FO020
ENDOTHELIAL DYSFUNCTION IN UREMIA: EFFECT OF FLAVONOIDS AND ANTIOXIDANTS Free

Introduction and Aims: Accelerated atherosclerosis in chronic kidney disease (CKD) is preceded by the development of endothelial dysfunction with the change of the endothelium exposed to uremic conditions to a proinflammatory and prothrombotic phenotype and enhanced oxidative stress. We evaluated the possible protective effect of several anti-inflammatory and antioxidant compounds in endothelial cells (EC) exposed to a uremic media in vitro.

Methods: Endothelial cells were incubated with the flavonoids apigenin, genistein and quercetin, and the chemically synthesized antioxidants Ebselen, EUK-134 and EUK-118 and subsequently exposed to culture medium supplemented with serum from CKD patients on dialysis, and to healthy donors, as the control group. The optimal concentrations were determined by cell viability studies (MTT). Changes in the expression of the adhesion receptor ICAM-1 and in the production of reactive oxygen species (ROS) were screened by immunofluorescence and fluorescence techniques. Activation of inflammation-related proteins, NFkappaB (NFκB), p38 MAP kinase (p38MAPK) was evaluated by ELISA and Western blot.

Results: Exposure of human umbilical endotelial cells (HUVECs) to uremic medium induced a significant increased expression of ICAM1 in the cell surface, activation of NFκB and p38MAPK, and ROS generation compared to the control group (fold increase of 1.9±0.2,1.9±0.1, 1.8±0.1 2.2±0.1,vs control, respectively p<0.05).Pretreatment with Ebselen, and EUK 134, EUK118 resulted in the restoration of normal levels in ICAM-1 expression and in a significant decrease of ROS production (56%, 25% and 30%,vs uremic group, respectively, p<0.01), whereas in relation with flavonoids, only Quercetin showed a moderate but significant inhibitory effect on both parameters (decrease of 37% for ICAM-1 and 33% for ROS production, vs uremic group, p<0.05). Both flavonoids and antioxidants reduced p38MAPK activation (Decreases of p38MAPK phosphorilation compared with the uremic condition were: Apigenin and Ebselen about 43% p<0.01; and Quercetin, Genistein, EUK 134 and EUK118 about 26%, p<0.05), while only antioxidant-enzyme mimetics were able to inhibit the activation of NFκB induced by uremia. (decreases of 31% for Ebselen, 22% for EUK134 and 17% for EUK118, vs uremic condition, p<0.05).

Conclusions: These results indicate that chemically synthesized antioxidant enzyme-mimetics exhibit not only a potent antioxidant effect, but also an anti-inflammatory effect in uremia-induced endothelial dysfunction, while only one of the flavonoids (quercetin) showed an anti-inflammatory effect under the studied conditions. This study demonstrates the efficacy in vitro of Ebselen, EUK118 and EUK134 molecules in preventing the development of endothelial dysfunction in uremia.