NO-Induced Lipofuscin Acumulation Was Caused by Inhibition of Cathepsin S

Abstract

Abstract: : Purpose:To investigate effects of Nitric oxide (NO) on the phagocytosis in RPE cells with the correlation of cathepsin S (Cat S) activity.Methods:RPE cells were isolated from 10-week-old Long evans rats. The RPE cells were cultured in the medium containing NO donor (S-nitroso-N-acetylpenicillamine,SNAP) with or without NO scavenger (Hydroxocobalamin) from 72hrs before the measurement of the phagocytic activities. Phagocytic activities of ROS were evaluated by a flow cytometric analysis and the activity of Cat S was measured by using a fluorogenic substrate. We also measured the activity of purified bovine spleen Cat S under the presence of SNAP. The Labeled or unlabeled ROS was used for monitoring the ingestion or digestion of ROS in RPE cells, respectively. Results:SNAP inhibited the phagocytic and Cat S activities of the RPE cells in a dose-dependent manner. Purified bovine spleen catS activity was also suppressed by co-incubation of SNAP in a dose dependent manner. These activities were partially recovered by the addition of NO scavenger in both RPE cells and purified Cat S. Conclusions: We demonstrated that NO donor inhibited the activity of catS and the phagosytosis of ROS. There were some reports about the direct S- nitrosylation of caspase catalytic cysteine residue by NO and Cat S is a cystein protease. The inhibitory effects of NO on phagocytosis may be due to S-nitrosylation which involves transfer of the NO group to the sulfhydryl of a cysteine residue.

Keywords: phagocytosis and killing • nitric oxide • enzymes/enzyme inhibitors