Early Activation of MAPK p44/42 Is Partially Involved in DON-Induced Disruption of the Intestinal Barrier Function and Tight Junction Network

Abstract

Deoxynivalenol (DON), produced by the plant pathogens Fusarium graminearum and Fusarium culmorum, is one of the most common mycotoxins, contaminating cereal and cereal-derived products. Although worldwide contamination of food and feed poses health threats to humans and animals, pigs are particularly susceptible to this mycotoxin. DON derivatives, such as deepoxy-deoxynivalenol (DOM-1), are produced by bacterial transformation of certain intestinal bacteria, which are naturally occurring or applied as feed additives. Intestinal epithelial cells are the initial barrier against these food- and feed-borne toxins. The present study confirms DON-induced activation of MAPK p44/42 and inhibition of p44/42 by MAPK-inhibitor U0126 monoethanolate. Influence of DON and DOM-1 on transepithelial electrical resistance (TEER), viability and expression of seven tight junction proteins (TJ), as well as the potential of U0126 to counteract DON-induced effects, was assessed. While DOM-1 showed no effect, DON significantly reduced TEER of differentiated IPEC-J2 and decreased expression of claudin-1 and -3, while leaving claudin-4; ZO-1, -2, and -3 and occludin unaffected. Inhibition of p44/42 counteracted DON-induced TEER decrease and restored claudin-3, but not claudin-1 expression. Therefore, effects of DON on TEER and claudin-3 are at least partially p44/42 mediated, while effects on viability and claudin-1 are likely mediated via alternative pathways.

Effect of Fusarium-Derived Metabolites on the Barrier Integrity of Differentiated Intestinal Porcine Epithelial Cells (IPEC-J2)

“…We recently demonstrated that DON does not significantly decrease TEER of differentiated IPEC-J2 at concentrations below 5 µM [87]. EC90 values of DON were equal to 8.08 ± 3.15 µM after 24 h, 4.53 ± 0.85 µM after 48 h and 4.45 ± 1.37 µM after 72 h of incubation.…”

“…Thus, we herein present the first investigation of the effect of such metabolites on intestinal barrier integrity of intestinal porcine epithelial cells. Such effects are well documented for the trichothecene mycotoxin DON [11,15,87,88,89], as well as for aflatoxins [90,91], zearalenone [92,93], ochratoxin A [94,95], patulin [96,97], fumonisin B1 [98,99,100] and T2 toxin [16,101]. Similarities with the original in vivo cell tissue and with humans and the fact that pigs are the most susceptible species to Fusarium mycotoxin DON, make porcine cell lines a highly suitable option for in vitro analysis of intestinal barrier integrity [102].…”

“…It has recently been reported, that beauvericin (3 and 10 µM) induces expression of phosphorylated mitogen activated protein kinase (MAPK) ERK44/42 within 24 h of exposure [109]. This is a documented feature of DON, where impairments of barrier function—shown by TEER decreases and compromised expression of TJ proteins claudin-3 and 4—are mediated through p44/42 phosphorylation [13,87]. It seems plausible that a potential beauvericin-induced activation of p44/42 could at least partially be responsible for the observed reductions of TEER in beauvericin-treated IPEC-J2.…”

“…We recently reported that DON concentrations below 5 µM do not negatively affect TEER of differentiated IPEC-J2 over a duration of 72 h [87]. Therefore we combined no-effect concentrations of metabolites (if applied individually) with no-effect DON concentrations (1.5 and 3 µM).…”

“…It presents an interesting starting point for further evaluations, for instance regarding effects of these metabolites on tight junctions or MAPK activity. In preliminary trials we were able to show that the selected metabolites—just like DON [87]—do not reduce protein expression of zona occludens (ZO)-1 and occludin. However, preliminary work suggests effects on the expression of tight junction protein, claudin-1 (data not shown).…”