Preliminary Characterization of the Transcriptional Response of the Porcine Intestinal Cell Line IPEC-J2 to Enterotoxigenic Escherichia coli, Escherichia coli, and E. coli Lipopolysaccharide

IPEC-J2, a promising in vitro model system, is not well characterized especially on the transcriptional level, in contrast to human counterparts. The aim of this study was to characterize the gene expression in IPEC-J2 cells when coincubated with enterotoxigenic Escherichia coli (ETEC), nonpathogenic E. coli, and E. coli endotoxin. Apical infection of polarized IPEC-J2 monolayers caused a time-dependent decrease in transepithelial electrical resistance (TEER). Microarray analysis showed up-regulation of interleukins when IPEC-J2 were cocultured with E. coli strains this has so far never been measured in this cell line. Highest IL8 expression was found with the ETEC strain possessing the F4 fimbrium, suggesting IPEC-J2 cells to be F4 receptor positive, confirmed in a brush border membrane adhesion assay. It is concluded that the innate immune responses to pathogens and LPS makes the IPEC-J2 cell line a suitable model for research on intestinal host pathogen interaction

Non-invasive intestinal biomarkers: a new ELISA test for Pancreatitis Associated Protein detection in pig

Feed additives are commonly used to improve pig performance and health, but they need to be tested so new biomarkers for intestinal health, non- or minimally invasive, are under investigations.The quantification of Myeloperoxidase (MPO) and Pancreatitis Associated Protein (PAP) in feces could prove useful to non-invasively monitor intestinal health (Niewold, 2015). MPO is an enzyme that permits to quantify the number of inflammatory cells present in tissues and feces (Prokopowicz et al., 2012) , while PAP is a protein mainly produced in the small intestine with anti-inflammatory and bactericidal activity (Cash et al., 2006; Mukherjee et al., 2014). Because of the lack of a commercial ELISA kit for porcine PAP detection, the main aim of this study was to develop and validate a new sandwich ELISA test for the quantification of PAP in pig fecal samples. Our study consisted of two phases: test development and test validation. During the development phase we used polyclonal antibodies previously immunized from rabbit serum with a pure peptide containing the N-terminus of pig PAP (Soler et al., 2015). The validation of the test was then performed with fecal extraction samples derived from animals with known high or low growth performance.Moreover, the temperature stability of PAP in feces and the optimal extraction method was tested. Even if only preliminary, our results seem to show a fair relationship between fecal consistency, used as health indicator, and PAP fecal concentrations. Furthermore, no relevant differences in PAP concentration after 24h of incubation at 37 °C, 4°C or room temperature were detected.To date, the present results suggest that PAP seems to be exceptionally stable in feces and is a very promising candidate as a non-invasive (fecal) biomarker for intestinal health and growth

The acute phase protein, haptoglobin : a potential parameter in welfare assessment?

Physiological parameters are important measures in animal welfare assessment. To assess the amount of stress an animal experiences, stress hormones like cortisol are frequently used. However, measuring cortisol has major disadvantages due to its rapid reactivity and decline and many influencing factors. Other potential alternative markers are acute phase proteins, since stress is known to affect the immune system. A pilot study was conducted to investigate the response of the acute phase protein, plasma haptoglobine (HP), in pigs subjected to a stressor (food deprivation) and to examine the correlation between HP levels and average daily growth (ADG). Forty grower pigs (25.1 ± 4.4 kg, mean ± SD) (sex and former pen mates balanced), were allocated to 4 conventional pens, 2 treatment (T) and 2 control (C) groups (10 pigs per pen). After 10 days of adaptation the experiment started and ran for 3 weeks. In the 2nd week, T groups were repeatedly subjected to an 8-hour food deprivation (day 1, 3, 5 and 7 of week 2), C groups had normal, unrestricted, access to food. Pigs were weighed twice a week and blood was collected once a week (every 5th day). Mean levels of plasma HP of C and T groups showed large variation between individuals (C groups, week 2: 1.84 ± 3.11 mg/ml; T groups, week 2: 1.40 ± 1.16 mg/ml). No significant differences (Kruskal-Wallis test) in HP levels or growth were found between the C and T groups or between the different weeks within the T groups. Significant negative weak to moderate correlations were found between ADG and HP levels (HP week 1 and ADG week 1: rs = -0.47, p=0.005; HP week 2 and ADG total; rs= -0.60, p=0.015; HP week 3 and ADG total: rs = -0.43, p=0.025; average HP total and ADG total: rs= -0.41, p=0.017). Large variations in HP levels between individuals were shown and no effect of treatment on HP levels or growth was found. Possibly, food deprivation had no apparent stress eliciting effect. Despite these results, interesting correlations between the level of HP and ADG were found, corroborating the inverse relationship between the acute phase response and growth. To further investigate the relation of the acute phase response and stress a successive experiment will be conducted in which we apply a stronger stressor (mixing pigs) and combine the physiological data with behavior

Importance of heat-stable enterotoxin B in the induction of early immune responses in piglets after infection with enterotoxigenic Escherichia coli

Enterotoxigenic Escherichia coli (ETEC) strains that produce heat-stable (STa, STb) and/or heat-labile (LT) enterotoxins are an important cause of post-weaning diarrhea in piglets [1]. However, the relative importance of the different enterotoxins in the pathogenesis of ETEC infection has been poorly defined. In the present study we assessed the contributions of different ETEC enterotoxins to the induction of small intestinal secretion and early innate immune responses in weaned piglets

Role of heat-labile and heat-stable enterotoxins in the induction of early immune responses in piglets after infection with enterotoxigenic Escherichia coli

C

Importance of heat-stable enterotoxin B in the induction of early immune responses in piglets after infection with enterotoxigenic Escherichia coli

Enterotoxigenic Escherichia coli (ETEC) strains that produce heat-stable (STa, STb) and/or heat-labile (LT) enterotoxins are an important cause of post-weaning diarrhea in piglets [1]. However, the relative importance of the different enterotoxins in the pathogenesis of ETEC infection has been poorly defined. In the present study we assessed the contributions of different ETEC enterotoxins to the induction of small intestinal secretion and early innate immune responses in weaned piglets

Занятие-общение - коммуникативно-ориентированная форма обучения научному стилю речи

The aim of the study was to determine the intraarticular set-Urn amyloid A (SAA) response pattern in horses with inflammatory arthritis. Inflammatory arthritis was induced by injection of lipopolysaccharide (LPS) into the radiocarpal joint of four horses. Serum and synovial fluid (SF) samples were collected before and at 4, 8, 12, 24, 48, 72, 96, and 144 h after injection. Concentrations of SAA were measured by immunoturbidometry, and expression of SAA isoforms was visualized by denaturing isoelectric focusing and Western blotting. The LPS injection caused systemic and local clinical signs of inflammation. Serum amyloid A appeared in serum and SF within 8 h after LPS injection. Isoelectric focusing showed three major SAA hands with apparent isoelectric points (pI) of 7.9, 8.6, and > 9.3 in serum and SF. Synovial fluid contained two additional isoforms with highly alkaline apparent pI values (apparent pl value extrapolated from standard curve = 10.0 and 10.2), which were not present in any of the serum samples. In conclusion, intraarticular injection of LPS induced systemic and local inflammatory responses in the horses. By demonstrating SF-specific SAA isoforms the results of the present Study suggest that SAA is synthesized locally in the equine inflamed joint, similar to what has been demonstrated in humans previously. The marked local SAA synthesis suggests an important pathophysiological role in inflammatory arthritis

Early transcriptional response in the jejunum of germ-free piglets after oral infection with virulent rotavirus

Germ-free piglets were orally infected with virulent rotavirus to collect jejunal mucosal scrapings at 12 and 18 hours post infection (two piglets per time point). IFN-gamma mRNA expression was stimulated in the mucosa of all four infected piglets, indicating that they all responded to the rotavirus infection. RNA pools prepared from two infected piglets were used to compare whole mucosal gene expression at 12 and 18 hpi to expression in uninfected germ-free piglets (n = 3) using a porcine intestinal cDNA microarray. Microarray analysis identified 13 down-regulated and 17 up-regulated genes. Northern blot analysis of a selected group of genes confirmed the data of the microarray. Genes were functionally clustered in interferon-regulated genes, proliferation/differentiation genes, apoptosis genes, cytoskeleton genes, signal transduction genes, and enterocyte digestive, absorptive, and transport genes. Down-regulation of the transport gene cluster reflected in part the loss of rotavirus-infected enterocytes from the villous tips. Data mining suggested that several genes were regulated in lower- or mid-villus immature enterocytes and goblet cells, probably to support repair of the damaged epithelial cell layer at the villous tips. Furthermore, up-regulation was observed for IFN-γ induced guanylate binding protein 2, a protein that effectively inhibited VSV and EMCV replication in vitro (Arch Virol 150:1213–1220, 2005). This protein may play a role in the small intestine’s innate defense against enteric viruses like rotavirus

Intestinal health biomarkers in vivo

© Wageningen Academic Publishers 2015. There is a definite need for biomarkers for intestinal health in vivo, which can be determined in samples obtained in a non-invasive or minimally invasive way. In humans, there are approximately fifteen biomarkers suggested, and test and reagents are available, whereas this is hardly the case for pigs and chicken. Here we review (possible) biomarkers for intestinal health, and their presence or absence in pig and chicken. There appears to be a striking lack of information on intestinal biomarkers in both species. It is also clear that certain biomarkers may not be present in all species, which is particularly true for chicken, or if present, are immunologically different. This usually means that reagents for assays are not available. For the pig there are at least some biomarkers and assays available such as for intestinal fatty acid binding protein (I-FABP), but essentially none are available for chicken. Given the importance of intestinal health in production animals, it is hoped that more effort is invested in this field.status: publishe