Skip to main content
. Author manuscript; available in PMC: 2015 Aug 1.
Published in final edited form as: Free Radic Biol Med. 2014 May 14;73:201–213. doi: 10.1016/j.freeradbiomed.2014.04.029

Figure 3. Cav-1 negatively regulates ROS production from NADPH oxidases in both heterologous expression systems and native cells.

Figure 3

(A–E) COS-7 cells were co-transfected with cDNAs encoding Nox1, 3, 4, 5 and either Cav-1 or a non-specific control gene (RFP), or peritoneal macrophages (source of Nox2) were transfected with either Cav-1 or RFP. Superoxide production was monitored by L-012-mediated chemiluminescence and hydrogen peroxide (H2O2) was measured using the Amplex® Red assay with excitation of 530–560 nm and emission detection at ~590 nm. (F–J) COS-7 cells or peritoneal macrophages expressing Nox1-Nox5 were treated with vehicle or the Cav-1 scaffolding domain peptide for 24 hrs and ROS production measured as described. (K) Knockdown of Cav-1 using siRNA (10, 30nM) in human lung fibroblasts increases ROS production. (L) Macrophages isolated from Cav-1−/− mice produce significantly higher amount of ROS compared to that of WT mice. (M) Cav-1 directly regulates ROS production from Nox5. COS-7 cells were co-transfected with cDNAs encoding Nox5 and either Cav-1 or a non-specific control gene (RFP). The activity of Nox5 in cell-free extracts was determined following injection of NADPH (arrow). * different from Control, WT or Vehicle, p < 0.05 (n = 5–6).