Figure 4.

ROS are involved in the COM crystal-induced tight junction disruption. MDCK cells were pretreated with 10 mM NAC for 2 h and then incubated with 1 mM COM crystals for 48 h. (a) ROS production induced by COM was inhibited by NAC. Intracellular ROS were determined by DCFH-DA assay using flow cytometry. (b) The apoptosis induced by COM crystals was alleviated by NAC. MDCK cells treated with or without NAC were detected using flow cytometry by Annexin-V/PI staining. (c) NAC treatment inhibited the down-regulation of ZO-1 and occludin induced by COM crystal, and repressed the phosphorylation of Akt, ASK1, and p38. Protein levels were detected by Western blot and the relative band intensities were analyzed by Image-Pro Plus 6.0. Illustrated is a representative of three separate experiments and the quantifications of data were represented as mean ± SD on the right panel. **p < .01 versus control; ##p < .01 versus COM.