Mechanism of growth inhibitory effect of mitomycin-C on cultured human retinal pigment epithelial cells: Apoptosis and cell cycle arrest

Citation
Sg. Kang et al., Mechanism of growth inhibitory effect of mitomycin-C on cultured human retinal pigment epithelial cells: Apoptosis and cell cycle arrest, CURR EYE R, 22(3), 2001, pp. 174-181
Citations number
35
Language
INGLESE
art.tipo
Article
Categorie Soggetti
da verificare
Journal title
CURRENT EYE RESEARCH
ISSN journal
0271-3683 → ACNP
Volume
22
Issue
3
Year of publication
2001
Pages
174 - 181
Database
ISI
SICI code
0271-3683(200103)22:3<174:MOGIEO>2.0.ZU;2-2
Abstract
Purpose. To investigate the therapeutic potential of Mitomycin-C (MMC) in t he management of proliferative vitreoretinopathy, the antiproliferative eff ect of MMC on cultured human retinal pigment epithelial (RPE) cells were in vestigated in vitro. Methods. Drug sensitivities of cultured human RPE cells to MMC were determi ned using the tetrazolium dye assay. In order to detect the presence of apo ptosis, DNA fragmentation was assessed by DAPI staining, and TdT-dUTP nick- end labeling (TUNEL) assay. The relative amount of DNA fragmentation was qu antified by flow cytometric analysis. To analyze the cell cycle response of RPE cells to MMC, flow cytometric analysis of propidium iodide stained nuc lei was performed. The levels of proteins related to DNA damage in the RPE cells were then determined by Western blot analysis. Results. MMC inhibited cell proliferation in a dose-dependent manner. The m ajority of RPE cells following treatment with 10 mug/ml of MMC exhibited fr agmented nuclei as observed by DAPI staining and TUNEL assay. Cell cycle an alysis demonstrated an accumulation of cells arrested in S and G2/M phase f ollowing treatment with 1 mug/ml of MMC. At 10 mug/ml of MMC, a dramatic in crease of the cell population in the sub G1 peak, which can be considered a marker of cell death by apoptosis, was observed by flow cytometry. Western blot analysis of p53 and p21 revealed a gradual increase in the level of t hese proteins when RPE cells were exposed to increasing concentrations of M MC. Conclusions. This study demonstrated that the response of RPE cells to MMC was bi-directional: 1) partial arrest of the cell cycle at S, G2/M phase, a nd 2) induction of apoptotic cell death.