Induction of apoptosis in human retinoblastoma cells by topoisomerase inhibitors

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Abstract

PURPOSE:To examine the apoptotic effect induced in human retinoblastoma Y79 cells by camptothecin, etoposide, and amsacrine, to examine the effect of these drugs on the expression of many apoptosis-related modulators, and to test the antiapoptotic effect exerted by insulin-like growth factor-I (IGF-I).METHODS:Morphologic features of apoptosis were demonstrated using acridine orange- ethidium bromide staining and electron microscopy. DNA fragmentation was determined by means of an in situ cell detection procedure (TdT-dUTP terminal nick-end labeling [TUNEL]) or by electrophoresis on agarose gels and was quantified by enzyme-linked immunosorbent assay. The expression of apoptosis-related modulators was studied by western blot analysis. The processing of latent p53 was examined by means of pulse- chase analysis.RESULTS:Camptothecin, etoposide, and amsacrine induced apoptosis in Y79 cells in a dose-dependent manner; camptothecin was the most efficacious compound. The effect, which was dependent on macromolecular synthesis, appeared after a lag of 8 hours and increased for as long as 24 hours. It was lower in cells treated with IGF-I, a potent mitogenic factor. Camptothecin and etoposide increased the p53 level after 4 hours of treatment, before the onset of apoptosis. This effect seemed to be a consequence of the conversion of latent p53 to one that is transcriptionally active. The drugs also induced an increase in p53-related proteins, such as p21, Bax, and IGF binding protein-3 (IGF-BP3), and caused a significant reduction of the Bcl-2 level. The latter effect was less evident in cells pretreated with IGF-I.CONCLUSIONS:Topoisomerase inhibitors induce apoptosis in Y79 cells. This event is accompanied by a decrease in the expression of Bcl-2, a death antagonist, and an increase in that of Bax, a death agonist. A probable consequence of these modifications is the activation of ICE-like activity with degradation of poly-(adenosine diphosphate [ADP] ribose)-polymerase. Insulin-like growth factor-I exerts an antiapoptotic action in Y79 cells, and this function is most likely reduced by the overexpression of IGF-BP3 that is induced by drug treatment.
Lingua originaleEnglish
pagine (da-a)1300-1311
Numero di pagine12
RivistaINVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE
Volume39
Stato di pubblicazionePublished - 1998

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Topoisomerase Inhibitors
Retinoblastoma
Camptothecin
Apoptosis
Insulin-Like Growth Factor I
Etoposide
Amsacrine
Insulin-Like Growth Factor Binding Protein 3
Pharmaceutical Preparations
Poly Adenosine Diphosphate Ribose
Acridine Orange
Ethidium
Agar Gel Electrophoresis
DNA Fragmentation
Electron Microscopy
Western Blotting
Enzyme-Linked Immunosorbent Assay
Staining and Labeling

All Science Journal Classification (ASJC) codes

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

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@article{e039046fdbc0408fa286ea27540561bd,
title = "Induction of apoptosis in human retinoblastoma cells by topoisomerase inhibitors",
abstract = "PURPOSE:To examine the apoptotic effect induced in human retinoblastoma Y79 cells by camptothecin, etoposide, and amsacrine, to examine the effect of these drugs on the expression of many apoptosis-related modulators, and to test the antiapoptotic effect exerted by insulin-like growth factor-I (IGF-I).METHODS:Morphologic features of apoptosis were demonstrated using acridine orange- ethidium bromide staining and electron microscopy. DNA fragmentation was determined by means of an in situ cell detection procedure (TdT-dUTP terminal nick-end labeling [TUNEL]) or by electrophoresis on agarose gels and was quantified by enzyme-linked immunosorbent assay. The expression of apoptosis-related modulators was studied by western blot analysis. The processing of latent p53 was examined by means of pulse- chase analysis.RESULTS:Camptothecin, etoposide, and amsacrine induced apoptosis in Y79 cells in a dose-dependent manner; camptothecin was the most efficacious compound. The effect, which was dependent on macromolecular synthesis, appeared after a lag of 8 hours and increased for as long as 24 hours. It was lower in cells treated with IGF-I, a potent mitogenic factor. Camptothecin and etoposide increased the p53 level after 4 hours of treatment, before the onset of apoptosis. This effect seemed to be a consequence of the conversion of latent p53 to one that is transcriptionally active. The drugs also induced an increase in p53-related proteins, such as p21, Bax, and IGF binding protein-3 (IGF-BP3), and caused a significant reduction of the Bcl-2 level. The latter effect was less evident in cells pretreated with IGF-I.CONCLUSIONS:Topoisomerase inhibitors induce apoptosis in Y79 cells. This event is accompanied by a decrease in the expression of Bcl-2, a death antagonist, and an increase in that of Bax, a death agonist. A probable consequence of these modifications is the activation of ICE-like activity with degradation of poly-(adenosine diphosphate [ADP] ribose)-polymerase. Insulin-like growth factor-I exerts an antiapoptotic action in Y79 cells, and this function is most likely reduced by the overexpression of IGF-BP3 that is induced by drug treatment.",
author = "Renza Vento and Marianna Lauricella and Michela Giuliano",
year = "1998",
language = "English",
volume = "39",
pages = "1300--1311",
journal = "INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE",
issn = "0146-0404",

}

TY - JOUR

T1 - Induction of apoptosis in human retinoblastoma cells by topoisomerase inhibitors

AU - Vento, Renza

AU - Lauricella, Marianna

AU - Giuliano, Michela

PY - 1998

Y1 - 1998

N2 - PURPOSE:To examine the apoptotic effect induced in human retinoblastoma Y79 cells by camptothecin, etoposide, and amsacrine, to examine the effect of these drugs on the expression of many apoptosis-related modulators, and to test the antiapoptotic effect exerted by insulin-like growth factor-I (IGF-I).METHODS:Morphologic features of apoptosis were demonstrated using acridine orange- ethidium bromide staining and electron microscopy. DNA fragmentation was determined by means of an in situ cell detection procedure (TdT-dUTP terminal nick-end labeling [TUNEL]) or by electrophoresis on agarose gels and was quantified by enzyme-linked immunosorbent assay. The expression of apoptosis-related modulators was studied by western blot analysis. The processing of latent p53 was examined by means of pulse- chase analysis.RESULTS:Camptothecin, etoposide, and amsacrine induced apoptosis in Y79 cells in a dose-dependent manner; camptothecin was the most efficacious compound. The effect, which was dependent on macromolecular synthesis, appeared after a lag of 8 hours and increased for as long as 24 hours. It was lower in cells treated with IGF-I, a potent mitogenic factor. Camptothecin and etoposide increased the p53 level after 4 hours of treatment, before the onset of apoptosis. This effect seemed to be a consequence of the conversion of latent p53 to one that is transcriptionally active. The drugs also induced an increase in p53-related proteins, such as p21, Bax, and IGF binding protein-3 (IGF-BP3), and caused a significant reduction of the Bcl-2 level. The latter effect was less evident in cells pretreated with IGF-I.CONCLUSIONS:Topoisomerase inhibitors induce apoptosis in Y79 cells. This event is accompanied by a decrease in the expression of Bcl-2, a death antagonist, and an increase in that of Bax, a death agonist. A probable consequence of these modifications is the activation of ICE-like activity with degradation of poly-(adenosine diphosphate [ADP] ribose)-polymerase. Insulin-like growth factor-I exerts an antiapoptotic action in Y79 cells, and this function is most likely reduced by the overexpression of IGF-BP3 that is induced by drug treatment.

AB - PURPOSE:To examine the apoptotic effect induced in human retinoblastoma Y79 cells by camptothecin, etoposide, and amsacrine, to examine the effect of these drugs on the expression of many apoptosis-related modulators, and to test the antiapoptotic effect exerted by insulin-like growth factor-I (IGF-I).METHODS:Morphologic features of apoptosis were demonstrated using acridine orange- ethidium bromide staining and electron microscopy. DNA fragmentation was determined by means of an in situ cell detection procedure (TdT-dUTP terminal nick-end labeling [TUNEL]) or by electrophoresis on agarose gels and was quantified by enzyme-linked immunosorbent assay. The expression of apoptosis-related modulators was studied by western blot analysis. The processing of latent p53 was examined by means of pulse- chase analysis.RESULTS:Camptothecin, etoposide, and amsacrine induced apoptosis in Y79 cells in a dose-dependent manner; camptothecin was the most efficacious compound. The effect, which was dependent on macromolecular synthesis, appeared after a lag of 8 hours and increased for as long as 24 hours. It was lower in cells treated with IGF-I, a potent mitogenic factor. Camptothecin and etoposide increased the p53 level after 4 hours of treatment, before the onset of apoptosis. This effect seemed to be a consequence of the conversion of latent p53 to one that is transcriptionally active. The drugs also induced an increase in p53-related proteins, such as p21, Bax, and IGF binding protein-3 (IGF-BP3), and caused a significant reduction of the Bcl-2 level. The latter effect was less evident in cells pretreated with IGF-I.CONCLUSIONS:Topoisomerase inhibitors induce apoptosis in Y79 cells. This event is accompanied by a decrease in the expression of Bcl-2, a death antagonist, and an increase in that of Bax, a death agonist. A probable consequence of these modifications is the activation of ICE-like activity with degradation of poly-(adenosine diphosphate [ADP] ribose)-polymerase. Insulin-like growth factor-I exerts an antiapoptotic action in Y79 cells, and this function is most likely reduced by the overexpression of IGF-BP3 that is induced by drug treatment.

UR - http://hdl.handle.net/10447/65756

M3 - Article

VL - 39

SP - 1300

EP - 1311

JO - INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE

JF - INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE

SN - 0146-0404

ER -