Apoptotic stress pathway activation mediated by iron on endothelial cells in vitro

Background. Iron sucrose (Fe-S) and low-molecular-weight iron dextran (Fe-D) have been used successfully in the treatment of anaemia in chronic kidney disease patients. However, some side effects, such as endothelial cell dysfunction have been reported. Mechanisms by which iron can induce endothelial cell damage have not been completely understood. This study was designed to examine the effect of Fe-S and Fe-D on bovine aortic endothelial cells in vitro. Methods. Cell proliferation was determined by [H-3] thymidine incorporation, cytotoxicity by lactate dehydrogenase, pro-Caspase-3 by immunoblotting; and Caspase-3 activity using a colorimetric assay. Expression of the apoptosis stress pathway proteins Bcl-2 and Bax and cycle arrest proteins p53 and p21(WAF/CIP1) were examined by immunoblot. Cell apoptosis was tested by terminal deoxynucleotidyltransferase-mediated nick-end labelling (TUNEL) and DNA fragmentation. Results. Both iron preparations inhibited cell proliferation. This effect was more important and occurred at lower concentrations in Fe-S than Fe-D cultured cells. Expression of p53 and p21(WAF/CIP1) increased in cells incubated with Fe-S, but not with Fe-D. Bcl-2 expression was significantly down-regulated in cells incubated with Fe-S in comparison with Fe-D, while Bax expression was not modified by the iron compounds. Pro-Caspase-3 expression and Caspase-3 activity increased only in cells treated with Fe-S. Apoptosis was present in cells treated with Fe-S. Conclusions. Our results demonstrate that Fe-S exerts a greater inhibitory effect on endothelial cell proliferation than Fe-D. The mechanisms involved in this process may be related, at least in part, to over expression of proteins related to the cell cycle arrest and apoptosis stress pathway.