Maximakinin reversed H 2 O 2 induced oxidative damage in rat cardiac H9c2 cells through AMPK/Akt and AMPK/ERK1/2 signaling pathways

Maximakinin (MK), a homolog of bradykinin (BK), is extracted from skin venom of the Chinese toad Bombina maxima . Although MK has a good antihypertensive effect, its effect on myocardial cells is unclear. This study investigates the protective effect of MK on hydrogen peroxide (H 2 O 2 )-induced oxidative damage in rat cardiac H9c2 cells and explores its mechanism of action. A 3-(4,5-Dimethyl-2-Thiazolyl)-2,5-Diphenyl Tetrazolium Bromide (MTT) assay was selected to detect the effect of MK on H9c2 cell viability, while flow cytometry was used to investigate the influence of MK and H 2 O 2 on intracellular reactive oxygen species (ROS) levels. Protein expression changes were detected by western blot. In addition, specific protein inhibitors were applied to confirm the induction of ROS-related signaling pathways by MK. MTT assay results show that MK significantly reversed H 2 O 2 -induced cell growth inhibition. Flow cytometry Dichlorodihydrofluorescein diacetate (DCFH-DA) staining shows that MK significantly reversed H 2 O 2 -induced increases in intracellular ROS production in H9c2 cells. Moreover, the addition of specific protein inhibitors suggests that MK reverses H 2 O 2 -induced oxidative damage by activating AMP -activated protein kinase (AMPK)/protein kinase B (Akt) and AMPK/extracellular-regulated kinase 1/2 (ERK1/2) pathways. Finally, an inhibitor of bradykinin B 2 receptors (B2Rs), HOE -140, was applied to investigate potential targets of MK in H9c2 cells. HOE -140 significantly blocked induction of AMPK/Akt and AMPK/ERK1/2 pathways by MK, suggesting a potentially important role for B2Rs in MK reversing H 2 O 2 -induced oxidative damage. Above all, MK protects against oxidative damage by inhibiting H 2 O 2 -induced ROS production in H9c2 cells. The protective mechanism of MK may be achieved by activation of B2Rs to activate downstream AMPK/Akt and AMPK/ERK1/2 pathways.