Preconditioning of carbon monoxide releasing molecule-derived CO attenuates LPS-induced activation of HUVEC

To investigate the effects and potential mechanisms of preconditioning of tricarbonyldichlororuthenium (III) dimer (CORM-2)-liberated CO on LPS-induced activation of endothelial cells (HUVEC).

HUVEC were pretreated with CORM-2 at the concentration of 50 or 100 microM for 2 hrs, washed and stimulated with LPS (10 microg/ml) for additional 4 hrs. Activation (oxidative stress) of HUVEC was assessed by measuring intracellular oxidation of DHR 123 or nitration of DAF-FM, specific H(2)O(2) and NO fluorochromes, respectively. The expression of HO-1, iNOS (Western blot) and ICAM-1 (cell ELISA) proteins and activation of inflammation-relevant transcription factor, NF-kappaB (EMSA) were assessed. In addition, PMN adhesion to HUVEC was also assessed.

The obtained data indicate that pretreatment of HUVEC with CORM-2 results in: 1) decrease of LPS-induced production of ROS and NO; 2) up-regulation of HO-1 but decrease in iNOS at the protein levels; 3) inhibition of LPS-induced activation of NF-kappaB; and 4) downregulation of expression of ICAM-1, and this was accompanied by a decrease of PMN adhesion to LPS-stimulated HUVEC.

Preconditioning of CO liberated by CORM-2 elicited its anti-inflammatory effects by interfering with the induction of intracellular oxidative stress. In addition, it also supports the notion that CO is a potent inhibitor of iNOS and NF-kappaB.