Purified recombinant mouse netrin-1 (R&D Systems, Minneapolis, MN, USA) was administered at doses ranging from 1 to 50 μg/kg, dissolved in normal saline, via injection into the LV lumen at the onset of reperfusion. Some mice were subjected to tail vein injection with netrin-1 (5 μg/kg) also immediately at the onset of reperfusion. UO126, a MEK1/2 inhibitor (200 μg/kg, dissolved in DMSO<0.01%), and PTIO (1 mg/kg, dissolved in normal saline), a specific NO chelator, were purchased from Sigma-Aldrich in highest purity (St. Louis, MO, USA), and administered via injection into the LV lumen at the onset of reperfusion.
Polyclonal antibodies specific for phosphorylated and total forms of ERK1/2, eNOS, were obtained from Cell Signaling Technology (CST, Danvers, MA, USA). NOX4 antibody was purchase from Abcam (Abcam, Cambridge, MA, USA). Anti-actin, antibody, and other chemicals in highest purity, were purchased from Sigma-Aldrich (Sigma-Aldrich, St. Louis, MO, USA).
Polyclonal antibodies specific for phosphorylated and total forms of ERK1/2, eNOS, were obtained from Cell Signaling Technology (CST, Danvers, MA, USA). NOX4 antibody was purchase from Abcam (Abcam, Cambridge, MA, USA). Anti-actin, antibody, and other chemicals in highest purity, were purchased from Sigma-Aldrich (Sigma-Aldrich, St. Louis, MO, USA).