title: Biochemical mechanisms of bornyl caffeate induced cytotoxicity in rat pheochromocytoma PC12 cells.
abstract: The chemopreventive and antineoplastic activities of caffeic acid derivatives are highly dependent on the chemical structures and cancer cell types. The objective of the present study was to investigate the cytotoxicity of bornyl caffeate and the underlying molecular mechanisms in rat pheochromocytoma PC12 cells. Our initial studies demonstrated that bornyl caffeate exhibited potent cytotoxicity in PC12 cells in a concentration- and time-dependent manner. By examining the cell morphology on a fluorescence microscope and detecting the cell surface phosphoserine with Annexin V-FITC, we proposed that bornyl caffeate could induce apoptosis in PC12 cells. We tested this hypothesis by investigating the effects of bornyl caffeate on several apoptosis-related biomarkers. These experiments showed that bornyl caffeate induced the up-regulation of Bax and down-regulation of Bcl-xl, the disruption of mitochondrial membrane potential, the activation of caspase 3 and the cleavage of PARP. Mechanistic studies further revealed that bornyl caffeate caused the depletion of glutathione (GSH), generation of superoxide ion and progressive activation of p38 mitogen-activate protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) in a concentration-dependent manner. In particular, GSH depletion appeared to be the most important mechanism underlying the cytotoxicity of bornyl caffeate. The preservation of the intracellular GSH contents with N-acetyl-L-cysteine (NAC), GSH and vitamin C abolished the effect of bornyl caffeate on the activation of p38 MAPK and JNK, preserved the integrity of mitochondrial membrane and ultimately rescued the cells from drug-induced cell death. These results suggest that bornyl caffeate induces apoptosis in PC12 cells via stimulating the depletion of GSH, the generation of reactive oxygen species (ROS) and the dissipation of mitochondrial transmembrane potential.
authors.affiliation: School of Chinese Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, 10 Sassoon Road, Pokfulam, Hong Kong, China.
authors.forename: Chuanbin
authors.initials: C
authors.lastname: Yang
authors.affiliation: School of Chinese Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, 10 Sassoon Road, Pokfulam, Hong Kong, China.
authors.forename: Jia
authors.initials: J
authors.lastname: Zhao
authors.affiliation: RITS and NWU Joint Laboratory for New Drugs Research of TCM, Research Institute of Tsinghua University in Shenzhen, Shenzhen 518000, China.
authors.forename: Weijing
authors.initials: W
authors.lastname: Pei
authors.affiliation: RITS and NWU Joint Laboratory for New Drugs Research of TCM, Research Institute of Tsinghua University in Shenzhen, Shenzhen 518000, China; Key Laboratory of Resource Biology and Biotechnology in Western China, School of Life Sciences, Northwest University, Xi'an, Shaanxi 710069, China.
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Zheng
authors.affiliation: School of Chinese Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, 10 Sassoon Road, Pokfulam, Hong Kong, China. Electronic address: jrong@hkucc.hku.hk.
authors.forename: Jianhui
authors.initials: J
authors.lastname: Rong
mesh_terms: D000310:Adrenal Gland Neoplasms / Q000188:drug therapy* / Q000201:enzymology; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D002470:Cell Survival / Q000187:drug effects*; D003373:Coumaric Acids / Q000494:pharmacology* / Q000627:therapeutic use; D005434:Flow Cytometry; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D005978:Glutathione / Q000032:analysis / Q000378:metabolism; D048670:MAP Kinase Kinase 4 / Q000032:analysis / Q000378:metabolism; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D016716:PC12 Cells; D010673:Pheochromocytoma / Q000188:drug therapy* / Q000201:enzymology; D051381:Rats; D051028:bcl-2-Associated X Protein / Q000032:analysis / Q000378:metabolism; D051020:bcl-X Protein / Q000032:analysis / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000032:analysis / Q000378:metabolism
chemical_list: C494813:Bcl2l1 protein, rat; D003373:Coumaric Acids; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; C115950:bornyl caffeate; D048051:p38 Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D005978:Glutathione
keywords: Bornyl caffeate; Cytotoxicity; GSH depletion; Mitochondrial dysfunction; ROS