interleukin-8 and Swine-Diseases

The stress caused by sound is inevitable. The stress caused by noise and the positive effects of music can affect the endocrine of animals and their welfare. In this study, a total of 72 hybrid piglets (Large White × Duroc × Min pig) were randomly divided into 3 groups, including music (Mozart K.448, 60-70 dB), noise (recorded mechanical noise, 80-85 dB), and control (natural background sound, <40 dB) groups. S-IgA (secretory immunoglobulin A), IL-6 (interleukin-6), IL-8 (interleukin-8), and positive emotion-related behaviors were used as indicators to discuss whether noise induced stress and inflammation in piglets or whether music could have positive effects. Six hours of auditory exposure were given daily (10:00-16:00), which lasted for 56 days. Behavioral responses of the piglets were observed, and the concentrations of salivary S-IgA and serum IL-6 and IL-8 were measured. The results showed that the concentration of S-IgA increased in the noise and control groups on the 57th day (P < 0.05); S-IgA concentration in the music group was unchanged after long-term music exposure. The concentrations of IL-6 and IL-8 showed that long-term noise exposure might lead to stress and inflammation in piglets. Tail-wagging and play behaviors of the piglets in the music group were significantly greater than those in the noise and control groups, which implied that long-term music exposure improved the emotional state of the piglets in a restricted and barren environment.

Topics: Animals; Emotions; Immunoglobulin A; Inflammation; Interleukin-6; Interleukin-8; Swine; Swine Diseases

Streptococcus suis serotype 2 (SS2) frequently colonizes the swine upper respiratory tract and can cause Streptococcal disease in swine with clinical manifestations of pneumonia, meningitis, and septicemia. Previously, we have shown that vimentin, a kind of intermediate filament protein, is involved in the penetration of SS2 through the tracheal epithelial barrier. The initiation of invasive disease is closely related to SS2-induced excessive local inflammation; however, the role of vimentin in airway epithelial inflammation remains unclear. Here, we show that vimentin deficient mice exhibit attenuated lung injury, diminished production of proinflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and the IL-8 homolog, keratinocyte-derived chemokine (KC), and substantially reduced neutrophils in the lungs following intranasal infection with SS2. We also found that swine tracheal epithelial cells (STEC) without vimentin show decreased transcription of IL-6, TNF-α, and IL-8. SS2 infection caused reassembly of vimentin in STEC, and pharmacological disruption of vimentin filaments prevented the transcription of those proinflammatory cytokines. Furthermore, deficiency of vimentin failed to increase the transcription of nucleotide oligomerization domain protein 2 (NOD2), which is known to interact with vimentin, and the phosphorylation of NF-κB protein p65. This study provides insights into how vimentin promotes excessive airway inflammation, thereby exacerbating airway injury and SS2-induced systemic infection.

Topics: Animals; Cytokines; Epithelium; Inflammation; Interleukin-6; Interleukin-8; Intermediate Filaments; Mice; Neutrophil Infiltration; Serogroup; Streptococcal Infections; Streptococcus suis; Swine; Swine Diseases; Trachea; Tumor Necrosis Factor-alpha; Vimentin

The effect of a water-soluble formulation of tylvalosin (Aivlosin® 625 mg/g granules) on disease caused by porcine reproductive and respiratory syndrome virus (PRRSV) and Mycoplasma hyopneumoniae (Mhyop) was investigated in two animal studies. In a PRRSV challenge model in pregnant sows (n = 18), six sows received water medicated at target dose of 5 mg tylvalosin/kg body weight/day from 3 days prior to challenge until the end of gestation. Six sows were left untreated, with a third group remaining untreated and unchallenged. Sows were challenged with PRRSV-2 at approximately 85 days of gestation. Cytokines, viremia, viral shedding, sow reproductive parameters and piglet performance to weaning were evaluated. In a dual infection study (n = 16), piglets were challenged with Mhyop on days 0, 1 and 2, and with PRRSV-1 on day 14 and euthanized on day 24. From day 10 to 20, eight piglets received water medicated at target dose of 20 mg tylvalosin/kg body weight/day and eight piglets were left untreated. Cytokines, viremia, bacteriology and lung lesions were evaluated.. Overall, tylvalosin reduced both local and systemic proinflammatory cytokines after challenge with respiratory pathogens in sows and in piglets. Tylvalosin was effective in reducing Mhyop recovery from the lungs and may reduce virus shedding in piglets following transplacental PRRSV infection in sows.

Topics: Animals; Body Weight; Cytokines; Female; Granulocyte-Macrophage Colony-Stimulating Factor; Interleukin-10; Interleukin-12; Interleukin-8; Mycoplasma hyopneumoniae; Porcine Reproductive and Respiratory Syndrome; Porcine respiratory and reproductive syndrome virus; Pregnancy; Swine; Swine Diseases; Tumor Necrosis Factor-alpha; Viremia

Enterotoxigenic Escherichia coli (ETEC) is an important cause of diarrhea in piglets, which leads to great economic losses. In this study, the ternary crossbred weaned piglets were orally administered with 1.5 × 10

Topics: Animals; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Interleukin-13; Interleukin-8; Intestinal Diseases; Intestinal Mucosa; Intestines; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

There have been limited studies focused on validation of swine microRNAs (miRNA) with mRNA targets. The objective of this study was to validate a defined set of targets using artificial miRNA mimics transfected into cell lines to confirm specific targets of endogenous miRNAs after administration of Escherichia coli lipopolysaccharide (LPS). Sixteen hours after mimic transfection of 3D4/21 cell lines, the cells were stimulated with 1 μg/ml LPS or phosphate-buffered saline (PBS). The cells were harvested and collected at 0, 1, 3, and 8 h post administration. The selected genes DAD1, IL8, and ESR, which are involved in known pathways of inflammation. and are predicted or validated human targets of either miR-146a, let-7a, or miR-22-3p. These were then evaluated by quantitative real-time-PCR (qRT-PCR) to verify microRNA-mRNA interaction in swine. Using the ROX reference dye, mRNA changes in expression were assessed using the comparative CT Method (ΔΔCT method) for normalization against the PBS control group. DAD1 and ESR1 were negatively regulated by miR-22-3p and miR-146a-5p, respectively in 3D4/21 cells after LPS stimulation. However, miR-146a-5p may play an indirect positive regulatory role of both DAD1 and IL8 mRNA expression. Furthermore, we found an inverse relationship between LPS stimulation compared with the let-7a-5p overexpression with DAD1. Our inflammation study provides new evidence on the roles and predicted targets of miR-146a, let-7a, and miR-22-3p in swine.

Topics: Animals; Humans; Inflammation; Interleukin-8; Lipopolysaccharides; MicroRNAs; RNA, Messenger; Swine; Swine Diseases

Topics: Animals; Cytoskeletal Proteins; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Interleukin-8; RNA-Seq; RNA, Long Noncoding; Swine; Swine Diseases

Infection induces the production of proinflammatory cytokines and chemokines such as interleukin-8 (IL-8) and interleukin-6 (IL-6). Although they facilitate local antiviral immunity, their excessive release leads to life-threatening cytokine release syndrome, exemplified by the severe cases of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In the present study, we found that interleukin-8 (IL-8) was upregulated by PDCoV infection. We then demonstrated that PDCoV E protein induced IL-8 production and that the TM domain and the C-terminal domain of the E protein were important for IL-8 production. Subsequently, we showed here that deleting the AP-1 and NF-κB binding motif in porcine IL-8 promoter abrogated its activation, suggesting that IL-8 expression was dependent on AP-1 and NF-κB. Furthermore, PDCoV E induced IL-8 production, which was also dependent on the NF-κB pathway through activating nuclear factor p65 phosphorylation and NF-κB inhibitor alpha (IκBα) protein phosphorylation, as well as inducing the nuclear translocation of p65, eventually resulting in the promotion of IL-8 production. PDCoV E also activated c-fos and c-jun, both of which are members of the AP-1 family. These findings provide new insights into the molecular mechanisms of PDCoV-induced IL-8 production and help us further understand the pathogenesis of PDCoV infection.

Topics: Animals; Antiviral Agents; COVID-19; Cytokines; Interleukin-6; Interleukin-8; NF-kappa B; NF-KappaB Inhibitor alpha; SARS-CoV-2; Swine; Swine Diseases; Transcription Factor AP-1

Glässer's disease in pigs is associated with infection by. La maladie de Glässer chez les porcs est associée avec une infection par

Topics: Animals; Anti-Inflammatory Agents; Antiviral Agents; Extracellular Signal-Regulated MAP Kinases; Haemophilus parasuis; Inflammation; Interleukin-6; Interleukin-8; JNK Mitogen-Activated Protein Kinases; Lipopolysaccharides; Macrophages, Alveolar; NF-kappa B; NF-KappaB Inhibitor alpha; Signal Transduction; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

Porcine epidemic diarrhea virus (PEDV) is a re-emerging pathogen that causes severe economic loss in the pig industry. The host's innate immune system is the first line of defense on virus invasion of the small intestinal epithelial cells. Chemokines, as a part of the innate immune system, play an important role in host immunity against infection, however, and their expression and chemotactic effect on key immune cells in PEDV infection remains unclear. In this study, cDNA microarray was firstly performed to analyzed ileum tissue of piglets on the third day after PEDV infection. The differentially expressed genes mainly involved in multiple biological processes, chemokine signaling pathway and cytokine receptor interaction signaling pathway had the highest enrichment according to GO and KEGG enrichment analysis. The expression levels of chemokines MCP-1, MIP-1β, IL-8, CXCL9, CXCL10 and CXCL13 in ileum of PEDV- infected piglets were significantly higher than those in the control group. The expression of chemokines in vivo experiment was further verified by RT-qPCR and ELISA using PEDV-infected IPEC-J2 cells. The results showed that the PEDV-infected IPEC-J2 cells had significantly induced protein expression of MCP-1, MIP-1β, IL-8, CXCL9, CXCL-10 and CXCL13. These results indicated that the changes of chemokines expressed in the ileum of piglets (in vivo) were consistent with those in IPEC-J2 cells (in vitro) after PEDV infection. Finally, the role of chemokines in immune cell migration during PEDV infection was illustrated by siRNA-mediated knock down method and the co-culture model of IPEC-J2 cells with peripheral blood leukocyte cells (PBLCs). The FACS analysis showed that MCP-1 induced by PEDV infection played a chemotactic effect on CD14

Topics: Animals; Cell Line; Chemokine CCL4; Coronavirus Infections; Interleukin-8; Monocytes; Porcine epidemic diarrhea virus; Swine; Swine Diseases

Newly emerging and re-emerging viral infectious diseases cause significant economic losses in swine production. Efficacious vaccines have not yet been developed for several major swine infectious diseases, including porcine epidemic diarrhea virus (PEDV). We used the PEDV-infected Vero cell model to screen lactic acid bacteria (LAB) strains with antiviral activity. Sixty LAB strains were isolated from the feces of nursing piglets. After the elimination of LAB strains with high cytotoxicity to Vero cells, the protective effects of the remaining 6 strains against PEDV infection were determined. Vero cells pretreated with the intracellular extracts or cell wall fractions of YM22 and YM33 strains for 24 h before infection with PEDV showed significantly higher cell viabilities and lower mRNA expression of PEDV nucleocapsid (PEDV-N) than the unpretreated cells, indicating that the intracellular extracts and cell wall fractions of YM22 and YM33 possessed prophylactic effects on Vero cells against PEDV infection. PEDV-infection significantly increased the mRNA expression of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-8 (IL-8) in Vero cells. However, pretreatment of Vero cells with the cell wall fractions of YM22 and YM33 decreased the mRNA expression of TNF-α and IL-8, which could be a mechanism associated with the protective effects of YM22 and YM33 against PEDV. Based on the biochemical characteristics and phylogenetic analyses, YM22 and YM33 were identified as Ligilactobacillus agilis (basonym: Lactobacillus agilis) and Ligilactobacillus salivarius (basonym: Lactobacillus salivarius), respectively. These findings suggest that L. agilis YM22 and L. salivarius YM33 could provide some levels of protective effects against PEDV infections.

Topics: Animals; Antiviral Agents; Chlorocebus aethiops; Coronavirus Infections; Diarrhea; Dysentery; Interleukin-8; Lactic Acid; Lactobacillales; Phylogeny; Plant Extracts; Porcine epidemic diarrhea virus; RNA, Messenger; Swine; Swine Diseases; Tumor Necrosis Factor-alpha; Vero Cells

Mycoplasma hyopneumoniae (Mhp) is the primary pathogen of porcine enzootic pneumonia (PEP). Consolidated lung tissue from the cranioventral lung lobes of 15 pigs with PEP was collected for quantitative polymerase chain reaction, histopathology and immunohistochemistry. Histopathology revealed the co-existence of bronchial-associated lymphoid tissue hyperplasia with intra-alveolar neutrophils and macrophage infiltration in lesions of suppurative bronchopneumonia. Immunolabelling of infiltrated macrophages with CD163/CD204 indicated the presence of M2-polarized macrophages. Mhp antigen was detected on respiratory epithelial cells and in phagocytosed neutrophils. The intensity of Mhp immunolabelling and number of CD163/CD204-positive macrophages were correlated with the Mhp load in lung tissue (r = 0.87, 0.56, P <0.05). IL-8 immunolabelling was mainly found in neutrophils and correlated with Mhp load, Mhp immunolabelling and histological lesion score (r = 0.70, 0.66, 0.64, P <0.05), respectively. Apoptosis was seen in intra-alveolar cells and was correlated with Mhp load (r = 0.62, P <0.05). It is postulated that IL-8 attracts neutrophils to the lesions, while M2-polarized macrophages are a major source of IL-10 and promote a Th2-type immune response.

Topics: Animals; Apoptosis; Interleukin-8; Macrophages; Mycoplasma hyopneumoniae; Neutrophils; Pneumonia of Swine, Mycoplasmal; Swine; Swine Diseases

Porcine hemagglutinating encephalomyelitis virus (PHEV) is a betacoronavirus that causes vomiting and wasting disease and/or encephalomyelitis in suckling pigs. This study characterized PHEV infection, pathogenesis, and immune response in cesarean-derived, colostrum-deprived (CDCD) neonatal pigs. Infected animals developed mild respiratory, enteric, and neurological clinical signs between 2 to 13 days postoronasal inoculation (dpi). PHEV did not produce viremia, but virus shedding was detected in nasal secretions (1 to 10 dpi) and feces (2 to 7 dpi) by reverse transcriptase quantitative PCR (RT-qPCR). Viral RNA was detected in all tissues except liver, but the detection rate and RT-qPCR threshold cycle (

Topics: Animals; Betacoronavirus 1; Cell Line; Coronavirus Infections; Interferon-alpha; Interleukin-8; Organ Specificity; Reverse Transcriptase Polymerase Chain Reaction; Swine; Swine Diseases; T-Lymphocytes

Dysfunction of endothelial cells and vascular system is one of the most important pathological changes of porcine circovirus disease (PCVD) caused by porcine circovirus type 2 (PCV2). PCV2-infected endothelial cells can upregulate the production of endothelial-derived IL-8, which can inhibit the maturation of dendritic cells. Endothelial-derived IL-8 has different structural and biological characteristics compared with monocyte-derived IL-8. However, the mechanism of endothelial-derived IL-8 production is still unclear.. Key molecules of RIG-I-like signaling pathway RIG-I, MDA-5, MAVS and a key molecule of JNK signaling pathway c-Jun in PCV2-infected porcine iliac artery endothelial cells (PIECs) were upregulated significantly detected with quantitative PCR, Western blot and fluorescence confocal microscopy, while no significant changes were found in NF-κB signaling pathway. Meanwhile, the expression of endothelial-derived IL-8 was downregulated after RIG-I, MDA-5, or MAVS genes in PIECs were knocked down and PIECs were treated by JNK inhibitor.. PCV2 can activate RIG-I/MDA-5/MAVS/JNK signaling pathway to induce the production of endothelial-derived IL-8 in PIECs, which provides an insight into the further study of endothelial dysfunction and vascular system disorder caused by PCV2.

Topics: Animals; Cells, Cultured; Circoviridae Infections; Circovirus; Endothelial Cells; Gene Knockdown Techniques; Iliac Artery; Interleukin-8; Signal Transduction; Swine; Swine Diseases

Chlamydia suis intestinal infection of single-animal experimental groups of gnotobiotic newborn piglets was previously reported to cause severe, temporary small intestinal epithelium damage. We investigated archived intestinal samples for pro-inflammatory nuclear factor kappa B (NF-κB) activation, Interleukin (IL)-6 and IL-8 production and immune cell influx. Samples were collected 2, 4 and 7 days post-inoculation with C. suis strain S45/6 or mock inoculum (control). Increased nuclear localization of epithelial NF-κB, representative of activation, in the jejunum and ileum of C. suis-infected animals, compared to uninfected controls, began by 2 days post-infection (dpi) and persisted through 7 dpi. Infected animals showed increased production of IL-8, peaking at 2 dpi, compared to controls. Infection-mediated CD45-positive immune cell influx into the jejunal lamina propria peaked at 7 dpi, when epithelial damage was largely resolved. Activation of NF-κB appears to be a key early event in the innate response of the unprimed porcine immune system challenged with C. suis. This results in an acute phase, coinciding with the most severe clinical symptoms, diarrhea and weight loss. Immune cells recruited shortly after infection remain present in the lamina propria during the recovery phase, which is characterized by reduced chlamydial shedding and restored intestinal epithelium integrity.

Topics: Animals; Chlamydia; Chlamydia Infections; Diarrhea; Feces; Germ-Free Life; Host-Pathogen Interactions; Immunity, Cellular; Immunohistochemistry; Interleukin-6; Interleukin-8; Intestinal Mucosa; Models, Animal; NF-kappa B; Swine; Swine Diseases

Porcine epidemic diarrhea virus (PEDV) causes severe economic loss in the pig industry each year. To better understand the relationship between cytokines and PEDV replication, in this study, pro-inflammatory cytokine and chemokine expression profiles in Vero cells infected with PEDV were analyzed. Real-time quantitative PCR assay indicated that IL-1α, IL-1β, TNF-α, CCL2, CCL5 and CXCL8 expression levels were significantly upregulated. Moreover, overexpression and siRNA silencing assays showed that overexpression of IL-1α, IL-1β, TNF-α, CCL2, CCL5 and CXCL8 could significantly inhibit PEDV replication, while silencing of IL-1α, IL-1β, TNF-α, CCL2, CCL5 and CXCL8 could significantly promote PEDV replication. Finally, a dual-luciferase reporter assay showed that nsp4 contributed to the expression of IL-1α, IL-1β, TNF-α, CCL2, CCL5 and CXCL8 via the NF-κB pathway. Together, these data determined that PEDV nsp4 could upregulate pro-inflammatory cytokine and chemokine expression, inhibiting viral replication in vitro. These results provided novel insights for understanding the roles of cytokines in PEDV replication.

Topics: Animals; Chemokine CCL2; Chemokine CCL5; Chemokines; Chlorocebus aethiops; Coronavirus Infections; Cytokines; Host-Pathogen Interactions; Interleukin-1beta; Interleukin-8; NF-kappa B; Porcine epidemic diarrhea virus; Swine; Swine Diseases; Tumor Necrosis Factor-alpha; Vero Cells; Viral Nonstructural Proteins; Virus Replication

Mammary lesions in sows can prevent suckling piglets from consuming colostrum that provides fundamental nutrients and protective immunity. Although mammary gross lesions are frequently found in sows at farms or slaughterhouses, with the exception of mastitis, they have received little research attention. In this study, we investigated mammary lesions observed in South Korean sows between 2015 and 2016. Mammary tissue samples of 82 sows showing gross lesions during meat inspection were histologically classified and immunohistochemical analysis was conducted to assess the expression of estrogen receptor (ER)-α, ER-β, and progesterone receptor (PR) for mammary hyperplastic lesions as well as that of cluster of differentiation (CD) 3, CD79a, interleukin (IL)-1α, IL-1β, IL-6, and IL-8 for mastitis. Furthermore, 20 swab samples were cultured, and the isolated bacteria were identified using polymerase chain reactions for 16S ribosomal RNA genes. The lesions were classified as hyperplasia, mastitis, or hyperplasia with mastitis. Immunohistochemistry results revealed that there was neither expression of ER-α nor of ER-β, but all examined hyperplastic samples expressed PR. In addition, there was a significant correlation between CD3 and IL-1β expressions, as well as between IL-1β and IL-6 expressions. Regarding the identity of the isolated bacteria,

Topics: Abattoirs; Animals; Breast Diseases; CD3 Complex; CD79 Antigens; Cytokines; Estrogen Receptor alpha; Estrogen Receptor beta; Female; Interleukin-1alpha; Interleukin-1beta; Interleukin-6; Interleukin-8; Mammary Glands, Animal; Mastitis; Pseudomonas; Receptors, Estrogen; Receptors, Progesterone; Swine; Swine Diseases

In the swine industry, Lawsonia intracellularis is one of the main enteric pathogens; it causes acute intestinal hemorrhage (proliferative hemorrhagic enteropathy) in naïve adult pigs and a wasting disease (proliferative enteropathy) in growing pigs. Among many kinds of cytokines, interferon-γ (IFN-γ) has previously been reported to play a significant role in limiting intracellular infection and increasing cellular proliferation associated with L. intracellularis. However, the levels of various circulating inflammatory cytokines, including IFN-γ, in animals infected with L. intracellularisis is still an area of considerable interest for understanding immunity against this bacterium. In addition, there has been no information on cytokine response in animals infected with any L. intracellularis isolate of South Korean origin or Asian origin. To determine the relationship between the changes in the systemic inflammatory cytokine response in the peripheral blood of the host after L. intracellularis infection, we measured the levels of some pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IFN-γ), anti-inflammatory cytokines (IL-4, IL-10, and transforming growth factor-β (TGF-β)), and a chemokine (IL-8) in pigs infected with L. intracellularis isolated from South Korea. This study demonstrated that a L. intracellularis isolate of South Korean origin induced cytokine (TNF-α, IL-6, and IFN-γ) responses in infected animals within 15 days post-infection although the circulating levels of IL-4, IL-10, IL-8 and TGF-β were induced relatively late.

Topics: Animals; Cytokines; Desulfovibrionaceae Infections; Feces; Interleukin-8; Intestinal Diseases; Lawsonia Bacteria; Republic of Korea; Swine; Swine Diseases

The obligate intracellular Lawsonia intracellularis (LI), the etiological agent of proliferative enteropathy (PE), is an economically important disease in the swine industry. Due to extreme difficulty of in vitro culture of the pathogen, molecular characterization of protein components of LI that are targets of the immune system, is difficult; thus, the scientific evidence to drive the development of preventive measures is lacking. In this work, we investigated the antigenic and functional characteristics of a putative flagellar-associated protein, LI0570, using in silico computational approaches for epitope prediction and an in vitro protein-based molecular assay. The amino acid sequence of LI0570 exhibited similarities to flagellar-associated proteins in four different bacterial strains. The presence of B cell linear confirmative epitopes of the protein predicted by a bioinformatics tool was validated by western blot analysis using anti-LI mouse hyperimmune serum, which implied that LI0570 induced production of antigen-specific antibodies in vivo. Further, TLR5-stimulating activity and IL-8 cytokine expression produced via downstream signaling were observed in HEK-Blue™-hTLR5 cells stimulated with LI0570. This result indicates that the LI0570 protein can trigger an innate immune response followed by a T-cell-related adaptive immune response in an infected host. Collectively, the data presented here support that the LI0570 protein which shows the antigenic potential could be a useful component of a recombinant vaccine against PE, providing progress toward an effective prevention strategy.

Topics: Adjuvants, Immunologic; Amino Acid Sequence; Animals; Desulfovibrionaceae Infections; Flagellin; HEK293 Cells; Humans; Interleukin-8; Lawsonia Bacteria; Sequence Alignment; Swine; Swine Diseases; Toll-Like Receptor 5

Porcine cytomegalovirus (PCMV) causes lifelong latent infections in swine. The pathogen is occasionally associated with inclusion body rhinitis and pneumonia in piglets, reproductive disorders in pregnant sows and respiratory disease complex in older pigs. Immunosuppressive potential of PCMV infection is discussed. Macrophages were recognised as one of target cell types where propagation of virus occurs. The aim of present study was to set up model PCMV infection of monocyte derived macrophages (MDMs) in vitro for PCMV immunobiology research. Obtained results showed that PCMV is able to infect and propagate in MDMs. Possible immunosuppressive effect of PCMV on infected macrophages was evaluated by measurement of immune relevant gene expression in MDMs. Infection decreased expression of IL-8 and TNF-α (pro-inflammatory cytokines) and increased expression of IL-10 (anti-inflammatory cytokine) on mRNA transcription level. Obtained data support hypothesis that higher sensitivity of animals to coinfection with other swine pathogens and its more severe clinical manifestations could potentially be the consequence of PCMV infection.

Topics: Animals; Cytokines; Cytomegalovirus; Cytomegalovirus Infections; Gene Expression; Immunity, Innate; Interleukin-10; Interleukin-8; Macrophages; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

The objective of this study was twofold: (i) to examine the effect of Clostridium butyricum on intestinal barrier function and (ii) to elucidate the mechanisms involved in enhanced intestinal barrier function.. Forty-eight weaned piglets were assigned randomly to either a basal diet or a C. butyricum-supplemented diet. On day 15, all pigs were orally challenged with enterotoxigenic Escherichia coli (ETEC) K88 or saline. Clostridium butyricum decreased serum diamine oxidase activity and d-lactic acid concentration, as well as increased intestinal tight junction proteins (ZO-1, claudin-3 and occludin) expression in ETEC K88-infected pigs. Moreover, C. butyricum decreased IL-1β and IL-18 levels in serum and gut, whereas it increased IL-10 levels. Furthermore, C. butyricum downregulated NLRP3 and caspase-1 expression in ETEC K88-challenged pig gut, but did not affect apoptosis-associated speck-like protein expression.. Clostridium butyricum enhanced intestinal barrier function and inhibited apoptosis-associated speck-like protein-independent NLRP3 inflammasome signalling pathway in weaned piglets after ETEC K88 challenge.. The novelty of this study lies in the beneficial effects of C. butyricum on intestinal health, likely by improving intestinal barrier function and alleviating inflammatory reactions.

Topics: Animal Feed; Animals; Clostridium butyricum; Diet; Dietary Supplements; Enterotoxigenic Escherichia coli; Escherichia coli Infections; Female; Interleukin-1beta; Interleukin-8; Intestinal Mucosa; Intestines; Male; Probiotics; Swine; Swine Diseases; Weaning

Gastric mRNA expression of markers for acid secretion and inflammation and presence of gastric ulceration was studied in naturally Helicobacter suis-infected and non-infected 2-3 months old, 6-8 months old and adult pigs. In H. suis-infected 2-3 months old pigs, IL-8 and IL-1β transcript levels were upregulated in the pyloric gland zone, indicating an innate immune response. A similar response was demonstrated in the fundic gland zone of adult pigs, potentially due to a shift of H. suis colonization from the pyloric to the fundic gland zone. A Treg response in combination with decreased expressions of IL-8, IL-17A and IFN-γ was indicated to be present in the H. suis-infected 6-8 months old pigs, which may have contributed to persistence of H. suis. In H. suis-infected adult pigs, a Treg response accompanied by a Th17 response was indicated, which may have played a role in the decreased number of H. suis bacteria in the stomach of this age group. The decreased G-cell mass and upregulated expression of somatostatin indicated decreased acid secretion in H. suis-infected 6-8 months old pigs. In H. suis-infected adult pigs, upregulation of most markers for gastric acid secretion and increased G-cell mass was detected. Presence of severe hyperkeratosis and erosions in the non-glandular part of the stomach were mainly seen in the H. suis-positive groups. These results show that H. suis infection affects the expression of markers for acid secretion and inflammation and indicate that these effects differ depending on the infection phase.

Topics: Age Factors; Animals; Female; Gastric Acid; Gastric Mucosa; Gastritis; Helicobacter heilmannii; Helicobacter Infections; Interferon-gamma; Interleukin-17; Interleukin-1beta; Interleukin-8; Stomach; Swine; Swine Diseases

Chlamydia is one of the most common sexually transmitted diseases in humans worldwide, causing chronic lesions in the reproductive tract. Due to its often asymptomatic course, there is limited knowledge about the initial changes in the genital tract following infection. This study employs a novel sexually mature minipig model to investigate the initial histopathological changes following vaginal infection with Chlamydia trachomatis serovar D.. A vaginal inoculation resulted in an infection primarily affecting the lower genital tract. The histopathological changes were characterized by a subepithelial inflammation consisting of neutrophils and mononuclear cells, followed by an increase in the number of plasma cells within the sub-epithelial stroma of the vagina. Detection of Chlamydia was associated with expression of cyclooxygenase-2 and interleukin-8 by superficial epithelial cells. The infection was self-limiting, with a duration of 7 days.. Neutrophils, plasma cells and IL-8 have been linked with Chlamydia genital infection of unknown duration in human patients. In this study, we observe a similar pattern of local immune response/inflammation following experimental inoculation suggesting this porcine model shows promise as a model for translational chlamydia research.

Topics: Animals; Chlamydia Infections; Chlamydia trachomatis; Cyclooxygenase 2; Epithelial Cells; Female; Interleukin-8; Serogroup; Swine; Swine Diseases; Swine, Miniature; Vagina

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.

Topics: Animals; Duodenum; Female; Glucagon-Like Peptide 2; Ileum; Inflammation; Interleukin-8; Intestinal Mucosa; Intestine, Small; Lipopolysaccharides; Microspheres; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

Glässer's disease in pigs caused by Haemophilus parasuis is characterized by a severe membrane inflammation. In our previous study, we have identified activation of the transcription factor NF-κB after H. parasuis infection of porcine epithelial cells. In this study, we found that H. parasuis infection also contributed to the activation of p38/JNK MAPK pathway predominantly linked to inflammation, but not the ERK MAPK pathway associated with growth, differentiation and development. Inhibition of NF-κB, p38 and JNK but not ERK activity significantly reduced IL-8 and CCL4 expression by H. parasuis. We also found TLR1, TLR2, TLR4 and TLR6 were required for NF-κB, p38 and JNK MAPK activation. Furthermore, MyD88 and TRIF signaling cascades were essential for H. parasuis-induced NF-κB activation. These results provided new insights into the molecular pathways underlying the inflammatory response induced by H. parasuis.

Topics: Animals; Cell Line; Chemokine CCL4; Haemophilus Infections; Haemophilus parasuis; Interleukin-8; MAP Kinase Kinase 4; MAP Kinase Signaling System; NF-kappa B; p38 Mitogen-Activated Protein Kinases; Swine; Swine Diseases; Toll-Like Receptors

Porcine circovirus type 2 (PCV2) is the essential infectious agent responsible for causing porcine circovirus-associated diseases in pigs. To date, eleven RNAs and five viral proteins of PCV2 have been detected. Here, we identified a novel viral gene within the PCV2 genome, termed ORF5, that exists at both the transcriptional and translational level during productive infection of PCV2 in porcine alveolar macrophages 3D4/2 (PAMs). Northern blot analysis was used to demonstrate that the ORF5 gene measures 180 bp in length and overlaps completely with ORF1 when read in the same direction. Site-directed mutagenesis was used to show that the ORF5 protein is not essential for PCV2 replication. To investigate the biological functions of the novel protein, we constructed a recombinant eukaryotic expression plasmid capable of expressing PCV2 ORF5. The results show that the GFP-tagged PCV2 ORF5 protein localizes to the endoplasmic reticulum (ER), is degraded via the proteasome, inhibits PAM growth and prolongs the S-phase of the cell cycle. Further studies show that the GFP-tagged PCV2 ORF5 protein induces ER stress and activates NF-κB, which was further confirmed by a significant upregulation in IL-6, IL-8 and COX-2 expression. In addition, five cellular proteins (GPNMB, CYP1A1, YWHAB, ZNF511 and SRSF3) were found to interact with ORF5 via yeast two-hybrid assay. These findings provide novel information on the identification and functional analysis of the PCV2 ORF5 protein and are likely to be of benefit in elucidating the molecular mechanisms of PCV2 pathogenicity. However, additional experiments are needed to validate the expression and function of the ORF5 protein during PCV2 infection in vitro before any definitive conclusion can be drawn.

Topics: Amino Acid Sequence; Animals; Blotting, Northern; Blotting, Western; Cell Cycle; Cell Proliferation; Cells, Cultured; Circoviridae Infections; Circovirus; Fluorescent Antibody Technique, Indirect; Green Fluorescent Proteins; Immunoenzyme Techniques; Interleukin-6; Interleukin-8; Macrophages, Alveolar; Molecular Sequence Data; NF-kappa B; Real-Time Polymerase Chain Reaction; Recombinant Fusion Proteins; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sequence Homology, Amino Acid; Swine; Swine Diseases; Two-Hybrid System Techniques; Viral Proteins

To identify a fast, economic and reliable method for preselecting lactic acid-producing bacterial (LAB) isolates to control enterotoxigenic Escherichia coli (ETEC).. Two assays with porcine intestinal epithelial IPEC-J2 cells or Caenorhabditis elegans for selecting effective probiotic candidates were compared. Both assays were based on measuring death of cells or worms caused by ETEC strain JG280. Six of 13 LAB isolates showed ≥50% protection in each assay, among which only four isolates (≥50% protection) were consistently selected by both assays. Isolate CL9 (Lactobacillus reuteri) was further studied. It reduced gene expression of estA, estB and elt in JG280 in both assays. Furthermore, the isolate protected IPEC-J2 and C. elegans from cell and worm death caused by STa, STb or LT enterotoxin expressed in E. coli DH5α. CL9 also promoted host defensive responses by decreasing IL-8 and increasing IL-10 production in IPEC-J2 cells and expression of antimicrobial peptide genes clec-60, clec-85 in C. elegans.. Caenorhabditis elegans is useful for preselecting probiotic candidates to control ETEC after initial screening with IPEC-J2 cells.. A combination of IPEC-J2 cell and C. elegans assays can improve the effectiveness in preselecting probiotic candidates.

Topics: Animals; Antibiosis; Antimicrobial Cationic Peptides; Caenorhabditis elegans; Cell Line; Enterotoxigenic Escherichia coli; Enterotoxins; Epithelial Cells; Escherichia coli Infections; Interleukin-10; Interleukin-8; Intestines; Limosilactobacillus reuteri; Models, Biological; Probiotics; Swine; Swine Diseases

Influenza A virus causes a highly contagious respiratory disease in a variety of avian and mammalian hosts, including humans and pigs. The primary means for preventing influenza epidemics is vaccination. Epitope-based vaccine represents a new approach to achieve protective immunity. The objective of this study was to construct and evaluate the immunogenicity of an epitope-based antigen for its potential application in future influenza vaccine development. The antigen, comprised of a set of consensus influenza A virus epitopes (IAVe), was genetically linked to a subunit of the bacterial heat-labile enterotoxin (LTB) as an adjuvant. Immunogenicity of this LTB-IAVe antigen was evaluated in a pig model. Despite an inability to detect neutralizing antibodies directed toward the whole virus, humoral immunity against the IAVe was demonstrated in both serum (IgA and IgG) and mucosal secretions (IgG) of immunized pigs. Specific cellular immunity was also induced after LTB-IAVe immunization, as evidenced by up-regulating of IL-1β, IL-8, and IL-4 expression in peripheral blood mononuclear cells (PBMCs) of vaccinated pigs. In comparison to the non-immunized pigs, pigs immunized with the LTB-IAVe showed improved protection against a pathogenic H1N1 swine influenza virus challenge, with about 50% decrease of pneumonic lesions and 10-fold reduction of the viral load in lung and nasal secretion at five days post challenge. This study establishes a platform for future construction of epitope-based vaccines against influenza A virus infection.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Neutralizing; Bacterial Toxins; Enterotoxins; Epitopes; Escherichia coli Proteins; Humans; Immunization; Immunogenetic Phenomena; Immunoglobulin A; Influenza A Virus, H1N1 Subtype; Influenza Vaccines; Interleukin-4; Interleukin-8; Leukocytes, Mononuclear; Orthomyxoviridae Infections; Swine; Swine Diseases

Haemophilus parasuis is a colonizer of healthy piglets and the etiological agent of Glässer's disease. Differences in virulence among strains of H. parasuis have been widely observed. In order to explore the host-pathogen interaction, snatch-farrowed colostrum-deprived piglets were intranasally infected with 4 strains of H. parasuis: reference virulent strain Nagasaki, reference nonvirulent strain SW114, field strain IT29205 (from a systemic lesion and virulent in a previous challenge), and field strain F9 (from the nasal cavity of a healthy piglet). At different times after infection, two animals of each group were euthanized and alveolar macrophages were analyzed for the expression of CD163, CD172a, SLA I (swine histocompatibility leukocyte antigen I), SLA II, sialoadhesin (or CD169), and CD14. At 1 day postinfection (dpi), virulent strains induced reduced expression of CD163, SLA II, and CD172a on the surfaces of the macrophages, while nonvirulent strains induced increased expression of CD163, both compared to noninfected controls. At 2 dpi, the pattern switched into a strong expression of CD172a, CD163, and sialoadhesin by the virulent strains, which was followed by a steep increase in interleukin 8 (IL-8) and soluble CD163 in serum at 3 to 4 dpi. The early increase in surface expression of CD163 induced by nonvirulent strains went along with higher levels of IL-8 in serum than those induced by virulent strains in the first 2 days of infection. Alpha interferon (IFN-α) induction was observed only in animals infected with nonvirulent strains. Overall, these results are compatible with a delay in macrophage activation by virulent strains, which may be critical for disease production.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Cell Shape; CHO Cells; Cricetinae; Disease Models, Animal; Haemophilus Infections; Haemophilus parasuis; Histocompatibility Antigens Class I; Histocompatibility Antigens Class II; Host-Pathogen Interactions; Interferon-alpha; Interleukin-8; Macrophage Activation; Macrophages, Alveolar; Phenotype; Receptors, Cell Surface; Receptors, Immunologic; Sialic Acid Binding Ig-like Lectin 1; Swine; Swine Diseases; Virulence

Preterm birth and formula feeding predispose to necrotizing enterocolitis (NEC) in infants. As mother's milk is often absent following preterm delivery, infant formula (IF) and human donor milk (HM) are frequently used as alternatives. We have previously shown that porcine and bovine colostrum (BC) provide similar NEC protection in preterm piglets relative to IF. We hypothesized that HM exerts similar effects and that this effect is partly species-independent. Preterm piglets (n = 40) received 2 days of total parenteral nutrition, followed by a rapid transition to full enteral feeding (15 ml·kg(-1)·2 h(-1)) for 2 days using BC (n = 13), HM (n = 13), or IF (n = 14). Intestinal passage time and hexose absorption were tested in vivo. Body and organ weights were recorded on day 5, and macroscopic NEC lesions in the gastrointestinal tract were assessed. Intestinal samples were collected for determination of histomorphology, histopathology, tissue IL-6 and IL-8, organic acids, bacterial adherence by fluorescence in situ hybridization score, and digestive enzyme activities. Relative to IF, pigs from BC and HM showed longer intestinal passage time; higher weight gain, hexose absorptive capacity, mucosal proportion, and enzyme activities; lower NEC incidence, organic acid concentration, and IL-8 concentration; and reduced histopathology lesions. Tissue IL-6 concentration and bacterial adherence score were lower for HM, relative to both BC and IF groups. We conclude that BC and HM are both superior to IF in stimulating gut structure, function, and NEC resistance in preterm piglets. BC may be a relevant alternative to HM when mother's milk is unavailable during the first week after preterm birth.

Topics: Animals; Animals, Newborn; Cattle; Colostrum; Disease Models, Animal; Enteral Nutrition; Enterocolitis, Necrotizing; Female; Humans; Incidence; Interleukin-6; Interleukin-8; Intestinal Mucosa; Intestines; Milk Banks; Milk, Human; Parenteral Nutrition, Total; Pregnancy; Pregnancy, Animal; Premature Birth; Swine; Swine Diseases

The triple reassortant H2N3 virus isolated from diseased pigs in the United States in 2006 is pathogenic for certain mammals without prior adaptation and transmits among swine and ferrets. Adaptation, in the H2 hemagglutinin derived from an avian virus, includes the ability to bind to the mammalian receptor, a significant prerequisite for infection of mammals, in particular humans, which poses a big concern for public health. Here we investigated the pathogenic potential of swine H2N3 in Cynomolgus macaques, a surrogate model for human influenza infection. In contrast to human H2N2 virus, which served as a control and largely caused mild pneumonia similar to seasonal influenza A viruses, the swine H2N3 virus was more pathogenic causing severe pneumonia in nonhuman primates. Both viruses replicated in the entire respiratory tract, but only swine H2N3 could be isolated from lung tissue on day 6 post infection. All animals cleared the infection whereas swine H2N3 infected macaques still presented with pathologic changes indicative of chronic pneumonia at day 14 post infection. Swine H2N3 virus was also detected to significantly higher titers in nasal and oral swabs indicating the potential for animal-to-animal transmission. Plasma levels of IL-6, IL-8, MCP-1 and IFNγ were significantly increased in swine H2N3 compared to human H2N2 infected animals supporting the previously published notion of increased IL-6 levels being a potential marker for severe influenza infections. In conclusion, the swine H2N3 virus represents a threat to humans with the potential for causing a larger outbreak in a non-immune or partially immune population. Furthermore, surveillance efforts in farmed pig populations need to become an integral part of any epidemic and pandemic influenza preparedness.

Topics: Animals; Chemokine CCL2; Female; Humans; Influenza A virus; Influenza A Virus, H2N2 Subtype; Interferon-gamma; Interleukin-6; Interleukin-8; Lung; Macaca fascicularis; Male; Orthomyxoviridae Infections; Pneumonia, Viral; Reassortant Viruses; Severity of Illness Index; Swine; Swine Diseases

NeuB, a sialic acid synthase catalyzes the last committed step of the de novo biosynthetic pathway of sialic acid, a major element of bacterial surface structure. Here we report a functional NeuB homologue of Streptococcus suis, a zoonotic agent, and systematically address its molecular and immunological role in bacterial virulence. Disruption of neuB led to thinner capsules and more susceptibility to pH, and cps2B inactivation resulted in complete absence of capsular polysaccharides. These two mutants both exhibited increased adhesion and invasion to Hep-2 cells and improved sensibility to phagocytosis. Not only do they retain the capability of inducing the release of host pro-inflammatory cytokines, but also result in the faster secretion of IL-8. Easier cleaning up of the mutant strains in whole blood is consistent with virulence attenuation seen with experimental infections of both mice and SPF-piglets. Therefore we concluded that altered architecture of S. suis surface attenuates its virulence.

Topics: Amino Acid Sequence; Animals; Bacterial Adhesion; Bacterial Capsules; Bacterial Proteins; Cell Line, Tumor; Host-Pathogen Interactions; Hydrogen-Ion Concentration; Interleukin-8; Mice; Models, Molecular; Molecular Sequence Data; Mutation; Oxo-Acid-Lyases; Phagocytosis; Sialic Acids; Streptococcal Infections; Streptococcus suis; Swine; Swine Diseases; Virulence

Since rotavirus is one of the leading pathogens that cause severe gastroenteritis and represents a serious threat to human and animal health, researchers have been searching for cheap, safe, and effective anti-rotaviral drugs. There is a widespread of interest in using natural products as antiviral agents, and among them, licorice derived from Glycyrrhiza spp. has exerted antiviral properties against several viruses. In this study, anti-rotaviral efficacy of Glycyrrhiza uralensis extract (GUE) as an effective and cheaper remedy without side-effects was evaluated in colostrums-deprived piglets after induction of rotavirus diarrhea.. Colostrums-deprived piglets were inoculated with porcine rotavirus K85 (G5P[7]) strain. On the onset of diarrhea, piglets were treated with different concentration of GUE. To evaluate the antiviral efficacy of GUE, fecal consistency score, fecal virus shedding and histological changes of the small intestine, mRNA expression levels of inflammation-related cytokines (IL8, IL10, IFN-β, IFN-γ and TNF-α), signaling molecules (p38 and JNK), and transcription factor (NFκB) in the small intestine and spleen were determined.. Among the dosages (100-400 mg/ml) administrated to animals, 400 mg/ml of GUE cured diarrhea, and markedly improved small intestinal lesion score and fecal virus shedding. mRNA expression levels of inflammation-related cytokines (IL8, IL10, IFN-β, IFN-γ and TNF-α), signaling molecules (p38 and JNK), and transcription factor (NFκB) in the small intestine and spleen were markedly increased in animals with RVA-induced diarrhea, but dose- dependently decreased in GUE treated animals after RVA-induced diarrhea.. GUE cures rotaviral enteritis by coordinating antiviral and anti-inflammatory effects. Therapy of this herbal medicine can be a viable medication for curing rotaviral enteritis in animals and humans.

Topics: Animals; Animals, Newborn; Anti-Inflammatory Agents, Non-Steroidal; Antiviral Agents; Cell Line; Colostrum; Diarrhea; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Feces; Glycyrrhiza uralensis; Interleukin-8; Intestine, Small; MAP Kinase Signaling System; Models, Animal; NF-kappa B; Phytotherapy; Plant Extracts; Plant Roots; RNA, Messenger; Rotavirus; Rotavirus Infections; Spleen; Swine; Swine Diseases; Tumor Necrosis Factor-alpha; Virus Shedding

Four efficiently translocating Escherichia coli (TEC) strains isolated from the blood of humans (HMLN-1), pigs (PC-1) and rats (KIC-1 and KIC-2) were tested for their ability to adhere and translocate across human gut epithelial Caco-2 and HT-29 cells, to elicit a proinflammatory response and for the presence of 47 pathogenic E. coli virulence genes. HMLN-1 and PC-1 were more efficient in adhesion and translocation than rat strains, had identical biochemical phenotype (BPT) and serotype (O77:H18) and phylogenetic group (D). KIC-2 adhered more than KIC-1, belonged to different BPT and serotype but the same phylogenetic group as KIC-1. TEC strains elicited significantly higher IL-8 response in both cell lines (P < 0.05) and monocytic THP-1 (P < 0.0001) cells than non-TEC strains. KIC-2 induced the highest IL-8 response which may be associated with its immunostimulatory flagellin. Apart from adhesin genes fimH and bmaE that were carried by all strains, HMLN-1 and PC-1 carried capsule synthesis gene kpsMT III and KIC-2 carried the EAST1 toxin gene. The lack of known virulence genes and the ability of TEC to efficiently adhere and translocate whilst causing proinflammatory response suggests that these strains may carry as yet unidentified genes that enable their translocating ability.

Topics: Animals; Bacterial Adhesion; Bacterial Translocation; Caco-2 Cells; Escherichia coli; Escherichia coli Infections; HT29 Cells; Humans; Interleukin-8; Molecular Sequence Data; Phylogeny; Rats; Rodent Diseases; Swine; Swine Diseases; Virulence; Virulence Factors

Actinobacillus pleuropneumoniae (A. pleuropneumoniae) causes fibrino-hemorrhagic necrotizing pleuropneumonia in pigs. Production of proinflammatory mediators in the lungs is an important feature of A. pleuropneumoniae infection. However, bacterial components other than lipopolysaccharide involved in this process remain unidentified. The goals of this study were to determine the role of A. pleuropneumoniae exotoxin ApxI in cytokine induction and to delineate the underlying mechanisms. Using real-time quantitative PCR analysis, we found native ApxI stimulated porcine alveolar macrophages (PAMs) to transcribe mRNAs of IL-1β, IL-8 and TNF-α in a concentration- and time-dependent manner. Heat-inactivation or pre-incubation of ApxI with a neutralizing antiserum attenuated ApxI bioactivity to induce cytokine gene expression. The secretion of IL-1β, IL-8 and TNF-α protein from PAMs stimulated with ApxI was also confirmed by quantitative ELISA. In delineating the underlying signaling pathways contributing to cytokine expression, we observed mitogen-activated protein kinases (MAPKs) p38 and cJun NH2-terminal kinase (JNK) were activated upon ApxI stimulation. Administration of an inhibitor specific to p38 or JNK resulted in varying degrees of attenuation on ApxI-induced cytokine expression, suggesting the differential regulatory roles of p38 and JNK in IL-1β, IL-8 and TNF-α production. Further, pre-incubation of PAMs with a CD18-blocking antibody prior to ApxI stimulation significantly reduced the activation of p38 and JNK, and subsequent expression of IL-1β, IL-8 or TNF-α gene, indicating a pivotal role of β2 integrins in the ApxI-mediated effect. Collectively, this study demonstrated ApxI induces gene expression of IL-1β, IL-8 and TNF-α in PAMs that involves β2 integrins and downstream MAPKs.

Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Bacterial Proteins; Blotting, Western; Hemolysin Proteins; Integrin beta Chains; Interleukin-1beta; Interleukin-8; Lipopolysaccharides; Macrophages, Alveolar; Mitogen-Activated Protein Kinases; Real-Time Polymerase Chain Reaction; Signal Transduction; Sus scrofa; Swine; Swine Diseases; Trypan Blue; Tumor Necrosis Factor-alpha

Influenza virus (Flu) infection and secondary complications are a leading cause of morbidity and mortality worldwide. The increasing number of annual Flu cases, coupled with the recent Flu pandemic, has amplified concerns about the impact of Flu on human and animal health. Similar to humans, Flu is problematic in pigs, not only as a primary pathogen but as an agent in polymicrobial pneumonia. Bordetella species play a role in mixed infections and often colonize the respiratory tract without overt clinical signs. Pigs serve as a valuable animal model for several respiratory pathogens, including Bordetella (Bb) and Flu. To investigate Flu/Bb coinfection pathogenesis, a study was completed in which pigs were inoculated with Flu-only, Bb-only or both agents (Flu/Bb). Results indicate that Flu clearance is not altered by Bb infection, but Flu does enhance Bb colonization. Pulmonary lesions in the Flu/Bb group were more severe when compared to Flu-only or Bb-only groups and Bb did not cause significant lesions unless pigs were coinfected with Flu. The type I interferon response was elevated in coinfected pigs, but increased expression of antiviral genes Mx and PKR did not appear to enhance Flu clearance in coinfected pigs, as viral clearance was similar between Flu/Bb and Flu-only groups. IL-1beta and IL-8 were elevated in lungs of coinfected pigs, correlating to the days enhanced lesions were observed. Overall, Flu infection increased Bb colonization and enhanced production of proinflammatory mediators that likely contribute to exacerbated pulmonary lesions.

Topics: Animals; Bordetella bronchiseptica; Bordetella Infections; Disease Models, Animal; eIF-2 Kinase; Female; GTP-Binding Proteins; Interferon Type I; Interleukin-1beta; Interleukin-8; Lung; Myxovirus Resistance Proteins; Orthomyxoviridae; Orthomyxoviridae Infections; Swine; Swine Diseases

Leukotrienes (LT) and chemokines are important chemotactic compounds in regulating the recruitment and activation of immune cells during pulmonary inflammatory reactions. Results showed that LTC4 release by porcine alveolar epithelial type II cells (AEC IIs) is significantly enhanced by either LTB4 or LPS stimulation. The basal level of IL-8 gene expression in AEC IIs was only 1/3 of that observed in alveolar macrophages (AMs) while AEC IIs expressed a higher basal level of monocyte chemotactic peptide-1 (MCP-1) and also in response to LPS stimulation than do AMs. The increasing basal and LT-induced MCP-1 gene expressions after 8h of incubation were observed in AEC IIs but decreased in AMs. These findings suggest that AEC IIs play an important role in initial inflammatory reactions of the lung by releasing LTC4, and that they also modulate later inflammatory reactions, evidenced by consistent elevation of MCP-1 gene expression after and during exogenous challenge in pigs.

Topics: Animals; Cell Communication; Chemokine CCL2; Enzyme-Linked Immunosorbent Assay; Gene Expression; Interleukin-8; Leukotriene C4; Macrophages, Alveolar; Male; Pneumonia; Pulmonary Alveoli; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Swine; Swine Diseases

Two serovars of Salmonella enterica, namely serovar Typhimurium (ST) and serovar Choleraesuis (SC) account for the vast majority of clinical cases of swine salmonellosis worldwide. These serovars are thought to be transmitted among pigs in production settings mainly through fecal-oral routes. Yet, few studies have evaluated effects of these serovars on expression of innate immune targets when presented to pigs via repeated oral dosing in an attempt to model transmission in production settings. Thus, a primary objective of the current experiments was to evaluate expression of Toll-like receptors (TLR) and selected chemoattractive mediators (interleukin 8, IL8; macrophage migration inhibitory factor, MIF; osteopontin, OPN) in tissues from pigs exposed to ST or SC that had been transformed with kanamycin resistance and green (STG) or red (SCR) fluorescent protein to facilitate isolation from pen fecal samples. In vitro studies confirmed that STG and SCR largely (though not completely) retained their ability to upregulate IL8 and CC chemokine ligand 20 (CCL20) in cultured swine jejunal epithelial cells. Transformed bacteria were then fed to pigs in an in vivo study to determine tissue specific effects on mRNA relative expression. Pigs were fed cookie dough inoculated with bacteria on days 0, 3, 7, and 10 with 10(8)CFU STG (n=8) or SCR (n=8), while control (CTL) pigs (n=8) received dough without bacteria. Animals were sacrificed 14 days from the initial bacterial challenge and samples of tonsil, jejunum, ileum, colon, mesenteric lymph node (MLN), spleen, and liver were removed for subsequent RNA isolation. Expression of mRNA in tissues was determined using real-time quantitative PCR and expressed relative to 18S rRNA. Within CTL pigs, when expressed relative to the content in liver, mRNA for all targets demonstrated substantial tissue effects (P<0.001 for all TLR; MIF, and OPN; P<0.05 for IL8). Feeding STG and SCR resulted in significant (P

Topics: Animals; Chemokines, CC; Interleukin-8; Intestinal Diseases; Luminescent Proteins; Macrophage Migration-Inhibitory Factors; Osteopontin; Random Allocation; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Salmonella arizonae; Salmonella Infections, Animal; Salmonella typhimurium; Swine; Swine Diseases; Toll-Like Receptors; Transformation, Genetic

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative intracellular bacterium which can infect and colonize pigs. After contact with enterocytes and macrophages, S. Typhimurium induces production of cytokines thus triggering the innate immune response. In this study we evaluated the cytokine response of two porcine cell lines, IPI-2I and 3D4/31, of epithelial or macrophage origins, respectively, to the wild-type S. Typhimurium and its hilA and ssrA mutants. We observed that the 3D4/31 cell line essentially did not respond to S. Typhimurium infection when a medium with foetal calf serum was used. However when the 3D4 cell line was incubated overnight in the presence of porcine serum, it efficiently responded to the wild-type strain and the ssrA mutant but not to the noninvasive hilA mutant as measured by mRNA quantification of TNF-alpha, IL-8 and GM-CSF by the real-time RT-PCR. In IPI-2I, all the cytokines were also induced by the wild-type S. Typhimurium and the ssrA mutant although the induction of TNF-alpha was lower than that induced by the wild-type strain. The hilA mutant was unable to induce any of the cytokines tested. The ssrA mutant can therefore be considered as more suitable for further vaccine development as the stimulation of innate immune response is important for animal protection against a challenge with virulent strains.

Topics: Animals; Bacterial Adhesion; Bacterial Proteins; Cells, Cultured; Cytokines; Epithelial Cells; Granulocyte-Macrophage Colony-Stimulating Factor; Interleukin-8; Macrophages; Mutation; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Salmonella enterica; Salmonella Infections, Animal; Swine; Swine Diseases; Trans-Activators; Tumor Necrosis Factor-alpha; Virulence

Salmonella enterica serovar Choleraesuis is an enteric pathogen of swine, producing septicemia, enterocolitis, pneumonia, and hepatitis. The initial molecular events at the site of Salmonella infection are hypothesized to be critical in the initiation of innate and adaptive immune responses; however, the acute immune response elicited by porcine intestinal tissues is not well understood. To address this need, we employed explants of jejunal Peyer's patch (JPP) mucosa from pigs to examine Salmonella-induced immune responses under controlled conditions as well as to overcome limitations of whole animal approaches. JPP explants mounted in Ussing chambers maintained normal histological structure for 2 h and stable short-circuit current and electrical conductance for 2.5 h. After ex vivo luminal exposure to Salmonella serovar Choleraesuis, JPP responded with an increase in mRNA expression of IL-1beta and IL-8, but not TNFalpha. Increased IL-1beta and IL-8 expression were dependent on efficient Salmonella adhesion and internalization, whereas mutant Salmonella did not induce inflammatory cytokine expression. Commensal enteric bacteria, present in some experiments, also did not induce inflammatory cytokine expression. These findings indicate that Salmonella uptake by Peyer's patch is important in the induction of an innate response involving expression of IL-1beta and IL-8, and that ex vivo intestinal immune tissue explants provide an intact tissue model that will facilitate investigation of mucosal immunity in swine.

Topics: Animals; Female; Histocytochemistry; In Vitro Techniques; Interleukin-1; Interleukin-8; Intestinal Mucosa; Jejunal Diseases; Male; Patch-Clamp Techniques; Peyer's Patches; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Salmonella enterica; Salmonella Infections, Animal; Swine; Swine Diseases; Tumor Necrosis Factor-alpha

Intestinal epithelial cells represent the first line of defence against pathogenic bacteria in the lumen of the gut. Besides acting as a physical barrier, epithelial cells orchestrate the immune response through the production of several innate immune mediator molecules including beta-defensins. Here, we establish the porcine intestinal cell line IPI-2I as a new model system to test the regulation of porcine beta-defensins 1 and 2. Gene expression of both defensins was highly upregulated by foetal calf serum components in normal growth medium. In serum-free medium, baseline expression remained low, but pBD-2 gene expression was increased 10-fold upon infection with Salmonella Typhimurium. Arcobacter cryaerophilus and Salmonella Enteritidis, pathogenic bacteria with comparable adhesion and invasion characteristics, failed to increase pBD-2 mRNA levels. Heat killed or colistin-treated Salmonella Typhimurium had no effect, showing that the upregulation of pBD-2 was dependent on the viability of the Salmonella Typhimurium. Gene expression of pBD-1 was regulated differently since an increase in pBD-1 mRNA was observed by Salmonella Enteritidis infection. We conclude that the IPI-2I cells can serve as a new model to study porcine beta-defensin regulation and that pBD-1 and pBD-2 are differentially regulated in this cell line.

Topics: Animals; Bacterial Adhesion; beta-Defensins; Cell Line; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; Gene Expression Regulation; Interleukin-8; Intestinal Diseases; Microscopy, Electron, Scanning; Reverse Transcriptase Polymerase Chain Reaction; RNA; Salmonella Infections, Animal; Salmonella typhimurium; Statistics, Nonparametric; Swine; Swine Diseases

Monocyte chemoattractant protein-1 (MCP-1) but not interleukin-8 (IL-8) was detected by in situ hybridization using a nonradioactive digoxigenin-labeled complementary DNA probe in granulomatous lesions of lymph nodes from 20 pigs with naturally occurring postweaning multisystemic wasting syndrome (PMWS). Complementary DNA probes of 375 and 266 base pairs for MCP-1 and IL-8, respectively, were generated by reverse transcription-polymerase chain reaction. The 20 pigs with PMWS had distinct positive hybridization signals for MCP-1 but not for IL-8. The hybridization signals for MCP-1 were strictly confined to the cells with granulomatous lesions, including macrophages and multinucleated giant cells. A very close cell-to-cell correlation between MCP-1 and porcine circovirus 2 was seen in serial sections of lymph nodes. Results of this study indicate that MCP-1 expression may play a role in the pathogenesis of granulomatous inflammation in pigs with PMWS.

Topics: Animals; Chemokine CCL2; Circoviridae Infections; Circovirus; Granuloma; In Situ Hybridization; Interleukin-8; Lymph Nodes; Reverse Transcriptase Polymerase Chain Reaction; RNA, Viral; Swine; Swine Diseases; Wasting Syndrome

Streptococcus suis capsular type 2 is an important aetiologic agent of swine meningitis, and it has been highlighted as a cause of occupational disease leading to meningitis and fulminant sepsis in humans. The objective of the present work was to study the ability of S. suis type 2 to induce the release of tumour necrosis factor alpha (TNF-alpha), interleukin-1 (IL-1), IL-6, IL-8 and monocyte chemotactic protein one (MCP-1) by human monocytic THP-1 cells. The induction of these five cytokines was dose- and incubation time-dependent, and it was significantly enhanced by pre-treatment of cells with interferon gamma. IL-8 levels were markedly higher compared with those obtained with the other cytokines. However, elevated levels of MCP-1 and IL-6 were also observed. Levels of cytokine induced by heat-killed or live bacteria were similar. Pre-treatment of cells with anti-CD14 monoclonal antibodies suggested that this important host receptor is partially implicated in TNF, IL-1, IL-6 and MCP-1 production, while CD14-independent pathways seem to be responsible for IL-8 production after S. suis stimulation. In addition, blocking studies with anti-TNF and anti-IL-1 antibodies revealed that these cytokines are involved in amplification of the S. suis-induced cytokine cascade. When several different S. suis strains of human or porcine origin were compared, a very heterogeneous pattern of cytokine production was observed. Human strains did not exhibit a clear tendency to induce higher cytokine release by human THP-1 monocytes. The synergistic effect of the up-regulation of cytokines during S. suis meningitis may mediate many of the inflammatory reactions, including the sequestration of leucocytes at the site of infection.

Topics: Animals; Antibodies, Monoclonal; Chemokine CCL2; Hot Temperature; Humans; Interferon-gamma; Interleukin-1; Interleukin-6; Interleukin-8; Interleukins; Lipopolysaccharide Receptors; Monocytes; Neoplasm Proteins; Recombinant Proteins; Species Specificity; Streptococcal Infections; Streptococcus suis; Swine; Swine Diseases; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha; Virulence

Pentoxifylline, a methylxanthine derivative and nonspecific type 4 phosphodiesterase inhibitor, has been used to improve survival of animals with sepsis and to attenuate lung injury in acute lung inflammation. The purpose of this study was to examine whether pentoxifylline would inhibit the expression of inflammatory cytokines, particularly tumor necrosis factor alpha (TNF), and thereby decrease the pathophysiology of acute porcine pleuropneumonia. E. coli lipopolysaccharide (LPS) and bacterial extracts of A. pleuropneumoniae--induced elevations in TNF mRNA which were fully abrogated by addition of pentoxifylline in both alveolar macrophage and neutrophil cultures. A 30% reduction in the level of LPS-induced interleukin (IL)-1beta mRNA levels also was achieved in macrophages. Pentoxifylline did not affect either IL-1alpha or IL-8 expression in vitro. Pentoxifylline therapy in vivo significantly reduced the number of band neutrophils in swine but did not reduce the pathology associated with pleuropneumonia, including changes in serum zinc, iron, or haptoglobin. Neither did it alter TNF, IL-1, IL-6, or IL-8 expression. Measurement of pentoxifylline and its metabolites in pig sera suggested that efficacious doses of pentoxifylline were probably not achieved in vivo. However, subcutaneous doses of pentoxifylline higher than 25 mg/kg produced transient diarrhea, vomiting, and tremors. These results suggest that pentoxifylline is an effective pharmacological tool for the dissection of cytokine regulation in vitro, but inhibitory concentrations may not be achievable for in vivo pharmacological use in swine.

Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Animals; Bronchoalveolar Lavage Fluid; Cells, Cultured; Diarrhea; Dose-Response Relationship, Drug; In Vitro Techniques; Interleukin-1; Interleukin-6; Interleukin-8; Macrophages, Alveolar; Neutrophils; Pentoxifylline; Phosphodiesterase Inhibitors; Swine; Swine Diseases; Tremor; Tumor Necrosis Factor-alpha; Vomiting

Inflammatory mediators that are induced by gram-negative bacteria in the course of intrauterine infections threaten successful pregnancy. To compare the effect of two different routes of cytokine induction, bacterial lipopolysaccharide (LPS) was administered in vivo either into the cord vein or into the amniotic cavity of pig fetuses in the second half of gestation for 20 h and cytokines were detected in the amnion.Tumor necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8) were induced in the amniotic epithelium after intra-amniotic but not after intra-venous administration of LPS. The presence of IL-8 was confirmed by RT-PCR. In contrast, transforming growth factor-beta1 (TGF-beta1) was expressed constitutively and was found in all samples of the amniotic epithelium. Amniotic fluid contained only minute levels of TNF-alpha. IL-8 levels in amniotic fluid increased after the treatment with LPS and the highest IL-8 levels were found in dead LPS-treated fetuses.

Topics: Amnion; Animals; Chorioamnionitis; Female; Fetus; Immunohistochemistry; Interleukin-8; Lipopolysaccharides; Pregnancy; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Swine; Swine Diseases; Swine, Miniature; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

Swine neutrophils were quantitatively examined for the direct and indirect migratory responses to Actinobacillus pleuropneumoniae (APP) in vitro and the effects of pseudorabies virus (PrV), frequently co-infecting with APP, were also observed. About 30% of swine neutrophils responded to viable APP, while 3.2% of the neutrophils responded to 0.1% casein which served as the control. The migration of APP was not affected by preincubation of neutrophils with PrV, which inhibited the random migration. When the random migration was normalized to 1, the chemotactic indices for APP, opsonized-APP and casein were 64, 70 and 8.5, respectively. Heat-killed APP or E. coli lipopolysaccharide stimulated the production of interleukin-8 activity by adherent peripheral blood mononuclear cells (PBMC). Preincubation of PBMC with PrV inhibited the production of neutrophil attractant activity when stimulated with heat-killed APP. The results suggested that the direct chemotaxis of neutrophils to viable APP might contribute to early infiltration in Actinobacillus pleuropneumonia, and that PrV might inhibit indirect recruitment of neutrophils to infected lungs by compromising the functions of PBMC.

Topics: Actinobacillus pleuropneumoniae; Animals; Caseins; Cell Adhesion; Chemotaxis, Leukocyte; Culture Media, Conditioned; Herpesvirus 1, Suid; Interleukin-8; Kinetics; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Swine; Swine Diseases

To develop and evaluate an in vivo model to study early events in the pathogenesis of acute porcine pleuropneumonia.. Thirty-six 6- to 8-week-old pigs.. Pigs were inoculated intranasally or endotracheally with Actinobacillus pleuropneumoniae; inoculation routes were compared by evaluation of clinical signs, gross and microscopic lung lesions, hematologic changes, serum zinc, iron, and haptoglobin concentrations, and inflammatory cytokines.. The 2 inoculation routes resulted in similar findings, although intranasal inoculation caused unilateral gross lung lesions, whereas endotracheal inoculation caused bilateral gross lesions. Clinical signs of disease were observed < 2 hours after endotracheal inoculation and 6 to 8 hours after intranasal inoculation. Total WBC counts did not differ significantly after inoculation by either inoculation route, although band neutrophils increased significantly. The earliest findings associated with A pleuropneumoniae inoculation, irrespective of route, were decreased serum zinc and iron concentrations. Serum haptoglobin concentrations were significantly increased after inoculation. Inoculation induced rapid influx of macrophages into the lung and local induction of proinflammatory cytokines. Northern blot analysis of total RNA from lung tissue indicated that inoculated pigs had increased concentrations of interleukin (IL)-1beta, IL-1alpha, and IL-8; tumor necrosis factor messenger RNA concentration was not increased.. Endotracheal inoculation with A pleuropneumoniae rapidly and consistently induced diffuse bilateral pneumonia; thus, this method may be useful for the study of acute pathophysiologic changes associated with bacterial pneumonia and may provide an experimental model for testing modalities for prevention and treatment of this and other respiratory tract diseases of pigs.

Topics: Actinobacillus Infections; Actinobacillus pleuropneumoniae; Acute Disease; Administration, Intranasal; Animals; Antibodies, Monoclonal; Blotting, Northern; Cytokines; Disease Models, Animal; DNA Probes; Haptoglobins; Immunohistochemistry; Interleukin-1; Interleukin-8; Intubation, Intratracheal; Iron; Lung; Pleuropneumonia; RNA, Bacterial; Swine; Swine Diseases; Tumor Necrosis Factor-alpha; Zinc

To evaluate in situ expression of inflammatory cytokine mRNA in lymphoid tissue of swine experimentally infected with Mycobacterium avium serovar 2.. 7 noninfected pigs and 7 pigs infected with M. avium serovar 2.. Expression of mRNA of inflammatory cytokines such as tumor necrosis factor alpha (TNFalpha), interleukin (IL)-1beta IL-6, and IL-8 in formalin-fixed paraffin-embedded blocks of lymphoid tissue (lymph nodes and tonsil) of swine experimentally infected with M. avium serovar 2 was compared with that of noninfected pigs. Tissues were evaluated by use of morphologic localization of cytokine mRNA, using in situ hybridization at 160 days after inoculation.. A noticeable increase in mRNA expression for TNFalpha and mild increases in mRNA expression of IL-8 and IL-1beta were detected in mandibular lymph nodes from infected swine, compared with noninfected swine. Mild increase in mRNA expression for 1L-6 also was observed in tonsils from infected swine. Cytokine mRNA was detected in macrophages and lymphocytes, primarily within cortical follicles and adjacent mantle zones.. Expression of mRNA for inflammatory cytokines was increased in lymphoid tissue of infected swine, possibly resulting from local factors on, or secreted by, M. avium. These results suggest that alterations in cytokine mRNA expression are important in the pathogenesis and clinical course of mycobacteriosis in swine. Modulation of the immune response by vaccines that selectively target cytokine expression and secretion in response to mycobacterial challenge may be effective in prevention of mycobacteriosis in swine.

Topics: Animals; Cytokines; Gene Expression Regulation; Interleukin-1; Interleukin-6; Interleukin-8; Lymph Nodes; Lymphoid Tissue; Mycobacterium avium; Palatine Tonsil; Reference Values; RNA, Messenger; Swine; Swine Diseases; Transcription, Genetic; Tuberculosis; Tumor Necrosis Factor-alpha