A microencapsulated blend of botanicals supports weaning piglets during a lipopolysaccharide challenge by modulating liver inflammation and intestinal integrity.

This study examined the action of a blend of botanicals (BOT) against lipopolysaccharide (LPS)-induced inflammation on cultured hepatocytes and weaning piglets. In vitro studies examined HepG2 cells treated with BOT and challenged with Escherichiacoli LPS for 8 d. BOT treatment reduced IL-6 concentration in cell culture media across time (P < 0.05) and decreased pro-inflammatory cytokine expression on days 1 and 8 of experiment (TNFα, IL-1ß; P < 0.05). BOT also increased the expression of antioxidant enzymes (GPX-2, SOD, CAT) on day 8 (P < 0.05), which was supported by lowered reactive oxygen species concentration after LPS challenge (P < 0.1). The in vivo study was conducted with 72 weaning pigs, allotted into 24 pens and divided into 3 groups: a negative control (CTR-, basal diet), a challenged control (CTR+) that received an intraperitoneal injection of E. coli O55:B5 LPS on days 14 and 16, and a challenged treated group which received a diet containing 1.5 g/kg of microencapsulated BOT (BOT+) for the whole duration of the study. Growth performance was determined weekly and, on days 21 (1 animal per pen) and 28 (remaining animals), pigs were sacrificed to collect liver and jejunal tissues. After the challenge, BOT+ pigs had increased BW on days 21 (P < 0.05) and 28 (P < 0.1) compared to CTR+. Similar improvements in average daily gain and FCR on days 14 to 21 (P < 0.05) and 21 to 28 (P < 0.1) were also seen in BOT+ group. In the liver, compared to CTR+ pigs, BOT+ pigs had downregulated expression of TLR-4, IL-6, IFN-γ on day 21 (P < 0.05), and TLR-4, TNF-α, IL-8 on day 28 (P < 0.05). BOT+ also increased GPX-2 expression on days 21 and 28 (P < 0.05), while also upregulating SOD-1 and SOD-2 on day 21 (P < 0.05) and CAT on day 28 (P < 0.05) compared to CTR+. In the jejunum, BOT+ reduced inflammation by affecting cytokine expression (P < 0.05) and increasing the expression of tight-junction proteins, ZO-1 on day 21 and CLD-1 on day 28 (P < 0.05). Furthermore, BOT+ pigs had lower crypt depth on days 21 (P < 0.1) and 28 (P < 0.05), and increased villi-to-crypt ratio on days 21 and 28 (P < 0.05). By day 28, BOT+ intestinal measurements were restored to values similar to the CTR-. Finally, BOT+ also reduced mast cell activation on day 21 (P < 0.05) compared to CTR+. Considering all the findings, BOT controlled inflammatory activation and oxidative stress in liver cells, enhanced intestinal integrity, and as a result improved the growth performance of weaning piglets challenged with LPS.

Piglets are particularly susceptible to stress due to the abrupt changes they face during weaning. These stressors cause a surge of oxidation and inflammation, particularly in the intestinal tract. Inflammation in the intestine causes a loss in its barrier function and facilitates the translocation of harmful compounds. Of particular concern is the translocation of lipopolysaccharide (LPS), which elicits an immune response in the liver, diverting energy from growth to inflammatory processes. Exposure to LPS also has the potential to have long-lasting detrimental effects on piglets' health. Research has identified the potential of many botanicals to minimize weaning stress through diverse modes of action. This study investigated the efficacy of a blend of botanicals (BOT) to help hepatocytes control inflammatory stress in vitro and to ameliorate the effects of an LPS challenge in piglets in vivo. Our in vitro and in vivo models successfully generated an inflammatory state. In vitro, BOT decreased inflammation and oxidation, and similar effects were seen in vivo, where BOT supplementation modulated the expression of cytokines in the liver and maintained intestinal integrity. These effects validate BOT ability to improve the performance of LPS-challenged piglets and support its utilization as a feed supplement to mitigate weaning stress.

Calcium trafficking and gastrointestinal physiology following an acute lipopolysaccharide challenge in pigs.

The influence of systemic immune activation on whole-body calcium (Ca) trafficking and gastrointestinal tract (GIT) physiology is not clear. Thus, the study objectives were to characterize the effects of lipopolysaccharide (LPS) on Ca pools and GIT dynamics to increase understanding of immune-induced hypocalcemia, ileus, and stomach hemorrhaging. Twelve crossbred pigs [44 ±â€…3 kg body weight (BW)] were randomly assigned to 1 of 2 intramuscular treatments: (1) control (CON; 2 mL saline; n = 6) or (2) LPS (40 µg LPS/kg BW; n = 6). Pigs were housed in metabolism stalls to collect total urine and feces for 6 h after treatment administration, at which point they were euthanized, and various tissues, organs, fluids, and digesta were weighed, and analyzed for Ca content. Data were analyzed with the MIXED procedure in SAS 9.4. Rectal temperature and respiration rate increased in LPS relative to CON pigs (1.4 °C and 32%, respectively; P ≤ 0.05). Inflammatory biomarkers such as circulating alkaline phosphatase, aspartate aminotransferase, and total bilirubin increased in LPS compared with CON pigs whereas albumin decreased (P ≤ 0.02). Plasma glucose and urea nitrogen decreased and increased, respectively, after LPS (43% and 80%, respectively; P < 0.01). Pigs administered LPS had reduced circulating ionized calcium (iCa) compared to CON (15%; P < 0.01). Considering estimations of total blood volume, LPS caused an iCa deficit of 23 mg relative to CON (P < 0.01). Adipose tissue and urine from LPS pigs had reduced Ca compared to CON (39% and 77%, respectively; P ≤ 0.05). There did not appear to be increased Ca efflux into GIT contents and no detectable increases in other organ or tissue Ca concentrations were identified. Thus, while LPS caused hypocalcemia, we were unable to determine where circulating Ca was trafficked. LPS administration markedly altered GIT dynamics including stomach hemorrhaging, diarrhea (increased fecal output and moisture), and reduced small intestine and fecal pH (P ≤ 0.06). Taken together, changes in GIT physiology suggested dyshomeostasis and alimentary pathology. Future research is required to fully elucidate the etiology of immune activation-induced hypocalcemia and GIT pathophysiology.

Lipopolysaccharide (LPS) activates the immune system and this is accompanied with hypocalcemia and altered gastrointestinal tract (GIT) physiology. The study objectives were to characterize whole-body calcium (Ca) trafficking and evaluate GIT dynamics during LPS-induced immune activation. Ca concentrations were analyzed after intramuscular LPS injection. Administering LPS caused marked alterations in metabolic and inflammatory biomarkers and GIT dynamics, characterized by increased lower GIT motility and stomach hemorrhaging. Circulating Ca and adipose tissue and urine Ca output were decreased after LPS. Ca concentrations in other tissues and GIT contents were not detectably different. Thus, we were unable to account for about 110 mg Ca following LPS. Where and how circulating Ca is partitioned during immune activation remains unclear.

The mechanism of cadmium exposure-induced lymph node necroptosis and inflammation in piglets: Activation of CYP450 through VDR / CREB1 pathway.

Cadmium (Cd) is toxic non-essential heavy metal that precipitates adverse health effects in humans and animals, but the effect of Cd on lymph node toxicity of piglets is still unclear. In order to explore the possible molecular mechanism of Cd toxicity to lymph nodes of piglets, ten 6-week-old male weaned piglets were randomly divided into two groups, C group and Cd group. Group C was fed with basal diet, while group Cd was fed with basal diet supplemented with CdCl2 (20 mg/kg) for 40 days, the pigs were euthanized and the mesenteric, inguinal and submandibular lymph nodes (MLN, ILN, SLN) were collected. The results indicated that Cd could induce the inflammatory cell infiltration, microvascular hemorrhage, microthrombosis and cell necrosis in MLN, ILN and SLN of piglets, induced Cytochrome P450 proteins (CYP1A1、CYP2E1、CYP2A1 and CYP3A2) mRNA levels and the protein levels of Vitamin D receptor (VDR) and cAMP response element binding protein 1 (CREB1). In addition, Cd exposure upregulated the mRNA and protein levels of dynamin-related protein 1 (DRP1), receptor-interacting protein kinase 3 (RIP3), mixed lineage kinase domain-like protein (MLKL), and increased tumor necrosis factor-α (TNFα), interferon-γ (IFNγ), interleukin-2 (IL-2), interleukin-4 (IL-4), cyclooxygenase 2 (COX-2) protein levels, and the damage degree of three kinds of lymph nodes was similar after Cd exposure. In general, these results manifest that Cd exposure regulates VDR/CREB1 pathway, activates CYP450s, induces necroptosis of lymph nodes, and leads to inflammation.

Trace amounts of antibiotic exacerbated diarrhea and systemic inflammation of weaned pigs infected with a pathogenic Escherichia coli.

The experiment was conducted to investigate the effects of trace amounts of antibiotic on growth performance, diarrhea, systemic immunity, and intestinal health of weaned pigs experimentally infected with an enterotoxigenic Escherichia coli. Weaned pigs (n = 34, 6.88 ± 1.03 kg body weight [BW]) were individually housed in disease containment rooms and randomly allotted to one of the three dietary treatments: nursery basal diet (CON) and two additional diets supplemented with 0.5 or 50 mg/kg carbadox to the nursery basal diet (TRA or REC), respectively. The experiment lasted 18 d with 7 d before and 11 d after the first E. coli inoculation. The E. coli F18 inoculum was orally provided to all pigs with a dose of 1010 colony-forming unit (CFU)/3 mL for three consecutive days. Fecal and blood samples were collected on day 0 before inoculation and days 2, 5, 8, and 11 postinoculation (PI) to test the percentage of ß-hemolytic coliforms in total coliforms and complete blood cell count, respectively. Sixteen pigs were euthanized on day 5 PI, whereas the remaining pigs were euthanized at the end of the experiment to collect the jejunal and ileal mucosa and mesenteric lymph node for gene expression and bacterial translocation, respectively. Pigs in REC had greater (P < 0.05) final BW and lower (P < 0.05) overall frequency of diarrhea compared with pigs in the CON and TRA groups. Pigs in TRA had the lowest (P < 0.05) average daily gain and feed efficiency from day 0 to 5 PI, highest (P < 0.05) percentage of ß-hemolytic coliforms in fecal samples on days 2 and 5 PI, and greatest (P < 0.05) bacterial colonies in mesenteric lymph nodes on day 11 PI compared with pigs in the CON and REC groups. Pigs in TRA had the greatest (P < 0.05) neutrophils on day 5 PI and higher (P < 0.05) white blood cell counts and lymphocytes than other groups on day 11 PI. Pigs in TRA had the greatest (P < 0.05) serum C-reactive protein on days 2 and 5 PI and serum tumor necrosis factor-α on day 5 PI, compared with pigs in the CON and REC groups. Pigs fed REC had increased (P < 0.05) mRNA expression of zona occludens-1 (ZO-1) and occludin (OCDN) and reduced (P < 0.05) interleukin-1 beta (IL1B), interleukin-6 (IL6), and tumor necrosis factor-alpha (TNFA) in ileal mucosa on day 5 PI, compared with the CON, whereas TRA upregulated (P < 0.05) mRNA expression of IL1B, IL6, and cyclooxygenase-2 (COX2) in the ileal mucosa on day 11 PI, compared with the REC. In conclusion, trace amounts of antibiotic may exacerbate the detrimental effects of E. coli infection on pig performance by increasing diarrhea and systemic inflammation of weanling pigs.

Eugenol attenuates inflammatory response and enhances barrier function during lipopolysaccharide-induced inflammation in the porcine intestinal epithelial cells.

Eugenol (4-allyl-2-methoxyphenol) is an essential oil component, possessing antimicrobial, anti-inflammatory, and antioxidative properties; however, the effect of eugenol on porcine gut inflammation has not yet been investigated. In this study, an in vitro lipopolysaccharide (LPS)-induced inflammation model in porcine intestinal epithelial cells (IPEC-J2) has been set up. Cells were pretreated with 100 µM (16.42 mg/L) eugenol for 2 h followed by 10 µg/mL LPS stimulation for 6 h. Proinflammatory cytokine secretion; reactive oxygen species; gene expression of proinflammatory cytokines, tight junction proteins, and nutrient transporters; the expression and distribution of zonula occludens-1 (ZO-1); transepithelial electrical resistance (TEER); and cell permeability were measured to investigate the effect of eugenol on inflammatory responses and gut barrier function. The results showed that eugenol pretreatment significantly suppressed the LPS-stimulated interleukin-8 level and the mRNA abundance of tumor necrosis factor-α and restored the LPS-stimulated decrease of the mRNA abundance of tight junction proteins, such as ZO-1 and occludin, and the mRNA abundance of nutrient transporters, such as B0 1 system ASC sodium-dependent neutral amino acid exchanger 2, sodium-dependent glucose transporter 1, excitatory amino acid transporter 1, and peptide transporter 1. In addition, eugenol improved the expression and even redistribution of ZO-1 and tended to increase TEER value and maintained the barrier integrity. In conclusion, a low dose of eugenol attenuated inflammatory responses and enhanced selectively permeable barrier function during LPS-induced inflammation in the IPEC-J2 cell line.

Holly polyphenols alleviate intestinal inflammation and alter microbiota composition in lipopolysaccharide-challenged pigs.

The effect of holly polyphenols (HP) on intestinal inflammation and microbiota composition was evaluated in a piglet model of lipopolysaccharide (LPS)-induced intestinal injury. A total of twenty-four piglets were used in a 2 × 2 factorial design including diet type and LPS challenge. After 16 d of feeding with a basal diet supplemented with or without 250 mg/kg HP, pigs were challenged with LPS (100 µg/kg body weight) or an equal volume of saline for 4 h, followed by analysis of disaccharidase activities, gene expression levels of several representative tight junction proteins and inflammatory mediators, the SCFA concentrations and microbiota composition in intestinal contents as well as proinflammatory cytokine levels in plasma. Our results indicated that HP enhanced intestinal disaccharidase activities and reduced plasma proinflammatory cytokines including TNF-α and IL-6 in LPS-challenged piglets. Moreover, HP up-regulated mRNA expression of intestinal tight junction proteins such as claudin-1 and occludin. In addition, bacterial 16S rRNA gene sequencing showed that HP altered hindgut microbiota composition by enriching Prevotella and enhancing SCFA production following LPS challenge. These results collectively suggest that HP is capable of alleviating LPS-triggered intestinal injury by improving intestinal disaccharidase activities, barrier function and SCFA production, while reducing intestinal inflammation.

Effects of dietary supplementation of Lycium barbarum polysaccharides on growth performance, immune status, antioxidant capacity and selected microbial populations of weaned piglets.

Lycium barbarum polysaccharides (LBPs) are a complex mixture of highly branched and partially characterised polysaccharides and proteoglycans extracted from the goji berry. This mixture has great potential as a novel feed supplement for pigs. Two trials were conducted to evaluate the effects of supplementation with LBPs on the growth performance, immune status, antioxidant capacity and selected intestinal microbial populations in weaned piglets. In trial 1, a total of 400 weaned piglets [(Yorkshire × Landrace) × Duroc] with an average body weight (BW) of 6.34 ± 0.16 kg (21 days of age) were divided into five groups and fed a basal diet (control group) or a basal diet containing 1,000, 2,000, 4,000 or 6,000 mg/kg LBPs (supplemented at the expense of corn). Supplementation with 4,000 or 6,000 mg/kg LBPs for 2 weeks significantly increased the average daily gain (ADG) and average daily feed intake (ADFI) of the pigs compared with the control group (p < .05). In trial 2, thirty-two 21-days-old weaned piglets (BW: 6.33 ± 0.11 kg) were allotted to a control group (fed with a basal diet) or an experimental group (basal diet containing 4,000 mg/kg LBPs). The experiment lasted for 14 days. Pigs fed LBP diets exhibited an increased ADG and ADFI, and a decreased diarrhoeal incidence compared with those fed the basal diets (p < .05). Supplementation with LBPs increased the serum IgG and IgM levels (p < .05). Dietary LBPs effectively promoted antioxidant defence properties through enhancing the activities of serum, liver superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px), in addition to decreasing the malondialdehyde (MDA) content (p < .05). The addition of LBPs increased the amounts of Bacteroidetes in the ileum and caecum and the caecal contents of Lactobacillus spp. and Bifidobacterium spp. (p < .05), while decreased the populations of Escherichia coli and Firmicutes in the ileum and caecum (p < .05) compared with the control group. Our results suggest that dietary supplementation with LBPs can enhance growth performance, immune status and antioxidant capacity, and improve the intestinal microbial populations of weaned piglets.

Porcine Model of Infrarenal Abdominal Aortic Aneurysm.

Large animal models to study abdominal aortic aneurysms are sparse. The purpose of this model is to create reproducible, clinically significant infrarenal abdominal aortic aneurysms (AAA) in swine. To achieve this, we use a combination of balloon angioplasty, elastase and collagenase, and a lysyl oxidase inhibitor, called ß-aminopropionitrile (BAPN), to create clinically significant infrarenal aortic aneurysms, analogous to human disease. Noncastrated male swine are fed BAPN for 7 days prior to surgery to achieve a steady state in the blood. A midline laparotomy is performed and the infrarenal aorta is circumferentially dissected. An initial measurement is recorded prior to aneurysm induction with a combination of balloon angioplasty, elastase (500 units)/collagenase (8000 units) perfusion, and topical elastase application. Swine are fed BAPN daily until terminal procedure on either postoperative day 7, 14, or 28, at which time the aneurysm is measured, and tissue procured. BAPN + surgery pigs are compared to pigs that underwent surgery alone. Swine treated with BAPN and surgery had a mean aortic dilation of 89.9% ± 47.4% at day 7, 105.4% ± 58.1% at day 14, and 113.5% ± 30.2% at day 28. Pigs treated with surgery alone had significantly smaller aneurysms compared to BAPN + surgery animals at day 28 (p < 0.0003). The BAPN + surgery group had macroscopic and immunohistochemical evidence of end stage aneurysmal disease. Clinically significant infrarenal AAA can be induced using balloon angioplasty, elastase/collagenase perfusion and topical application, supplemented with oral BAPN. This model creates large, clinically significant AAA with hallmarks of human disease. This has important implications for the elucidation of AAA pathogenesis and testing of novel therapies and devices for the treatment of AAA. Limitations of the model include variation in BAPN ingested by swine, quality of elastase perfusion, and cost of BAPN.

Beta-hydroxy beta-methyl butyrate decreases muscle protein degradation via increased Akt/FoxO3a signaling and mitochondrial biogenesis in weanling piglets after lipopolysaccharide challenge.

The aim of this study was to investigate the effects of dietary ß-hydroxy-ß-methylbutyrate (HMB) on lipopolysaccharide (LPS)-induced muscle atrophy and to investigate the mechanisms involved. Sixty pigs (21 ± 2 days old, 5.86 ± 0.18 kg body weight) were used in a 2 × 3 factorial design and the main factors included diet (0, 0.60%, or 1.20% HMB) and immunological challenge (LPS or saline). After 15 d of treatment with LPS and/or HMB, growth performance, blood parameters, and muscle protein degradation rate were measured. The results showed that in LPS-injected pigs, 0.60% HMB supplementation increased the average daily gain and average daily feed intake and decreased the feed : gain ratio (P < 0.05), with a concurrent increase of lean percentage. Moreover, 0.60% HMB supplementation decreased the serum concentrations of blood urea nitrogen, IL-1ß, and TNF-α and the rate of protein degradation as well as cell apoptosis in selected muscles (P < 0.05). In addition, dietary HMB supplementation (0.60%) regulated the expression of genes involved in mitochondrial biogenesis and increased the phosphorylation of Akt and Forkhead Box O3a (FoxO3a) in selected muscles, accompanied by decreased protein expression of muscle RING finger 1 and muscle atrophy F-box. These results indicate that HMB may exert protective effects against LPS-induced muscle atrophy by normalizing the Akt/FoxO3a axis that regulates ubiquitin proteolysis and by improving mitochondrial biogenesis.

The influence of doxazosin on the contractility of the urinary bladder in female pigs with experimentally induced cystitis.

The present in vitro study investigated the influence of doxazosin on the contractility of the urinary bladder in female pigs with experimentally induced cystitis. Fifteen juvenile female piglets (18-20 kg body weight) were randomly assigned into three groups (n=5 animals each): i) control (clinically healthy animals, without doxazosin treatment), ii) animals with induced inflammation of the urinary bladder, but without doxazosin treatment (experimental group I) and iii) animals with inflamed bladder, treated orally with doxazosin (0.1 mg/kg body weight for 30 days; experimental group II). Thereafter, the pigs were sacrificed and strips of the bladder trigone were suspended in organ baths. The tension and amplitude of the smooth muscles was measured before and after exposition to 5-hydroxytryptamine (5-HT; 10-6-10-4 M), acetylocholine (ACh; 10-5-10-3 M) and norepinephrine (NE; 10-9-10-7 M). 5-HT caused an increase in the tension of contractions in all the groups and the amplitude in the experimental groups, however, the effect was higher in the experimental group I than in group II as compared to that found in the pre-treatment period. ACh caused an increase in the tension in the control group and a decrease in the amplitude in both experimental groups; these changes significantly differed between the control and doxazosin-treated group. NE caused a decrease in the tension in both experimental groups and amplitude in all the groups, however, the effect was most strongly expressed in doxazosine-treated group. The present study has revealed that long-term administration of doxazosin causes a desensitization of the detrusor smooth muscle to in vitro applied mediators in the autonomic nervous system.

Overexpression of pig selenoprotein S blocks OTA-induced promotion of PCV2 replication by inhibiting oxidative stress and p38 phosphorylation in PK15 cells.

Porcine circovirus type 2 (PCV2) is the primary cause of porcine circovirus disease, and ochratoxin A (OTA)-induced oxidative stress promotes PCV2 replication. In humans, selenoprotein S (SelS) has antioxidant ability, but it is unclear whether SelS affects viral infection. Here, we stably transfected PK15 cells with pig pCDNA3.1-SelS to overexpress SelS. Selenium (Se) at 2 or 4 µM and SelS overexpression blocked the OTA-induced increases of PCV2 DNA copy number and infected cell numbers. SelS overexpression also increased glutathione (GSH), NF-E2-related factor 2 (Nrf2) mRNA, and γ-glutamyl-cysteine synthetase mRNA levels; decreased reactive oxygen species (ROS) levels; and inhibited p38 phosphorylation in PCV2-infected PK15 cells, regardless of OTA treatment. Buthionine sulfoximine reversed all of the above SelS-induced changes. siRNA-mediated SelS knockdown decreased Nrf2 mRNA and GSH levels, increased ROS levels, and promoted PCV2 replication in OTA-treated PK15 cells. These data indicate that pig SelS blocks OTA-induced promotion of PCV2 replication by inhibiting the oxidative stress and p38 phosphorylation in PK15 cells.

Porcine glucagon-like peptide-2 microspheres ameliorate inflammation in lipopolysaccharide-challenged weaning piglets.

This study aimed to investigate the effects of porcine [gly2] glucagon-like peptide-2 (p[gly2]GLP-2) microspheres on lipopolysaccharide-challenged piglets and to evaluate efficacy of microspheres for administration compared with more conventional administration. Eighteen 21-d-old Duroc female piglets were randomly assigned into 3 groups: the control group (intraperitoneal injection with 3 mL saline solution daily), the glucagon-like peptide-2 (GLP-2) group (intraperitoneal injection with 3 mL p[gly2]GLP-2 at 20 nmol/kg BW daily), and the microsphere (MS) group (intraperitoneal injection with 100 mg p[gly2]GLP-2 microsphere suspension at Day 1). On Day 8, all piglets were injected with 100 µg lipopolysaccharide/kg BW. Results showed that administration of p[gly2]GLP-2 microspheres decreased the -lactic acid and methane dicarboxylic aldehyde content of the serum and increased the villus height and villus crypt ratio in the duodenum and ileum. Inducible nitric oxide synthase activity in the duodenum and ileum decreased, whereas enzyme activity for sucrose and Na-K adenosine triphosphatase in the ileum increased with treatment of p[gly2]GLP-2 microspheres. In the MS group, we observed downregulation of IL-8, TNF-α, IFN-γ, and GLP-2R mRNA expression in the ileum, upregulation of positive cell expression in the duodenum and positive cell expression in the ileum, and downregulation of GLP-2 receptor positive cell expression in the ileum. One injection of p[gly2]GLP-2 microspheres was as effective as p[gly2]GLP-2 administered for 7 d. Results suggested that p[gly2]GLP-2 can be a candidate agent for ameliorating weaning stress in piglets and that the use of microspheres is an ideal delivery system for GLP-2.

Modulation by gamithromycin and ketoprofen of in vitro and in vivo porcine lipopolysaccharide-induced inflammation.

The immunomodulatory properties of gamithromycin (GAM), ketoprofen (KETO) and their combination (GAM-KETO) were investigated after both in vitro and in vivo lipopolysaccharide (LPS)-induced inflammation. The influence of these drugs was measured on the production of prostaglandin E2 (PGE2) and the pro-inflammatory cytokines tumour necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1ß in both LPS-stimulated porcine peripheral blood mononuclear cells (PBMCs) and LPS-challenged pigs. Additionally, effects on the production of acute phase proteins (APPs), including pig major acute phase protein (pig-MAP) and C-reactive protein (CRP), as well as on the development of fever, pulmonary symptoms and sickness behaviour were investigated. Dexamethasone was included as a positive control in the in vitro research. Following an 18h-incubation period with 1.25µg/mL LPS, the levels of TNF-α, IL-1ß and IL-6 (p<0.05) measured in the PBMC supernatants were significantly increased. Incubation with a high concentration of both GAM and KETO significantly reduced the in vitro levels of all three cytokines. Maximal plasma concentrations of TNF-α and IL-6 were observed at 1h and 2.5h following LPS challenge in pigs, respectively. Neither GAM, nor KETO nor the combination GAM-KETO was able to inhibit the in vivo LPS-induced cytokine production. Furthermore, none of the drugs influenced the subsequent APPs production. In contrast, administration of KETO significantly reduced PGE2 production both in vitro and in vivo (p<0.05 and p<0.001, respectively) and prevented the development of fever and severe symptoms, including dyspnoea, anorexia, vomiting and lateral decubitus.

Diosmectite-zinc oxide composite improves intestinal barrier restoration and modulates TGF-ß1, ERK1/2, and Akt in piglets after acetic acid challenge.

The present study evaluated the beneficial effect of diosmectite-zinc oxide composite (DS-ZnO) on improving intestinal barrier restoration in piglets after acetic acid challenge and explored the underlying mechanisms. Twenty-four 35-d-old piglets (Duroc × Landrace × Yorkshire), with an average weight of 8.1 kg, were allocated to 4 treatment groups. On d 1 of the trial, colitis was induced via intrarectal injection of acetic acid (10 mL of 10% acetic acid [ACA] solution for ACA, DS-ZnO, and mixture of diosmectite [DS] and ZnO [DS+ZnO] groups) and the control group was infused with saline. Twenty-four hours after challenged, piglets were fed with the following diets: 1) control group (basal diet), 2) ACA group (basal diet), 3) DS-ZnO group (basal diet supplemented with DS-ZnO), and 4) DS+ZnO group (mixture of 1.5 g diosmectite [DS]/kg and 500 mg Zn/kg from ZnO [equal amount of DS and ZnO in the DS-ZnO treatment group]). On d 8 of the trial, piglets were sacrificed. The results showed that DS-ZnO supplementation improved (P < 0.05) ADG, ADFI, and transepithelial electrical resistance and decreased (P < 0.05) fecal scores, crypt depth, and fluorescein isothiocyanate-dextran 4 kDa (FD4) influx as compared with ACA group. Moreover, DS-ZnO increased (P < 0.05) occludin, claudin-1, and zonula occluden-1 expressions; reduced (P < 0.05) caspase-9 and caspase-3 activity and Bax expression; and improved (P < 0.05) Bcl2, XIAP, and PCNA expression. Diosmectite-zinc oxide composite supplementation also increased (P < 0.05) TGF-ß1 expression and ERK1/2 and Akt activation. These results suggest that DS-ZnO attenuates the acetic acid-induced colitis by improving mucosa barrier restoration, inhibiting apoptosis, and improving intestinal epithelial cells proliferation and modulation of TGF-ß1 and ERK1/2 and Akt signaling pathway.

Individual and combined effects of Fusarium toxins on the mRNA expression of pro-inflammatory cytokines in swine jejunal epithelial cells.

Fusarium toxins have been arousing public interest in recent years because of their potential health hazards for humans and agricultural livestock. It was hypothesized that selected pro-inflammatory cytokines might serve as sensitive biomarkers of the predicted adverse effects of Fusarium toxins on the basis of their potential ability to induce immune and intestinal alterations comparable to those in human chronic inflammatory infection. Consequently, the aim of this study was to elucidate individual and combined effects of four common Fusarium toxins, deoxynivalenol (DON), nivalenol (NIV), zearalenone (ZEA) and fumonisin B1 (FB1) on the mRNA expression of pro-inflammatory cytokines (IL1α, IL1ß, IL6, IL8, TNFα and MCP-1) using a porcine jejunal epithelial cell line, IPEC-J2. Based on a dose-response relationship between individual mycotoxins and cell viability (MTT assay) that was previously established, cytotoxic and non-cytotoxic concentrations were selected to investigate combinations of two, three and all four of the mycotoxins. In general, up-regulation of pro-inflammatory cytokine mRNA expression occurred for both individual and mixtures of Fusarium toxins at cytotoxic concentrations, whereas significant up-regulation of pro-inflammatory cytokine mRNA mostly obtained when the toxins existed in mixtures at non-cytotoxic concentrations and these mixtures were found to cause cytotoxicity from MTT assay determined previously. Therefore, it may be concluded that some of the changes in the mRNA expression of IL1α, IL1ß, IL6, IL8, TNFα and MCP-1 could be cytotoxicity-related. It was also noted that additive effects were not always observed for the mixtures. These data suggest that individual or mixtures of Fusarium toxins could cause or exacerbate intestinal inflammation. These also provide a better understanding of the possible effects of Fusarium toxins, alone or in combinations on the immunological defense mechanisms of IECs, which would contribute to the risk assessment of these toxins.

Late gestational hyperprolactinemia accelerates mammary epithelial cell differentiation that leads to increased milk yield.

The growth rate of piglets is limited by sow milk yield, which reflects the extent of epithelial growth and differentiation in the mammary glands (MG) during pregnancy. Prolactin (PRL) promotes both the growth and differentiation of the mammary epithelium, where the lactational success of pigs is absolutely dependent on PRL exposure during late gestation. We hypothesized that inducing hyperprolactinemia in primiparous gilts during late gestation by administering the dopamine antagonist domperidone (DOM) would increase MG epithelial cell proliferation and differentiation, subsequent milk yield, and piglet growth. A total of 19 Yorkshire-Hampshire gilts were assigned to receive either no treatment (CON, n = 9) or DOM (n = 10) twice daily from gestation d 90 to 110. Serial blood sampling during the treatment period and subsequent lactation confirmed that plasma PRL concentrations were increased in DOM gilts on gestation d 91 and 96 (P < 0.001). Piglets reared by DOM-treated gilts gained 21% more BW during lactation than controls (P = 0.03) because of increased milk production by these same gilts on d 14 (24%, P = 0.02) and 21 (32%, P < 0.001) of lactation. Milk composition did not differ between the 2 groups on d 1 or 20 of lactation. Alveolar volume within the MG of DOM-treated gilts was increased during the treatment period (P < 0.001), whereas epithelial proliferation was unaffected by treatment. Exposure to DOM during late gestation augmented the postpartum increase in mRNA expression within the MG for ß-casein (P < 0.03), acetyl CoA carboxylase-α (P < 0.01), lipoprotein lipase (P < 0.06), α-lactalbumin (P < 0.08), and glucose transporter 1 (P < 0.06). These findings demonstrate that late gestational hyperprolactinemia enhances lactogenesis within the porcine MG and increases milk production in the subsequent lactation.

Porcine intestinal epithelial barrier disruption by the Fusarium mycotoxins deoxynivalenol and T-2 toxin promotes transepithelial passage of doxycycline and paromomycin.

BACKGROUND: The gastrointestinal tract is the first target for the potentially harmful effects of mycotoxins after intake of mycotoxin contaminated food or feed. With deoxynivalenol (DON), T-2 toxin (T-2), fumonisin B1 (FB1) and zearalenone (ZEA) being important Fusarium toxins in the northern hemisphere, this study aimed to investigate in vitro the toxic effect of these mycotoxins on intestinal porcine epithelial cells derived from the jejunum (IPEC-J2 cells). Viability of IPEC-J2 cells as well as the proportion of apoptotic and necrotic IPEC-J2 cells was determined by flow cytometry after 72 h of exposure to the toxins. Correlatively, the integrity of the intestinal epithelial cell monolayer was studied using Transwell(®) inserts, in which the trans-epithelial electrical resistance (TEER) and passage of the antibiotics doxycycline and paromomycin were used as endpoints. RESULTS: We demonstrated that the percentage of Annexin-V-FITC and PI negative (viable) cells, Annexin-V-FITC positive and PI negative (apoptotic) cells and Annexin-V-FITC and PI positive (necrotic) IPEC-J2 cells showed a mycotoxin concentration-dependent relationship with T-2 toxin being the most toxic. Moreover, the ratio between Annexin-V-FITC positive and PI negative cells and Annexin-V-FITC and PI positive cells varied depending on the type of toxin. More Annexin-V-FITC and PI positive cells could be found after treatment with T-2 toxin, while more Annexin-V-FITC positive and PI negative cells were found after exposure to DON. Consistent with the cytotoxicity results, both DON and T-2 decreased TEER and increased cellular permeability to doxycycline and paromomycin in a time- and concentration-dependent manner. CONCLUSIONS: It was concluded that Fusarium mycotoxins may severely disturb the intestinal epithelial barrier and promote passage of antibiotics.

Anaesthetic complications in pigs undergoing MRI guided convection enhanced drug delivery to the brain: a case series.

OBSERVATIONS: A total of 13 intracerebral infusions were performed at approximately 1 month intervals in three NIH miniature pigs over the age range of 31-59 weeks. Pigs received azaperone and ketamine premedication to allow venous cannulation and propofol induction of anaesthesia. Anaesthesia was maintained with isoflurane throughout cranial surgery and MRI scanning. Physiological monitoring during surgery consisted of blood pressure, pulse, temperature and oxygen saturation monitoring, ECG and capnography. Analgesia consisted of meloxicam and morphine. However, during MRI scanning blood pressure and ECG monitoring had to be discontinued. Anaesthetized pigs underwent intermittent intraputamenal convection enhanced delivery (CED) of gadolinium with real-time magnetic resonance imaging. Progressive tachycardia was consistently observed in all pigs during CED with a mean ± SD maximum increase of 41 ± 22 beats minute(-1) from a baseline heart rate of 96 ± 9 minute(-1) . The heart rate remained elevated until recovery. A mean reduction in body temperature of 2.8 ± 0.6 °C from the start of anaesthesia was also observed during the period of MRI scanning. All pigs recovered from anaesthesia smoothly and heart rates returned to normal during the recovery period. CONCLUSIONS: Hypothermia is common in pigs undergoing this sedation and anaesthesia protocol. Convection enhanced delivery of drugs in healthy anaesthetized pigs may result in tachycardia.

Effect of dietary fermented garlic by Weissella koreensis powder on growth performance, blood characteristics, and immune response of growing pigs challenged with Escherichia coli lipopolysaccharide.

Two experiments were conducted to investigate the effect of fermented garlic by Weissella koreensis powder (WKG) on pig growth performance and immune responses after an Escherichia coli lipopolysaccharide (LPS) challenge. In Exp. 1, 120 growing barrows (23.5 ± 0.5 kg of BW and 56 d of age) were used in a 35-d experiment to determine the optimal amounts of WKG. Pigs were randomly allotted to 1 of 5 treatments with 6 replicate pens and 4 pigs per pen. Dietary treatments included 1) NC (negative control; basal diet without antibiotics), 2) PC (positive control; basal diet + 1 g of tylosin/kg), 3) WKG1 (basal diet + 1 g of WKG/kg), 4) WKG2 (basal diet + 2 g of WKG/kg), and 5) basal diet + 4 g of WKG/kg. At the end of the feeding period, 12 pigs each were selected from the NC and WKG2 treatment groups, and 6 pigs were injected with LPS (50 µg/kg of BW) and the other 6 pigs with an equivalent amount of sterile saline, resulting in a 2 × 2 factorial arrangement of treatments. Blood samples and rectal temperature data were collected at 0, 2, 4, 6, 8, and 12 h after challenge. The ADG of pigs fed WKG- and antibiotic-supplemented diets was greater (P<0.05) than NC from d 14 to 35 and the overall phase, but no dosage-dependent effects were observed. At the end of the experiment, the fecal E. coli count was linearly reduced by the increasing amounts of WKG at d 35 (P=0.01). Challenge with LPS increased white blood cell counts at 6 and 8 h (P<0.01) and depressed lymphocyte concentration at 4, 8, and 12 h (P<0.01). During challenge, LPS injection increased rectal temperature at 2, 4, 6, and 8 h postchallenge (P<0.05), and WKG2 alleviated (P<0.05) the increase in the temperature at 2 h postchallenge. The LPS injection increased plasma tumor necrosis factor-α and IGF-1 concentrations at 2, 4, 6, 8, and 12 h (P<0.01), whereas an alleviating effect of WKG was observed at 4, 6, and 8 h after LPS challenge (P<0.05). At 2, 4, and 6 h postchallenge, concentration of cluster of differentiation-antigen-4-positive cells and cluster of differentiation-antigen-8-positive cells (CD4(+) and CD8(+), respectively) increased in the LPS treatments (P<0.05), and the WKG2 boosted this effect (P<0.05). In conclusion, dietary supplementation of WKG2 in growing pigs can improve ADG and have a beneficial effect on the immune response during an inflammatory challenge.

The effects of acute exposure to deoxynivalenol on some inflammatory parameters in miniature pigs.

Seven miniature pigs were injected intravenously with deoxynivalenol (DON) at 1 mg/kg body weight; afterward, the number of leukocytes in peripheral blood, the luminol-dependent chemiluminescence of neutrophils, the serum or plasma concentration of cytokines and acute-phase proteins were evaluated to determine the effects of acute exposure to DON on inflammatory responses. White blood cell counts were transiently increased at 3, 6, and 12 hr post-injection (PI) due to the increased number of neutrophils. The luminol-dependent chemiluminescence value of neutrophils was significantly elevated at 24 hr PI, indicating the activation of the bactericidal function of neutrophils. Significant increases of interleukin (IL)-8 and tumor necrosis factor-α at 3 hr PI and IL-6 at 6 hr PI were detected in the serum. The concentration of haptoglobin and serum amyloid A was significantly increased at 24 hr PI. These results suggest that acute exposure to DON induced a temporary recruitment of neutrophils in the peripheral blood by IL-8 and subsequent activation of the bactericidal function, and a transient increase of proinflammatory cytokines and acute-phase proteins, indicating the immunomodulatory effects of DON in pigs.