title: Prooxidative effects of green tea polyphenol (-)-epigallocatechin-3-gallate on the HIT-T15 pancreatic beta cell line.
abstract: Epigallocatechin-3-gallate (EGCG) is the main polyphenolic constituent in green tea and is believed to function as an antioxidant. However, increasing evidence indicates that EGCG produces reactive oxygen species (ROS) and subsequent cell death. In this study, we investigated the prooxidative effects of EGCG on the HIT-T15 pancreatic beta cell line. Dose-dependent cell viability was monitored with the cell counting kit-8 assay, while the induction of apoptosis was analyzed by a cell death ELISA kit and comet assay. Extracellular H(2)O(2) was determined using the Amplex Red Hydrogen Peroxide Assay Kit. Intracellular oxidative stress was measured by fluorometric analysis of 2',7'-dichlorofluorescin (DCFH) oxidation using DCFH diacetate (DA) as the probe. Treatment with EGCG (5-100 microM) decreased the viability of pancreatic beta cells, caused concomitant increases in apoptotic cell death, and increased the production of H(2)O(2) and ROS. Catalase, the iron-chelating agent diethylenetriaminepentaacetic acid, and the Fe(II)-specific chelator o-phenanthroline all suppressed the effects of EGCG, indicating the involvement of both H(2)O(2) and Fe(II) in the mechanism of action of EGCG. The antioxidant N-acetyl-cysteine and alpha-lipoic acid also suppressed the effects of EGCG. Furthermore, EGCG did not scavenge exogenous H(2)O(2), but rather, it synergistically increased H(2)O(2)-induced oxidative cell damage in pancreatic beta cells. Together, these findings suggest that in the HIT-T15 pancreatic beta cell line, EGCG mediated the generation of H(2)O(2), triggering Fe(II)-dependent formation of a highly toxic radical that in turn induced oxidative cell damage.
authors.affiliation: Department of Endocrinology and Metabolism, Kyung Hee University School of Medicine, 1 Hoegi-dong, Dongdaemun-gu, Seoul 130-701, South Korea.
authors.forename: Kwang Sik
authors.initials: KS
authors.lastname: Suh
authors.forename: Suk
authors.initials: S
authors.lastname: Chon
authors.forename: Seungjoon
authors.initials: S
authors.lastname: Oh
authors.forename: Sung Woon
authors.initials: SW
authors.lastname: Kim
authors.forename: Jin-Woo
authors.initials: JW
authors.lastname: Kim
authors.forename: Young Seol
authors.initials: YS
authors.lastname: Kim
authors.forename: Jeong-Taek
authors.initials: JT
authors.lastname: Woo
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002392:Catechin / Q000031:analogs & derivatives* / Q000633:toxicity; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D020552:Comet Assay; D004305:Dose-Response Relationship, Drug; D005419:Flavonoids / Q000633:toxicity; D006861:Hydrogen Peroxide / Q000378:metabolism; D050417:Insulin-Secreting Cells / Q000187:drug effects* / Q000378:metabolism; D016877:Oxidants / Q000633:toxicity*; D018384:Oxidative Stress / Q000187:drug effects; D010636:Phenols / Q000633:toxicity; D059808:Polyphenols; D017382:Reactive Oxygen Species / Q000378:metabolism; D013662:Tea / Q000737:chemistry
chemical_list: D005419:Flavonoids; D016877:Oxidants; D010636:Phenols; D059808:Polyphenols; D017382:Reactive Oxygen Species; D013662:Tea; D002392:Catechin; D006861:Hydrogen Peroxide; C045651:epigallocatechin gallate