Nutritional Mitigation of Deoxynivalenol-Induced Endocytosis and Degradation of Intestinal Tight Junction Proteins

Enkai Li, Purdue University

Abstract

The presence of trichothecene mycotoxins in feed and food has emerged as a major threat to both animal and human health, as up to 25% of the cereal production across the globe could be contaminated with these mycotoxins. The intestine has been shown as the first line of defense against external contaminants and the major target organ for these mycotoxins. Deoxynivalenol (DON), the most prevalent mycotoxin detected in cereal products, has been shown to decrease protein expression of tight junction proteins (TJP) and induce intestinal barrier loss. However, the overall mechanism by which DON regulates specific intestinal TJP turnover and epithelial barrier function remains unclear. In the first study, our data showed that DON exposure decreased the protein stability and accelerated the endocytosis and degradation of TJP in the lysosome and this is partly regulated by the activation of p38 MAPK and JNK signaling pathways. We then tested the effect of a natural JNK inhibitor, resveratrol against DON-induced stress. Resveratrol prevented the endocytic degradation of claudin-4 caused by DON and protected against DONinduced decrease in transepithelial electrical resistance (TEER) and increase in fluorescein isothiocyanate-labeled dextran (FITC-dextran) permeability in IPEC-J2 cells. Intestinal tight junction structures are known to be sensitive to mitogenic stimuli that are regulated by multiple signaling pathways. Recent transcriptomic studies suggested that intestinal exposure to DON suppressed peroxisome proliferator activated receptor gamma (PPAR) signaling pathway. The role of PPAR in the regulation of adipocyte differentiation has been well characterized. Besides adipose tissue, PPAR is also highly expressed in the intestine. However, the potential role of PPAR in regulating intestinal function still remains unclear. In the second and third studies, we investigated the protective effects of both pharmacological and natural PPAR agonists against DON-induced intestinal barrier loss. PPAR activation by pharmacological PPAR ligands reversed DON-induced reduction of claudin-3 and 4 in vitro and in vivo. These protective effects were lost in the presence of a specific PPAR antagonist or in PPAR knockout cells, confirming the importance of PPAR in the regulation of intestinal barrier function and integrity. Consistent with the beneficial effects observed obtained from pharmacological PPAR ligands, both eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), two natural PPAR agonists, also blocked DON-induced endocytic degradation of claudin-4. EPA and DHA also restored the membrane presence of claudin-4 and ZO-1 that was disrupted by DON. Effect of PPAR activation by EPA and DHA also included the restoration of TEER and reduction of FITC-dextran permeability that have been perturbed by DON. However, the effectiveness of EPA and DHA in opposing DON-induced decrease in TEER and the increase in FITC-dextran permeability was not affected by PPAR inhibition, potentially suggesting the involvement of other PPAR-independent mechanisms in the observed benefits from EPA and DHA. Recent studies revealed that DON exposure suppressed nuclear factor erythroid 2-like 2 (Nrf2) signaling pathway, suggesting that Nrf2 might also play a role in the response to mycotoxins exposure. Given the demonstrated protective effect of Nrf2 activation in several conditions, we hypothesized that activation of Nrf2 might exert a protective effect against DON-induced intestinal barrier dysfunction. In the fourth study, we investigated the effects of a natural Nrf2 activator, quercetin in alleviating the DON-induced barrier loss and intestinal injuries in vitro and in vivo. Quercetin treatment dose-dependently increased mRNA expression of Nrf2 target gene, NQO-1, and concomitantly increased the expression of claudin-4 at both mRNA and protein levels. Quercetin supplementation also reversed the reduction of claudin-4 caused by DON exposure in vivo and in vitro. In vivo study in weaned pigs showed that DON exposure impaired villus-crypt morphology. Notably, intestinal injuries caused by DON administration were partly mitigated by quercetin supplementation.

Degree

Ph.D.

Advisors

Ajuwon, Purdue University.

Subject Area

Animal sciences

COinS