title: Hepatocyte-protective effect of nectandrin B, a nutmeg lignan, against oxidative stress: Role of Nrf2 activation through ERK phosphorylation and AMPK-dependent inhibition of GSK-3β.
abstract: Oxidative stress can contribute to the development and progression of liver diseases, such as drug-induced or alcoholic liver injury, nonalcoholic fatty liver disease, and nonalcoholic steatohepatitis. Nectandrin B is a bioactive lignan isolated from nutmeg extract. To date, little information is available about its pharmacological activities in the liver. This study investigated the hepatocyte-protective effect of nectandrin B against tert-butylhydroperoxide-induced oxidative injury and the underlying molecular mechanism. The cell viability assay revealed that nectandrin B prevents apoptosis stimulated by tert-butylhydroperoxide in both HepG2 cells and primary mouse hepatocytes. Nectandrin B also attenuated ROS production and restored the depleted glutathione level. Real-time PCR and immunoblot analyses showed that the expression of glutamate-cysteine ligase, an enzyme responsible for the glutathione biosynthesis, was induced by nectandrin B, indicating its indirect antioxidative effect. The NF-E2-related factor-2 (Nrf2) regulates gene expression of an array of antioxidant enzymes in hepatocytes. Nectandrin B stimulated Nrf2 activation as evidenced by its enhanced nuclear accumulation and increased antioxidant response element (ARE)-luciferase activity. Intriguingly, the hepatocyte-protective effect of nectandrin B against oxidative damage was completely abrogated by Nrf2 knockdown using Nrf2 specific siRNA. Nectandrin B promoted ERK activation, but inactivated GSK-3β through the AMPK-mediated inhibitory phosphorylation. The enforced overexpression of dominant-negative mutant of MEK1 or AMPKα, or wild-type GSK-3β inhibited the increase in the NQO1-ARE-luciferase activity stimulated by nectandrin B, suggesting that both ERK and AMPK-GSK-3β signalings are involved in the activation of Nrf2/ARE pathway by nectandrin B. Consistent with this, cytoprotection and restoration of glutathione level by nectandrin B was also blocked by the overexpression of dominant-negative MEK1 or wild-type GSK-3β. Finally, our data demonstrate that nectandrin B has the ability to protect hepatocytes against oxidative injury through the activation of Nrf2/ARE pathway mediated by ERK phosphorylation and AMPK-dependent inactivation of GSK-3β.
authors: Department of Pharmacy, College of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan, Gyeonggi-do 15588, Republic of Korea.
authors: Jae-Sook
authors: JS
authors: Song
authors: Department of Pharmacy, College of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan, Gyeonggi-do 15588, Republic of Korea.
authors: Eun-Kyung
authors: EK
authors: Kim
authors: Department of Pharmacy, College of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan, Gyeonggi-do 15588, Republic of Korea.
authors: Yong-Won
authors: YW
authors: Choi
authors: College of Pharmacy and Research Institute of Pharmaceutical Sciences, Seoul National University, Republic of Korea.
authors: Won Keun
authors: WK
authors: Oh
authors: Department of Pharmacy, College of Pharmacy and Institute of Pharmaceutical Science and Technology, Hanyang University, Ansan, Gyeonggi-do 15588, Republic of Korea. Electronic address: ymikim12@hanyang.ac.kr.
authors: Young-Mi
authors: YM
authors: Kim
mesh_terms: D055372:AMP-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005978:Glutathione / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta / Q000235:genetics / Q000378:metabolism; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D017705:Lignans / Q000494:pharmacology*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D048369:MAP Kinase Kinase 1 / Q000235:genetics; D020935:MAP Kinase Signaling System / Q000187:drug effects; D008810:Mice, Inbred C57BL; D026323:Myristica; D051267:NF-E2-Related Factor 2 / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects; D010766:Phosphorylation / Q000187:drug effects; D020011:Protective Agents / Q000494:pharmacology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D020122:tert-Butylhydroperoxide
chemical_list: D017705:Lignans; D051267:NF-E2-Related Factor 2; D020011:Protective Agents; D017382:Reactive Oxygen Species; C524653:nectandrin-B; D020122:tert-Butylhydroperoxide; D008156:Luciferases; D000071679:Glycogen Synthase Kinase 3 beta; D055372:AMP-Activated Protein Kinases; D048369:MAP Kinase Kinase 1; C482125:MAP2K1 protein, human; D005978:Glutathione
keywords: Antioxidant enzymes; ERK; GSK-3β; Hepatocyte protection; Nectandrin B; Nrf2