Contribution of lysosomal vesicles to the formation of lipid raft redox signaling platforms in endothelial cells. Antioxid Redox Signal 9

We have demonstrated that the formation of lipid raft (LR)-redox signaling platforms membrane is associ-ated with activation of acid sphingomyelinase (ASMase) in coronary arterial endothelial cells (CAECs). Given that the trafficking of lysosomal vesicles might play an essential role in ASMase activation, the present study tested whether lysosomal vesicles contribute to the formation of LR redox signaling platforms. By confocal microscopy, we found that Fas ligand (FasL) induced the formation of LR clusters in the plasma membrane of CAECs, accompanied by aggregation of NAD(P)H oxidase subunits, gp91phox and p47phox, and ROS pro-duction. When the cells were pretreated with two structurally different lysosomal vesicle function inhibitors, bafilomycin A1 (Baf) and glycyl-L-phenylalanine--naphthylamide (GPN), the FasL-induced LRs clustering was substantially blocked, and corresponding ROS production significantly decreased. By confocal micro-scopic observations in living CAECs by using LysoTracker, a colocalization of LRs and lysosomal vesicles was found around the cell membrane, which was abolished by Baf or GPN. Functionally, FasL-induced inhibition of endothelium-dependent vasorelaxation was also reduced by both inhibitors of lysosome function. These re-sults suggest that lysosomal vesicles importantly contribute to the formation of LR-redox signaling platforms and thereby participate in the oxidative injury of endothelial function during activation of death receptor-Fa