Safrole induces G0/G1 phase arrest via inhibition of cyclin E and provokes apoptosis through endoplasmic reticulum stress and mitochondrion-dependent pathways in human leukemia HL-60 cells

Anticancer Res. 2012 May;32(5):1671-9.

Abstract

Safrole, a component of Piper betle inflorescence, is a carcinogen which has been demonstrated to induce apoptosis on human oral cancer HSC-3 cells in vitro and to inhibit HSC-3 cells in xenograft tumor cells in vivo. In our previous study, safrole promoted phagocytosis by macrophages and natural killer cell cytotoxicity in normal BALB/c mice. The cytotoxic effects of safrole on HL-60 cells were investigated by using flow cytometric analysis, comet assay, 4',6-diamidino-2-phenylindole (DAPI) staining, western blotting and confocal laser microscopy. The obtained results indicate that safrole induced a cytotoxic response through reducing the percentage of viable cells and induction of apoptosis in HL-60 cells in a dose-dependent manner. DAPI staining and comet assay also showed that safrole induced apoptosis (chromatin condensation) and DNA damage in HL-60 cells. The flow cytometric assay showed that safrole increased the production of reactive oxygen species (ROS) and Ca(2+) and reduced the mitochondrial membrane potential in HL-60 cells. Safrole enhanced the levels of the pro-apoptotic protein BAX, inhibited those of the anti-apoptotic protein BCL-2 and promoted the levels of apoptosis-inducing factor (AIF) and endonuclease G (Endo G) in HL-60 cells. Furthermore, safrole promoted the expression of glucose-regulated protein 78 (GRP78), growth arrest- and DNA damage-inducible gene 153 (GADD153) and of activating transcription factor 6α (ATF-6α). Based on these findings, we suggest that safrole-induced apoptosis in HL-60 cells is mediated through the ER stress and intrinsic signaling pathways.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis / drug effects*
  • Apoptosis Inducing Factor / metabolism
  • Calcium / metabolism
  • Cell Cycle Checkpoints / drug effects*
  • Cyclin E / antagonists & inhibitors*
  • DNA Damage
  • Endoplasmic Reticulum Chaperone BiP
  • Endoplasmic Reticulum Stress / drug effects*
  • G1 Phase / drug effects
  • HL-60 Cells
  • Humans
  • Membrane Potential, Mitochondrial / drug effects
  • Mice
  • Mitochondria / physiology*
  • Resting Phase, Cell Cycle / drug effects
  • Safrole / pharmacology*
  • Transcription Factor CHOP / metabolism

Substances

  • Apoptosis Inducing Factor
  • Cyclin E
  • DDIT3 protein, human
  • Endoplasmic Reticulum Chaperone BiP
  • HSPA5 protein, human
  • Hspa5 protein, mouse
  • Transcription Factor CHOP
  • Safrole
  • Calcium