Casticin potentiates TRAIL-induced apoptosis of gastric cancer cells through endoplasmic reticulum stress

PLoS One. 2013;8(3):e58855. doi: 10.1371/journal.pone.0058855. Epub 2013 Mar 11.

Abstract

Background: Casticin is one of the main active components obtained from Fructus Viticis and has been reported to exert anti-carcinogenic activity on a variety of cancer cells but the precise mechanism underlying this activity remains unclear.

Materials and methods: Apoptotic activities of casticin (1.0 µmol/l) and TRAIL (25, 50 ng/ml) alone or in combination in the gastric cancer cell lines BGC-823, SGC-7901 and MGC-803 were detected by the use of a cell apoptosis ELISA detection kit, flow cytometry (FCM) with propidium iodide (PI) staining and activities of caspase-3, -8 and -9 by ELISA and cleavage of polyADP-ribose polymerase (PARP) protein using western blot analysis. Death receptors (DR) expression levels were evaluated using FCM analysis and western blotting. 2', 7'-dichlorofluorescein diacetate (DCFH-DA) was used as a probe to measure the increase in reactive oxygen species (ROS) levels in cells. Multiple interventions, such as siRNA transfection and pharmacological inhibitors were used to explore the mechanisms of these actions.

Results: Subtoxic concentrations of casticin significantly potentiated TRAIL-induced cytotoxicity and apoptosis in BGC-823, SGC-7901 and MGC-803 cells. Casticin dramatically upregulated DR5 receptor expression but had no effects on DR4 or decoy receptors. Deletion of DR5 by siRNA significantly reduced the apoptosis induced by the co-application of TRAIL and casticin. Gene silencing of the CCAAT/enhancer binding protein homologous protein (CHOP) and pretreatment with salubrinal, an endoplasmic reticulum (ER) stress inhibitor, attenuated casticin-induced DR5 receptor expression, and apoptosis and ROS production. Casticin downregulated the expression levels of the cell survival proteins cFLIP, Bcl-2, XIAP, and survivin. In addition, casticin also induced the expressions of DR5 protein in other gastric cancer cells (SGC-7901 and MGC-803).

Conclusion/significance: Casticin enhances TRAIL-induced apoptosis through the downregulation of cell survival proteins and the upregulation of DR5 receptors through actions on the ROS-ER stress-CHOP pathway.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / drug effects*
  • Apoptosis / genetics
  • Apoptosis Regulatory Proteins / genetics
  • Apoptosis Regulatory Proteins / metabolism
  • Cell Line, Tumor
  • Endoplasmic Reticulum Stress / drug effects*
  • Flavonoids / pharmacology*
  • Flavonoids / toxicity
  • Gene Expression Regulation, Neoplastic / drug effects
  • Gene Knockout Techniques
  • Humans
  • RNA Interference
  • Reactive Oxygen Species / metabolism
  • Receptors, TNF-Related Apoptosis-Inducing Ligand / genetics
  • Receptors, TNF-Related Apoptosis-Inducing Ligand / metabolism
  • Stomach Neoplasms / genetics
  • Stomach Neoplasms / metabolism*
  • TNF-Related Apoptosis-Inducing Ligand / pharmacology*
  • TNF-Related Apoptosis-Inducing Ligand / toxicity

Substances

  • Apoptosis Regulatory Proteins
  • Flavonoids
  • Reactive Oxygen Species
  • Receptors, TNF-Related Apoptosis-Inducing Ligand
  • TNF-Related Apoptosis-Inducing Ligand
  • casticin

Grant support

This work was supported by the Project of NSFC (number 30760248), the Project of Scientific Research of Hunan Province the Administration Bureau of Traditional Chinese Medicine (number 2010081), the Project of Scientific Research of Hunan Province, Department of Education (number 10C0975), Major Project Item of Scientific Research of Hunan Province the Department of Education (number 09A054) and the Project of Scientific Research of Changsha City Bureau of Science and Technology (number K1104060-31). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.