cP) and protein carbonyl (0.40 to 0.17 nmol/mg), increased the relative abundance of Prevotellaceae (20.22 to 61.56%), increased villus height (433 to 462 µm) and villus height to crypt depth ratio (1.64 to 1.99) in the jejunum, and increased (P &amp;lt; 0.05) AID of DM (47.3 to 58.7%), and gross energy (42.9 to 56.0%). In conclusion, reducing ME (100 kcal/kg below the requirement) by reducing supplemental fat (by 2% unit) impaired intestinal health, reducing ability of the pigs to digest feeds whereas xylanase supplementation appeared to mediate some of the negative effects seen by energy reduction by improving intestinal morphology, reducing oxidative damage products, and diversity of the mucosa-associated microbiota.

258 Effects of Dietary Supplementation of Myristic Acid on Intestinal Health and Growth Performance of Nursery Pigs

Abstract The objective of this study was to investigate the effects of myristic acid on intestinal health and growth performance of nursery pigs. Thirty-six newly weaned pigs (6.6 ± 0.4 kg BW) were assigned to 3 treatments (n = 12) in a randomized complete block design with initial BW and sex as blocks. Pigs were fed for 35 d in 3 phases (10, 10, and 15 d respectively). Treatments were 1) NC: basal diet; 2) PC: NC + 0.25% bacitracin methylene disalicylate (antibiotic; bacitracin: 276 g/t feed) in all phases; and 3) MA: NC + myristic acid at 0.075% in phases 1 and 2 and 0.045% in phase 3. Diets contained nutrients meeting the requirement. Growth performance and fecal score were measured for each phase. Pigs were euthanized to collect jejunal mucosa, jejunal tissues, and ileal digesta. Data were analyzed using preplanned contrasts (NC vs. PC and NC vs. MA) in the Proc MIXED procedure of SAS. Compared with the NC, the PC improved (P &amp;lt; 0.05) ADG in phase 1 (43 to 101 g/d) and ADFI in phase 2 (458 to 578 g/d), whereas the MA improved (P &amp;lt; 0.05) ADG (541 to 668 g/d) and ADFI (801 to 996 g/d) in phase 3. Overall, the PC tended to improve ADG (353 to 419 g/d; P = 0.063), and ADFI (515 to 606 g/d; P = 0.051), whereas the MA improved (P &amp;lt; 0.05) ADG (353 to 431 g/d) and ADFI (515 to 623 g/d) compared with the NC group. The PC decreased (P &amp;lt; 0.05) fecal score (3.17 to 3.03) in phase 2 compared with the NC. The PC tended to decrease IL8 (P = 0.053; 0.53 to 0.37 ng/mg of protein) and protein carbonyl (P = 0.075; 2.65 to 1.66 nmol/mg of protein) in jejunal mucosa compared with the NC. The PC increased (P &amp;lt; 0.05) the villus height to crypt depth ratio (2.61 to 3.02) compared with that of NC. The MA tended to decrease IgG (P = 0.051; 2.44 to 1.67 µg/mg of protein) and IL8 (P = 0.090; 0.53 to 0.39 ng/mg of protein) in jejunal mucosa compared with that of the NC. The MA tended to increase relative abundance of Bacteroidetes (P = 0.075; 3.8 to 13.9) at phylum level, Prevotellaceae (P = 0.073; 3.7 to 13.8) at family level, and Prevotella (P = 0.085; 4.2 to 15.4) at genus level compared with the NC. In this study, it was shown that both bacitracin and myristic supplementation could improve the intestinal health and growth performance of nursery pigs. Effects of bacitracin on intestinal health and growth performance was rather immediate whereas the effects of myristic acid were obtained after a 3-week feeding.

PSV-29 Plasma Citrulline Is Strongly Correlated with Jejunum and Ileum Citrulline Concentrations in Pigs at Weaning

Abstract Pigs at weaning are abruptly exposed to numerous stressors that negatively affect gut health. While the term gut health lacks a clear definition, it is best measured by its key functions, including nutrient digestion, absorption, and metabolism, mucosal immunity, and barrier function. The gut is the main endogenous source of citrulline, a potential biomarker of gut function. The objective of this study was to determine the relationship between plasma and intestinal concentrations of citrulline and citrulline precursors in pigs. Pigs (initial BW = 7,233 ± 683 g) were either weaned at 21 d age and euthanized at 23 d age (W; n = 9) or were not weaned, remained with the sow, and euthanized at 23 d age (NW; n = 9). At euthanasia, blood, jejunum, and ileum tissue was collected and snap-frozen in liquid nitrogen; segments of mid jejunum and distal ileum were collected for morphological analysis. Plasma and tissue glutamine (Gln), glutamate (Glu), proline (Pro), ornithine (Orn), and citrulline (Cit) concentrations were determined by reverse-phase HPLC following pre-column derivatization with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate. Data were analyzed with the generalized linear mixed model and correlation procedures of SAS (version 9.4). Pearson correlation coefficients between 0.50 and 0.69 were considered as moderate, between 0.70 and 0.89 as strong, and between 0.90 to 1 as very strong. Jejunum villus height was reduced in W compared with NW pigs (370 versus 246 ± 18 µm; P &amp;lt; 0.001), whereas ileum villus height was not different between NW and W pigs (257 versus 215 ± 27 µm; P = 0.27). Among precursors for gut Cit synthesis, plasma Pro was strongly correlated with jejunum Pro (r = 0.83; P &amp;lt; 0.001) and very strongly correlated with ileum Pro (r = 0.91; P &amp;lt; 0.001). Plasma Glu was moderately correlated with ileum Glu (r = 0.58; P = 0.014), but not with jejunum Glu (r = 0.18, P = 0.48), whereas plasma Gln was moderately correlated with ileum Gln (r = 0.57; P = 0.016) and strongly correlated with jejunum Gln (r = 0.80; P &amp;lt; 0.001). Plasma Orn was strongly correlated with both jejunum Orn (r = 0.73; P &amp;lt; 0.001) and ileum Orn (r = 0.78; P &amp;lt; 0.001). Plasma Cit was strongly correlated with jejunum Cit (r = 0.77; P &amp;lt; 0.001) and very strongly correlated with ileum Cit (r = 0.95; P &amp;lt; 0.001). Plasma Cit can be used to estimate gut Cit content and may be predictive of gut function in nursery pigs undergoing weaning stress.

262 Weaning-Induced Changes in Tissue Cysteine Utilization in Pigs

Abstract Weaning stress in pigs is associated with low and variable feed intake, poor nutrient utilization, and high feed costs. Cysteine is a sulfur amino acid with key roles in pig production, but how cysteine metabolism and requirements are affected by weaning stress should be better defined. The objective of this study was to determine weaning-induced changes in tissue cysteine metabolism. Pigs (initial BW = 7.23 ± 0.68 kg) were either weaned at 21 d age and euthanized at 23 d age (W; n = 9) or remained with the sow and euthanized at 23 d age (NW; n = 9). Weaned pigs were fasted to simulate acute nutritional stress but had free access to water. At euthanasia, blood, liver, jejunum, and ileum tissues were collected. Plasma and tissue amino acids and amino thiols were analyzed by HPLC. The abundance of proteins central to liver cysteine utilization were determined by immunoblot and enzyme activity assays. Glutathione (GSH) concentrations were less in jejunum (1,379 versus 1,720 ± 70 nmol/g; P &amp;lt; 0.01) and ileum (1,497 versus 1,740 ± 74 nmol/g; P &amp;lt; 0.05) in W than NW pigs. However, liver GSH concentration was not different between W and NW pigs (P &amp;gt; 0.10). While cysteine in jejunum and ileum were not affected by weaning (P &amp;gt; 0.10), γ-glutamylcysteine (γ-GC), the immediate precursor of GSH, declined in both jejunum (9.7 versus 14.3 ± 1.4 nmol/g; P &amp;lt; 0.01) and ileum (6.4 versus 11.2 ± 0.8 nmol/g; P &amp;lt; 0.001) in W compared with NW pigs. In liver, cysteine concentration was greater in W compared with NW pigs (417 versus 298 ± 33 nmol/g; P &amp;lt; 0.05). Although liver cysteine was greater in W pigs, liver activity of glutamate cysteine ligase (GCL; 2.49 versus 4.15 ± 0.25 nmol γ-GC/(mg protein•min); P &amp;lt; 0.001) and glutathione synthetase (GSS; 1.54 versus 1.92 nmol GSH/(mg protein•min); P &amp;lt; 0.001), enzymes needed for GSH production, were less in W than NW pigs. Immunoblot analysis of the catalytic subunit of GCL (GCLC; P &amp;lt; 0.05) and modifier subunit of GCL (GCLM; P = 0.057) were also lower in W compared with NW pigs, but GSS was not different between groups (P &amp;gt; 0.10). While plasma taurine was lower in W compared with NW pigs (64 versus 128 ± 7 µmol/L, P &amp;lt; 0.001), liver taurine was greater in W compared with NW pigs (4,726 versus 2,250 ± 471 nmol/g; P &amp;lt; 0.01). Despite an increase systemic and liver availability of cysteine, the rate-limiting amino acid for GSH synthesis, small intestine GSH concentrations decline. This suggests that systemic cysteine is not effectively utilized for gut GSH production in pigs after weaning.

256 Effects of Lactobacillus Fermentate Replacing Bacitracin on Biomarkers Associated with Bacterial Cell Wall Recognition and Intestinal Integrity in Pigs

Abstract Two experiments were conducted to evaluate the efficacy of Lactobacillus fermentate (LBF) as postbiotic on immune response and barrier function in jejunal mucosa of nursery pigs challenged with F18+Escherichia coli. Sixty-four pigs (32 barrows and 32 gilts) weaned at 21 d of age with 7.9 ± 0.5 kg in Exp. 1 (n = 32) and 6.6 ± 0.7 kg in Exp. 2 (n = 32) were allotted based on a randomized complete block design with sex and initial BW as blocks. The treatments were NC: no challenge/no supplement; PC: E. coli challenge/no supplement; AGP: E. coli challenge/bacitracin (30 g/t feed); and PBT: E. coli challenge/postbiotic (2 kg/t feed). Pigs were fed a basal diet for 28 d, formulated to meet the nutrient requirements of NRC (2012). At d 7, the challenged pigs were orally inoculated with F18+E. coli. At d 28, pigs were euthanized to collect jejunal tissue to analyze gene expressions involved in bacterial cell wall recognition, immune response, and intestinal barrier function. Data were analyzed using the Proc MIXED of SAS 9.4. The PC tended to reduce (P = 0.097) the relative expression of PGLYRP2A (1.12 to 0.76) and increased (P &amp;lt; 0.05) TLR4 (0.98 to 1.67) and NOD1 (0.99 to 1.56) compared with NC. The AGP tended to increase (P = 0.074) PGLYRP2A (0.76 to 1.15) and tended to reduce (P = 0.099) PGLYRP3 (1.21 to 0.82) compared with PC. The PBT increased (P &amp;lt; 0.05) PGLYRP4 (0.86 to 1.53) compared with PC. The AGP tended to reduce (P = 0.085) TLR4 (1.67 to 0.98) compared with PC. The AGP and the PBT reduced (P &amp;lt; 0.05) NOD1 (1.56 to 0.98 and 0.97, respectively) compared with PC. The PC tended to increase (P = 0.079) CD14 (1.13 to 2.13) compared with NC, however, PBT tended to reduce (P = 0.091) it (2.13 to 1.27) compared with PC. The AGP increased (P &amp;lt; 0.05) interferon-γ (1.13 to 1.21) and the PBT tended to increase (P = 0.096) it (1.13 to 1.63) compared with PC. The AGP tended to increase (P = 0.082) claudin-1 (0.85 to 1.81) compared with PC. In conclusion, F18+E. coli challenge upregulated the expression of pathogen recognition genes, including TLR4, CD14, and NOD1, which can increase the production of cytokines associated with pro-inflammatory response. The AGP showed a trend towards increasing the intestinal barrier function, possibly by increasing the expression of interferon-γ and reducing the expression of genes associated with pathogen recognition. The PBT enhanced the immunocompetence of nursery pigs by increasing the expression of interferon-γ and PGLYRP4, and reducing the expression of genes associated with pathogen recognition including NOD1 and CD14, which may indicate a reduction of the pathogen invasion.

Lactiplantibacillus plantarum GL001 alleviates jejunal oxidative damage induced by intestinal ischemia-reperfusion injury by influencing jejunal tissue metabolism through the improvement of jejunal microbial composition

Intestinal ischemia-reperfusion (IIR) injury is associated with inflammation and oxidative stress, yet its precise mechanisms remain not fully understood. IIR injury is closely linked to the gut microbiota and its metabolites. The anti-inflammatory and antioxidant effects of Lactiplantibacillus plantarum are specific to IIR. In our study, we conducted a 30-day pre-treatment of SD rats with both a standard strain of Lactiplantibacillus plantarum and Lactiplantibacillus plantarum GL001. After a 7-day cessation of treatment, we induced an IIR injury model to investigate the mechanisms by which Lactiplantibacillus plantarum alleviates IIR damage. The results demonstrate that Lactiplantibacillus plantarum effectively mitigates the inflammatory and oxidative stress damage induced by IIR. Lactiplantibacillus plantarum GL001 can improve the gut microbiota by reducing the abundance of harmful bacteria and increasing the abundance of beneficial bacteria. In IIR intestinal tissue, the levels of secondary bile acids are elevated. The content of the bacterial metabolite Calcimycin increases. Annotations of metabolic pathways suggest that Lactiplantibacillus plantarum GL001 can alleviate IIR damage by modulating calcium-phosphorus homeostasis through the regulation of parathyroid hormone synthesis, secretion, and action. Microbiota-metabolite correlation analysis reveals a significant negative correlation between calcimycin and Lactonacillus and a significant positive correlation between calcimycin and Shigella. There is also a significant positive correlation between calcimycin and secondary bile acids. Lactiplantibacillus plantarum GL001 can alleviate oxidative damage induced by IIR through improvements in gut microbiota and intestinal tissue metabolism.

Effects of fermented feed on growth performance, serum biochemical indexes, antioxidant capacity, and intestinal health of lion-head goslings.

The aim of this study was to evaluate the effects of fermented feed on growth performance, antioxidant indexes and intestinal health in lion-head goslings. 288 male lion-head goslings (one-day-old) were randomly divided into four groups (6 replicates per group, 12 samples per replicate): control group (basal diet) and fermented feed (FF) groups (basal diet supplemented with 2.5, 5.0 and 7.5% FF, respectively). The experimental period lasted 28 days. The results showed that 5.0 and 7.5% FF groups decreased feed conversion rate (FCR) when compared with the control group (p < 0.05). The 5.0% FF group reduced the activity of alkaline phosphatase (ALP) and lactate dehydrogenase (LDH) in serum; while the 7.5% FF group decreased the concentration of total cholesterol (TC), ALP and LDH activity (p < 0.05). Furthermore, the 7.5% FF group significantly increased total antioxidant capacity (T-AOC) in serum (p < 0.05); 2.5% and 5.0% FF groups significantly increased glutathione peroxidase (GSH-Px) in serum (p < 0.05); all FF groups increased the activity of superoxide dismutase (T-SOD) in serum (p < 0.05). For intestinal health, the villous height and villi/crypt ratio in jejunum were increased in all FF groups, but crypt depth was decreased (p < 0.05); The 5.0% FF groups enhanced T-AOC activity in jejunum (p < 0.05); The 2.5% and 5.0% FF groups enhanced GSH-Px activity (p < 0.05) in jejunum; All FF groups reduced malondialdehyde (MDA) level in jejunum (p < 0.05). LEfSe analysis showed that the cecum microbiota was significantly dominant in the 2.5% FF group compared to the control group including Firmicutes, Lactobacillales, Lactobacillus, and Prevotella; the flora that were significantly dominant in the 5.0% FF group compared to the control group included Bacteroidaceae, Bacteroides, Megamonas, and Prevotella; and the groups that were significantly dominant in the 7.5% FF group compared to the control group included Bacteroidota, Bacteroides, Bacteroidaceae, and Ruminococcaceae. In summary, dietary FF supplementation improved growth performance, serum biochemical parameters and antioxidant capacity of lion-head goslings, as well as improved jejunal tissue morphology and optimized intestinal flora structure. In particular, the FF addition at a dose of 7.5% was relatively more effective for lion- head goslings.