Involvement of Smac, p53, and caspase pathways in induction of apoptosis by gossypol in human retinoblastoma cells
Resource
Mol. Vis., 18(211-14), 2033-2042
Journal
Mol. Vis.
Journal Volume
18
Journal Issue
211-14
Pages
2033-2042
Date Issued
2012
Date
2012
Author(s)
Hsiao, Wei-Ting
Tsai, Ming-Dar
Jow, Guey-Mei
Tien, Lu-Tai
Lee, Yih-Jing
Abstract
Purpose: Retinoblastoma is a malignant tumor of the retina usually occurring in young children. To date, the conventional treatments for retinoblastoma have been enucleation, cryotherapy, external beam radiotherapy, or chemotherapy. Most of these treatments, however, have possible side effects, including blindness, infections, fever, gastrointestinal toxicity, and neurotoxicity. More effective treatments are therefore imperative. Gossypol has been reported as a potential inhibitor of cell proliferation in various types of cancers, such as prostate cancer, breast cancer, leukemia, and lung cancer. This study investigates the possible antiproliferative effect of gossypol on retinoblastoma. ;Methods: Human retinoblastoma cells were cultured with various concentrations of gossypol and checked for cell viability with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Nuclear condensation caused by cell apoptosis was detected by staining retinoblastoma cells with 4',6-diamidino-2-phenylindole (DAPI), counting those with condensed nuclei, and determining the percentage of apoptotic cells. In addition, the stages of apoptosis and phases in cell cycles were examined with flow cytometry. The possible signal transduction pathways involved were examined with a protein array assay and western blot analysis. ;Results: After incubation, the cell survival rate was significantly lower after treatment with 5, 10, and 20 mu M of gossypol. The maximum antisurvival effect of gossypol was observed at 20 mu M, and the number of apoptotic cells was higher in the preparations cultured with 10 and 20 mu M of gossypol. The results in flow cytometry indicated that at concentrations of 10 and 20 mu M, gossypol increased the proportion of early- and late-apoptotic retinoblastoma cells and induced cell arrest of retinoblastoma cells at the same concentrations. This antiproliferative effect was later confirmed by upregulating the expression of death receptor 5 (DR5), caspase 8, caspase 9, caspase 3, cytochrome C, tumor protein 53 (p53), and second mitochondria-derived activator of caspases (Smac) in the signal transduction pathways. ;Conclusions: We concluded that gossypol has an antiproliferative effect on retinoblastoma cells.
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