Mitochondria as the primary target of resveratrol-induced apoptosis in human retinoblastoma cells.
OBJECTIVE
To determine the molecular mechanisms by which resveratrol induces retinoblastoma tumor cell death.


METHODS
After resveratrol treatment, Y79 tumor cell viability was measured using a fluorescence-based assay, and proapoptotic and antiproliferative effects were characterized by Hoechst stain and flow cytometry, respectively. Mitochondrial transmembrane potential (DeltaPsim) was measured as a function of drug treatment using 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-benzamidazolocarbocyanin iodide (JC-1), whereas the release of cytochrome c from mitochondria was assayed by immunoblotting and caspase activities were determined by monitoring the cleavage of fluorogenic peptide substrates.


RESULTS
Resveratrol induced a dose- and time-dependent decrease in Y79 tumor cell viability and inhibited proliferation by inducing S-phase growth arrest and apoptotic cell death. Preceding cell death, resveratrol evoked a rapid dissipation of DeltaPsim. This was followed by the release of cytochrome c into the cytoplasm and a substantial increase in the activities of caspase-9 and caspase-3. Additionally, in a cell-free system, resveratrol directly induced the depolarization of isolated mitochondria.


CONCLUSIONS
These results demonstrate that resveratrol, a nontoxic natural plant compound, inhibits Y79 cell proliferation and stimulates apoptosis through activation of the mitochondrial (intrinsic) apoptotic pathway and may warrant further exploration as an adjuvant to conventional anticancer therapies for retinoblastoma.
Sareen
Dhruv
D
Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, WI 53792, USA.
van Ginkel
Paul R
PR
Takach
Jennifer C
JC
Mohiuddin
Ayesha
A
Darjatmoko
Soesiawati R
SR
Albert
Daniel M
DM
Polans
Arthur S
AS
D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D045304:Cytochromes c / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects* / Q000201:enzymology; D011808:Quinone Reductases / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D000077185:Resveratrol; D019572:Retinal Neoplasms / Q000378:metabolism / Q000473:pathology*; D012175:Retinoblastoma / Q000378:metabolism / Q000473:pathology*; D012264:Ribonucleotide Reductases / Q000037:antagonists & inhibitors; D013267:Stilbenes / Q000494:pharmacology*; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000972:Antineoplastic Agents, Phytogenic; D012333:RNA, Messenger; D013267:Stilbenes; D045304:Cytochromes c; D012264:Ribonucleotide Reductases; C419642:NRH - quinone oxidoreductase2; D011808:Quinone Reductases; C505407:CASP3 protein, human; C506433:CASP9 protein, human; D053148:Caspase 3; D053453:Caspase 9; D020169:Caspases; D000077185:Resveratrol