Effects of thiol antioxidants on hepatocyte growth factor signaling in cardiac myocytes

We describe here novel antioxidant-sensitive events in which activation kinetics are delayed, leading to inhibition of cell signaling. Hepatocyte growth factor (HGF) transiently phosphorylated p44/42 mitogen-activated protein kinase (MAPK) with a peak at 3-5 min in HL-1 adult cardiac myocytes. Pretreatment of cells with thiol antioxidants, N-acetylcysteine or a-lipoic acid attenuated MAPK phosphorylation induced by a 3-min incubation with HGF. However, kinetic analysis revealed that the apparent inhibition of HGF signaling was due to a delay in the activation because HGF phosphorylated MAPK with a peak at 5-7 min in cells treated with thiol antioxidants. This 2-min delay in HGF activation of MAPK resulted in >5-min delay in phosphorylation of MAPK targets such as p90RSK and GATA-4. Hydrogen peroxide did not mimic HGF signaling, and HGF did not induce reactive oxygen species production. Thus, in cardiac myocytes, thiol antioxidants delay HGF-mediated MAPK activation and suppress subsequent signaling events via reactive oxygen species-independent mechanism.