Disruption of mitochondria during tocotrienol-induced apoptosis in MDA-MB-231 human breast cancer cells.
Tocotrienols, which are Vitamin E isoforms, are known to inhibit the growth of human breast cancer cells due partly to apoptosis. However, the characterization of tocotrienol-induced apoptosis is incomplete, particularly what happens during the initiation phase that precedes execution of the cells. The objective of this study was to clarify the apoptotic effects of tocotrienols, with especial emphasis in determining if the mitochondria-mediated death pathway is activated when human breast cancer cells are incubated with a specific tocotrienol isomer. During incubation with gamma-tocotrienol, MDA-MB-231 human breast cancer cells showed membrane blebbing, and apoptotic bodies were present. Upon 4',6-diamidino-2-phenylindole staining of the cells, chromatin condensation and fragmentation were observed. Additionally, the annexin V-binding assay detected the translocation of membrane phospholipid during earlier analysis of the cells. Taken together, these results further establish that gamma-tocotrienol can induce apoptosis in human breast cancer cells. To help elucidate how gamma-tocotrienol induced the apoptosis, some important parameters related to the mitochondria-mediated death pathway were examined next. In gamma-tocotrienol-treated cells, the mitochondria were disrupted. Collapse of the mitochondrial membrane potential was detected, and cytochrome c was released later from mitochondria. However, expression of Bax and Bcl-2 (mRNA and protein) did not change. Furthermore, poly-(ADP-ribose)-polymerase cleavage was not detected, suggesting that caspases were not involved in the gamma-tocotrienol-induced apoptosis. These results imply that cytochrome c is not the critical protein released from mitochondria that triggers gamma-tocotrienol-induced apoptosis in MDA-MB-231 cells.
Takahashi
Keiko
K
Cellular and Molecular Nutrition Research Laboratory, Graduate Program in Nutrition, The University of North Carolina at Greensboro, Greensboro, NC 27402-6170, USA.
Loo
George
G
D017304:Annexin A5 / Q000378:metabolism; D000975:Antioxidants / Q000494:pharmacology*; D017209:Apoptosis; D001943:Breast Neoplasms / Q000473:pathology; D020169:Caspases / Q000378:metabolism; D002843:Chromatin / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D006801:Humans; D006834:Hydrazines / Q000378:metabolism; D020782:Matrix Metalloproteinases / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D011506:Proteins / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D024508:Tocotrienols / Q000494:pharmacology*; D014407:Tumor Cells, Cultured; D051028:bcl-2-Associated X Protein
C417561:Alexa 488 hydrazide; D017304:Annexin A5; D000975:Antioxidants; C494826:BAX protein, human; D002843:Chromatin; D006834:Hydrazines; D011506:Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D024508:Tocotrienols; D051028:bcl-2-Associated X Protein; D045304:Cytochromes c; C499320:PARP1 protein, human; D000071137:Poly (ADP-Ribose) Polymerase-1; D011065:Poly(ADP-ribose) Polymerases; D020169:Caspases; D020782:Matrix Metalloproteinases