Abstract
ObjectiveTo investigate the protective effects and mechanisms of carbon monoxide-releasing molecule-2 (CORM-2) on barrier function of intestinal epithelial cells.Materials and MethodsAfter pre-incubation with CORM-2 for 1 hour, cultured intestinal epithelial IEC-6 cells were stimulated with 50 µg/ml lipopolysaccharides (LPS). Cytokines levels in culture medium were detected using ELISA kits. Trans-epithelial electrical resistance (TER) of IEC-6 cell monolayers in Transwells were measured with a Millipore electric resistance system (ERS-2; Millipore) and calculated as Ω/cm2 at different time points after LPS treatment. The permeability changes were also measured using FITC-dextran. The levels of tight junction (TJ) proteins (occludin and ZO-1) and myosin light chain (MLC) phosphorylation were detected using Western blotting with specific antibodies. The subsequent structural changes of TJ were visualized using transmission electron microscopy (TEM).ResultsCORM-2 significantly reduced LPS-induced secretion of TNF-α and IL-1β. The LPS-induced decrease of TER and increase of permeability to FITC-dextran were inhibited by CORM-2 in a concentration dependent manner (P<0.05). LPS-induced reduction of tight junction proteins and increase of MLC phosphorylation were also attenuated. In LPS-treated cells, TEM showed diminished electron-dense material and interruption of TJ and desmosomes between the apical lateral margins of adjoining cells, which were prevented by CORM-2 treatment.ConclusionsThe present study demonstrates that CORM-2, as a novel CO-releasing molecule, has ability to protect the barrier function of LPS-stimulated intestinal epithelial cells. Inhibition of inflammatory cytokines release, restoration of TJ proteins and suppression of MLC phosphorylation are among the protective effects of CORM-2.
Highlights
Carbon monoxide (CO) was considered as a toxic gas for a long time due to its higher affinity for hemoglobin than oxygen
The present study demonstrates that carbon monoxide-releasing molecule-2 (CORM-2), as a novel CO-releasing molecule, has ability to protect the barrier function of LPS-stimulated intestinal epithelial cells
Cytotoxic effects of CORM-2 on IEC-6 cells in vitro Previous studies have shown that CORM-2 is not toxic to airway epithelial cells, but it has not been applied to intestinal IEC-6 cells
Summary
Carbon monoxide (CO) was considered as a toxic gas for a long time due to its higher affinity for hemoglobin than oxygen. Some researchers have found that CO has additional functions in biological systems than was first thought. An elevated level of CO was detected in exhaled air of critically ill patients as a result of HO-1 induction, which was considered to be a self-protective mechanism of an organism [3]. It has been reported that induction of HO-1 or exogenous CO administration could improve the survival of septic mice [4,5]. Other biological functions of CO include anti-inflammation, antithrombosis and vasodilatation [7,8,9,10]. Increasing attention has been focused on this diatomic gas as a potential messenger
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