Optimization of a small intestinal segment perfusion model for heat-stable enterotoxin A induced secretion in pigs

Abstract

Enterotoxigenic Escherichia coli (ETEC) are a major cause of dehydrating diarrhoea in children and weaned piglets living under subhygienic conditions. After colonization of the small intestine, ETEC produce heat-labile (LT) and/or heat-stable (ST) enterotoxins. Individual human ETEC strains generally produce only one type of enterotoxin, and in symptomatic patients from developing countries the heat-stable enterotoxin A (STa) producing strains seem by far the most prevalent. STa binds to and activates guanylyl cyclase C (GC-C) in the brush border of intestinal epithelium. An increase in cGMP in turn activates cGMP-dependent protein kinase II (cGKII) leading to phosphorylation of the cystic fibrosis transmembrane regulator (CFTR) driving Cl-secretion and inhibition of NaCl absorption followed by net loss of water through osmotic diarrhoea. We used a small-intestinal segment perfusion model of ETEC diarrhea in pigs [1] in which we investigated the possibility of eliciting STa induced secretion. Five consecutive mid-jejunal segments of anaesthetized piglets were perfused for 6 hours with different concentrations of STa (150-0.62 nM) in a physiologic salt solution. Changes in intestinal net fluid absorption were measured. We concluded that the STa response was dose-dependent and that continuous perfusion with 50 nM of STa or more was required to reduce net absorption. In all piglets the control segments showed net absorption. Fifty nanomolar of STa was sufficient to reduce net absorption compared to control segments in 10 out of 12 piglets; in 8 of them net absorption was reversed to net secretion. STa-induced responses showed relative high variation between different jejunal segments similar to the inter-segment variation seen in control animals where segments were perfused with physiologic salt solution. Results indicate that more optimization is required before this model could be used to test compounds that could interfere with the STa induced fluid secretion. 1. Nabuurs, M.J., et al., A long-term perfusion test to measure net absorption in the small intestine of weaned pigs. Res Vet Sci, 1993. 55(1): p. 108-14.

Simultaneous Exposure to Escherichia coli Heat-Labile and Heat-Stable Enterotoxins Increases Fluid Secretion and Alters Cyclic Nucleotide and Cytokine Production by Intestinal Epithelial Cells

“…For instance, when Greenberg et al (42) examined adults with travelers' diarrhea, they demonstrated that fecal lactoferrin and the inflammatory cytokines interleukin-8 (IL-8), IL-1␤, and IL-1ra were elevated in individuals with ETEC travelers' diarrhea. Loos et al (43) examined the role of ST toxins in the induction of early immune responses in piglets after infection with ETEC and demonstrated that the physiological response to ETEC is also accompanied by a marked change in mucosal expression of innate immunity genes. Additionally, LT has been shown to induce the production of cytokines and chemokines from intestinal cells both in vivo and in vitro (35).…”

Synbiotic Effects of Enzyme and Probiotics on Intestinal Health and Growth of Newly Weaned Pigs Challenged With Enterotoxigenic F18+Escherichia coli

“…McLamb et al ( 24 ) previously reported that pigs challenged with ETEC have the immune response activated. According to Loos et al ( 46 ), the secretion of IL-6 in the lumen of the small intestine is stimulated by STa produced by E. coli F18 + . In this study, the ETEC challenge increased the concentration of IL-6 as previous reported ( 46 , 47 ).…”

“…According to Loos et al ( 46 ), the secretion of IL-6 in the lumen of the small intestine is stimulated by STa produced by E. coli F18 + . In this study, the ETEC challenge increased the concentration of IL-6 as previous reported ( 46 , 47 ). High levels of IL-6 reduce the secretion of growth hormone ( 48 ) and damage the intestinal epithelium ( 49 , 50 ).…”

Heat-Stable Enterotoxins of Enterotoxigenic Escherichia coli and Their Impact on Host Immunity

“…In the latter, using the small intestinal segment perfusion (SISP) technique, STb was shown to play a dominant role during the early secretory response as compared to the contribution of STa and LT [20]. In younger piglets (two weeks old), LT was reported to be a more important virulence factor as compared to STb [20,48,49].…”