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

View Article: PubMed Central - PubMed

ABSTRACT

Deoxynivalenol (DON), produced by the plant pathogens Fusariumgraminearum 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.

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Effect of U0126 (10 µM) on reassembly of a tight IPEC-J2 monolayer after calcium deprivation of differentiated IPEC-J2. IPEC-J2, differentiated in 1.12 cm2 non-coated Transwell® polyester membrane inserts, were: (A) either pretreated U0126 (10 µM) or cultivation medium before calcium deprivation; or (B) exposed to calcium deprivation without pre-treatment. TEER was measured during recovery (0, 2, 4, 6 and 24 h) in complete calcium-containing cultivation medium with or without U0126 (10 µM). Test and control wells were tested in triplicate. Data represent mean ± SD, n = 4.
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toxins-08-00264-f004: Effect of U0126 (10 µM) on reassembly of a tight IPEC-J2 monolayer after calcium deprivation of differentiated IPEC-J2. IPEC-J2, differentiated in 1.12 cm2 non-coated Transwell® polyester membrane inserts, were: (A) either pretreated U0126 (10 µM) or cultivation medium before calcium deprivation; or (B) exposed to calcium deprivation without pre-treatment. TEER was measured during recovery (0, 2, 4, 6 and 24 h) in complete calcium-containing cultivation medium with or without U0126 (10 µM). Test and control wells were tested in triplicate. Data represent mean ± SD, n = 4.

Mentions: To further investigate the positive effect of U0126 pretreatment on TEER values of untreated IPEC-J2, a calcium switch assay was performed to determine reassembly of a tight IPEC-J2 monolayer after its deliberate destruction (Figure 4). U0126-supplemented cultivation medium, compared to cultivation medium alone, induced a more rapid increase of TEER in the 24 h following calcium deprivation. Even without a 24 h preincubation step with U0126, preceding the calcium switch, U0126 (10 µM) accelerated monolayer formation after calcium deprivation, even if less effectively.


Early Activation of MAPK p44/42 Is Partially Involved in DON-Induced Disruption of the Intestinal Barrier Function and Tight Junction Network
Effect of U0126 (10 µM) on reassembly of a tight IPEC-J2 monolayer after calcium deprivation of differentiated IPEC-J2. IPEC-J2, differentiated in 1.12 cm2 non-coated Transwell® polyester membrane inserts, were: (A) either pretreated U0126 (10 µM) or cultivation medium before calcium deprivation; or (B) exposed to calcium deprivation without pre-treatment. TEER was measured during recovery (0, 2, 4, 6 and 24 h) in complete calcium-containing cultivation medium with or without U0126 (10 µM). Test and control wells were tested in triplicate. Data represent mean ± SD, n = 4.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5037490&req=5

toxins-08-00264-f004: Effect of U0126 (10 µM) on reassembly of a tight IPEC-J2 monolayer after calcium deprivation of differentiated IPEC-J2. IPEC-J2, differentiated in 1.12 cm2 non-coated Transwell® polyester membrane inserts, were: (A) either pretreated U0126 (10 µM) or cultivation medium before calcium deprivation; or (B) exposed to calcium deprivation without pre-treatment. TEER was measured during recovery (0, 2, 4, 6 and 24 h) in complete calcium-containing cultivation medium with or without U0126 (10 µM). Test and control wells were tested in triplicate. Data represent mean ± SD, n = 4.
Mentions: To further investigate the positive effect of U0126 pretreatment on TEER values of untreated IPEC-J2, a calcium switch assay was performed to determine reassembly of a tight IPEC-J2 monolayer after its deliberate destruction (Figure 4). U0126-supplemented cultivation medium, compared to cultivation medium alone, induced a more rapid increase of TEER in the 24 h following calcium deprivation. Even without a 24 h preincubation step with U0126, preceding the calcium switch, U0126 (10 µM) accelerated monolayer formation after calcium deprivation, even if less effectively.

View Article: PubMed Central - PubMed

ABSTRACT

Deoxynivalenol (DON), produced by the plant pathogens Fusariumgraminearum 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.

No MeSH data available.


Related in: MedlinePlus