H2O2 CAUSES ENDOTHELIAL BARRIER DYSFUNCTION WITHOUT DISRUPTING THE ARGININE NITRIC-OXIDE PATHWAY

We have previously demonstrated that nitric oxide ((.)NO) donors atten uate and that inhibition of endogenous nitric oxide synthase (NOS) enh ances hydrogen peroxide (H2O2)-mediated porcine pulmonary artery endot helial cell (PAEC) injury. The current study investigates the hypothes is that oxidant-mediated inhibition of NOS contributes to PAEC injury. PAEC barrier function, measured as the transmonolayer clearance of al bumin, was significantly impaired by H2O2 (10-100 mu M) in the absence of cytotoxicity. Treatment with H2O2 did not alter NOS activity, meas ured as the conversion of [H-3]arginine to [H-3]citrulline in PAEC lys ates, either immediately after treatment with 0-250 mu M H2O2 for 30 m in or for up to 120 min after treatment with 100 mu M H2O2. H2O2 had l ittle effect on NOS activity in intact PAECs, measured as 1) the forma tion of [H-3]citrulline in [H-3]arginine-loaded PAECs, 2) PAEC guanosi ne 3',5'-cyclic monophosphate content, and 3) PAEC.NO release to the c ulture media. These results indicate that the arginine-.NO pathway rem ains intact after exposure to oxidant conditions sufficient to promote functional derangements of vascular endothelial cells.