Establishment of an In Vitro DSS-Induced Epithelial Injury Model to Investigate Dose-Dependent Effects of IL-6 and...

Dextran sodium sulfate (DSS) is widely used in vivo to induce epithelial injury; however, the epithelial-specific responses to DSS and mechanisms associated with barrier restoration remain incompletely understood. Here, we establish an in vitro DSS-induced epithelial damage model using differentiated Caco-2 cell monolayers. Exposure to 2.5%(w/v) DSS for 24 h resulted in significant upregulation of the pore-forming protein Claudin-2(CLDN2) (2.677-fold) and elevated oxidative marker protein levels of NADPH oxidase 1 (NOX1), accompanied by increased reactive oxygen species (ROS) production confirmed via dihydroethidium (DHE) staining. Increased epithelial permeability was observed by transepithelial electrical resistance (TEER) analysis, without significant cytotoxicity. Surprisingly, ELISA analysis revealed significant downregulation of Interleukin-6 (IL-6) (0.364 ± 0.034 pg/mL) following DSS exposure. Treatment with 1 ng/mL recombinant IL-6 (rIL-6) improved epithelial barrier function during continued DSS exposure, whereas a higher dose of 5 ng/mL did not produce the same restorative response. Furthermore, treatment of DSS-damaged Caco-2 monolayer with 10 mM butyrate significantly improved epithelial barrier function. Collectively, this study establishes a reproducible DSS-induced Caco-2 epithelial injury model and demonstrates concentration-dependent effects of rIL-6 and barrier-restorative effects of butyrate, providing an epithelial-specific platform for investigating intestinal barrier dysfunction relevant to IBD.