AbstractIntestinal oxidative stress produces pro-inflammatory cytokines, which increase tight junction (TJ) permeability, leading to intestinal and systemic inflammation. Cystine (Cys2) is a substrate of glutathione (GSH) and inhibits inflammation, however, it is unclear whether Cys2 locally improves intestinal barrier dysfunction. Thus, we investigated the local effects of Cys2 on oxidative stress-induced TJ permeability and intestinal inflammatory responses. Caco-2 cells were cultured in a Cys2-supplemented medium for 24 h and then treated with H2O2 for 2 h. We assessed TJ permeability by measuring transepithelial electrical resistance and the paracellular flux of fluorescein isothiocyanate–dextran 4 kDa. We measured the concentration of Cys2 and GSH after Cys2 pretreatment. The mRNA expression of pro-inflammatory cytokines was assessed. In addition, the levels of TJ proteins were assessed by measuring the expression of TJ proteins in the whole cells and the ratio of TJ proteins in the detergent-insoluble fractions to soluble fractions (IS/S ratio). Cys2 treatment reduced H2O2-induced TJ permeability. Cys2 did not change the expression of TJ proteins in the whole cells, however, suppressed the IS/S ratio of claudin-4. Intercellular levels of Cys2 and GSH significantly increased in cells treated with Cys2. Cys2 treatment suppressed the mRNA expression of pro-inflammatory cytokines, and the mRNA levels were significantly correlated with TJ permeability. In conclusion, Cys2 treatment locally reduced oxidative stress-induced intestinal barrier dysfunction possively due to the mitigation of claudin-4 dislocalization. Furthermore, the effect of Cys2 on the improvement of intestinal barrier function is related to the local suppression of oxidative stress-induced pro-inflammatory responses.
Inflammatory cytokines and intestinal barrier dysfunction in patients with severe acute pancreatitis