XII - 12th International Symposium - Digestive Physiology of Pigs

Digestive
Physiology
of Pigs
function. In this study, we treated human colon carcinoma
cells Caco-2/15 and porcine intestinal epithelial cells IPEC
J2 with bovine serocolostrum, milk whey, lactoferrin or
macropeptide, and verified their effect on cell proliferation,
migration, adhesion and inflammatory response. Caco-2/15
and IPEC J2 cells were treated with different concentrations
of milk fractions. Cell proliferation was determined using the
XTT assay, while migration was assessed by wound assay
and adhesion was observed following cell trypsinization.
Transient transfection and luciferase assay were done
to verify the impact of milk fractions on cell inflammatory
response using NF-kBluc, IL-8luc and IL-6luc as reporter
genes. After the transfection, Caco-2/15 and IPEC J2
cells were stimulated with heat-killed Escherichia coli and
Salmonella typhimurium (HK bacteria), in combination with
different concentrations of milk fractions. Cell proliferation
was increased by 20% in Caco-2/15 cells and up to 50%
in IPEC J2 cells after 24h treatment with serocolostrum (10
mg/mL), lactoserum (10 mg/mL) and lactoferrin (1 mg/mL).
Cell migration was increased (P < 0.05) by serocolostrum
in IPEC J2, while cell adhesion to the culture dish was
significantly impaired by serocolostrum and lactoserum. In
Caco-2/15 cells, lactoferrin decreased (P < 0.05) the cellcell
adhesion, while macropeptide increased cell-matrix
adhesion. Transient transfections and luciferase assay
showed a decrease (P < 0.05) in the induction of NF-kBluc,
IL-8luc and IL-6luc transcriptional activity by HK bacteria
in presence of serocolostrum. In conclusion, bovine milk
fractions modulate differently functional properties of
human and porcine intestinal epithelial cells by affecting
cell proliferation, migration and adhesion in vitro, while
inflammatory response is exclusively affected by bovine
serocolostrum.
Key words: milk fractions, intestinal epithelial cells
2019 Serocolostrum modulates gene expression in
porcine intestinal epithelial cells IPeC J2. M. Blais 1 , Y.
Pouliot 2 , S. Gauthier 2 , Y. Boutin 2 , and M. Lessard* 1 , 1 Dairy
and Swine Research and Development Centre, Sherbrooke,
Qc, Canada, 2 Institute of Nutraceuticals and Functional
Foods, U. Laval, Quebec, Qc, Canada.
Bovine serocolostrum contains nutrients and many bioactive
molecules including growth factors, immunoglobulins and
antimicrobial peptides. Previous studies have suggested the
therapeutical use of serocolostrum to treat gastrointestinal
disorders, but little is known about its specific impact on
intestinal epithelium. The aim of this study is to verify effect
of serocolostrum on gene expression, with or without
stimulation of the inflammatory response, in porcine
intestinal epithelial cells IPEC J2 by microarray analysis.
The IPEC J2 cells were treated for 2 h or 24 h with
serocolostrum (10mg/ml), in combination with heat-killed
Escherichia coli and Salmonella typhimurium (HK bacteria),
followed by RNA extraction. Gene expression was analyzed
by microarray using Agilent porcine array. The statistical
analysis of microarray data was done using the FlexArray
1.6.1 software package (Génome Québec, Canada) and
list of genes that were significantly regulated by either HK
bacteria or serocolostrum were classified according to their
Gene Ontology (GO) Biological Processes. Serocolostrum
XII INTERNATIONAL SYMPOSIUM ON
DIGESTIVE PHYSIOLOGY OF PIGS
99
Session III
treatment significantly increased the expression of
genes involved in cell proliferation, morphogenesis,
development, locomotion and response to wounding. More
than 50% of genes induced more than 2 times after HK
bacteria stimulation were downregulated by addition of
serocolostrum. These genes are mostly involved in immune
response, defense response and inflammatory response,
according to GO classifications. On the other hand,
approximately 25% of genes induced more than 2 times after
HK bacteria stimulation had increased levels of expression
when serocolostrum was added, and these genes are
rather involved in development. In conclusion, these results
show that serocolostrum could have positive properties on
intestinal epithelial integrity by stimulating basal levels of
genes involved in proliferation, repair and development in
intestinal epithelial cells in vitro. Also, serocolostrum acts
as an anti-inflammatory agent by decreasing the level of
induction of immune and inflammatory genes in presence
of pathogens.
Key words: microarray, porcine intestinal epithelial cells,
serocolostrum
2020 Influence of a phytogenic feed additive on digestive,
microbiological and immunological measurements
in weaned piglets. J. Zentek* 1 , S. Gaertner 1 , L.
Scharek-Tedin 1 , A. Mader 1 , K. R. Wendler 2 , and W. Vahjen 1 ,
1 Institut of Animal Nutrition, Freie Universitaet Berlin, Berlin,
Germany, 2 DELACON Biotechnik, Steyregg, Austria.
To identify possible anti-enteroadhesive effects against an
enteropathogenic Escherichia coli strain, flow cytometric
measurements were performed with IPEC-J2 cells, using
labeled E. coli (CFDA-SE). A feeding trial with 24 male
castrated pigs was conducted using a complete feed without
(control) or with 0.04% of a phytogenic feed additive (Fresta
F, Delacon, Steyregg, Austria). The piglets were orally
vaccinated with a live Salmonella Typhimurium vaccine to
determine the specific antibodies. On d 28 and 29 of the trial,
peripheral blood and digesta were sampled. Performance,
intestinal pH, dry matter content of digesta and the
intestinal microbiota (qPCR, Bifidobacteria, Lactobacillus
amylovorus, L. reuteri, L. johnsonii; Escherichia/Hafnia/
Shigella spp.; clostridial clusters I and XIVa; streptococci)
and bacterial metabolites (lactate, short chain fatty acids
(SCFA)) were determined. Leukocytes from the blood and
the lamina epithelialis of the proximal jejunum were isolated
and phenotyped by flow cytometry. The phagocytic activity
of isolated peripheral monocytes and granulocytes was
measured (Phagotest©, Orpegen, Heidelberg, Germany).
Proliferation of peripheral lymphocytes was analyzed with
3 mitogens (PWM, CON A, PHA-M). The adhesion of the
E. coli to IPEC-J2 cells was reduced by the plant extract
by 52.7% (P < 0.05). In piglets fed the diet with the additive
ileal pH (P < 0.05) and digesta dry matter was not affected.
Numbers of Escherichia spp. decreased in the jejunum
and clostridial cluster I increased (P < 0.05). Numbers of
clostridial cluster XIV decreased in the cecum. Lactate
and SCFA concentrations were similar in both groups. The
immunological measurements were unaffected, including
the concentrations of antibodies against S. Typhimurium,
the lymphocyte proliferation and the phagocytic activity