Vascular cells under peroxide induced oxidative stress: a balance study on in vitro peroxide handling by vascular endothelial and smooth muscle cells.

Abstract

Enzymes such as glutathione peroxidase and catalase play an important role in the cellular defence against (per)oxidative stress. Balance- and inhibitor-studies were undertaken with in vitro cultured human vascular endothelial cells (EC) and smooth muscle cells (SMC) to assay the relative importance of these enzymes in the handling of cumene hydroperoxide (Chp) and hydrogen peroxide (H2O2). Low concentrations of Chp (up to 80 microM) could be removed to near completion within the first hour of incubation by stimulation of the hexose monophosphate shunt (HMS) of both cell types. The HMS activity reached a plateau upon incubation with higher concentrations of Chp (> 80 microM). The non-converted Chp in the higher concentrations could be detected quantitatively in the incubation solution. After inhibition of the glutathione reductase by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), incubation with Chp (40 microM) did not result in a stimulation of the HMS activity. Moreover the added Chp could be recovered from the medium. So Chp is exclusively handled by the GSH-redox cycle. When low concentrations of H2O2 (up to 80 microM) were added to EC or SMC approximately 50% of the peroxide loss could not be accounted for. Inhibitor studies with aminotriazole proved that catalase was responsible for the handling of this unaccounted H2O2. In both ECs and SMCs at lower concentrations of H2O2 the GSH-redox cycle was as effective as catalase and at higher H2O2 concentrations the catalase pathway plays the major role.