Protection of Cultured Cortical Neurons by Luteolin against Oxidative Damage through Inhibition of Apoptosis and Induction of Heme Oxygenase-1

Luteolin, one of the most common flavonoids present in many types of natural products, possesses diverse biological properties including anti-oxidant activity. In this study, we investigated neuroprotective effect of luteolin and its underlying signaling pathways using primary cultured rat cortical cells. Luteolin was demonstrated to attenuate H2O2- or xanthine/xanthine oxidase-induced oxidative damage and generation of intracellular reactive oxygen species (ROS). It enhanced the phosphorylation of Bad at Ser112 and attenuated H2O2-induced activation of caspase 3, indicating anti-apoptotic action. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling (TUNEL) assay confirmed this finding, showing inhibition of H2O2-induced DNA fragmentation. We also found that luteolin significantly up-regulated the expression of anti-oxidant enzyme heme oxygenase (HO)-1. Treatment with tin protoporphyrin IX, a selective HO-1 inhibitor, abolished neuroprotective and anti-apoptotic effects of luteolin, suggesting a critical role of HO-1 up-regulation. It was also shown to increase the phosphorylation of mitogen-activated protein kinase (MAPKs) such as extracellular signal-regulated kinase (ERK), p38 MAPK and c-Jun N-terminal kinases (JNK) and Akt. Treatment of the cells with specific inhibitors including SB203580, SP600125, and LY294002 suppressed the luteolin-induced HO-1 expression, suggesting the involvement of p38 MAPK, JNK, and Akt in HO-1 induction. In contrast, HO-1 expression was not reduced by U0126, implying that ERK may not be directly involved in HO-1 induction. These results indicate that luteolin exhibits neuroprotective effect through the inhibition of ROS and apoptotic cell death. Furthermore, up-regulation of HO-1 expression via p38 MAPK, JNK and Akt may contribute, at least in part, to luteolin-mediated neuroprotection. Based on these findings, luteolin may serve as a potential intervention for neurodegenerative diseases associated with oxidative stress.