The expression of Bcl-XL, Bcl-XS and p27Kip1 in topotecan-induced apoptosis in hepatoblastoma HepG2 cell line.

Abstract

BACKGROUND:

To assess the efficacy of topotecan, a topoisomerase I specific inhibitor in S-phase, the reagent-induced apoptosis and cytotoxicity as well as related proteins expression, had been preliminarily investigated in human hepatoblastoma HepG2 cells.

METHODS:

Microculture tetrazolium assay (MTT), HE staining, transmission electron microscopy (TEM), flow cytometry (FCM), quantitative immunocytochemistry (QI), gene tranfection and RNAi technology were employed to carry out the exploration.

RESULTS:

Topotecan could potently kill HepG2 cells via inducing apoptosis and demonstrated strong cytotoxicity in a time, dose-dependent manner with IC50 of about 95 mu g/L. According to morphologic observation and FCM analyses, it was confirmed that the drug treatment, causing significant S-phase arrest, could trigger a typical interphase apoptosis, the main traits of which were identified as chromatin pycnosis and cytoplasm condensation. It was shown that the expression of Bcl-XL was simultaneously down-regulated with the up-regulation of Bcl-XS in cytoplasm, which was possibly a key downstream event following the topotecan-induced DNA damage in nucleus. The expression level of p27Kip1, a negative regulator in cell cycle at G1/S transient, was also elevated. Transfection of pcDNA 3.1-p27Kip1 into HepG2 cells could abrogate the cytotoxicity in a degree while silence of p27Kip1 with siRNA in drug treatments could significantly increased the chemosensitivity, strongly indicating that the up-regulation of p27Kip1 was not an apoptosis-promoting, but a self-rescue response against drug by moderate G0/G1 arrest.

CONCLUSION:

Topotecan had potent cytotoxicity against HepG2 cells by triggering an interphase apoptosis possibly mediated by increasing the ratio of Bcl-XS/Bcl-XL. Up-regulation of p27Kip1in TPT treatments could be a protective response for self-rescue and silence of the gene markedly augmented TPT cytotoxicity. Therefore, the experiment in vitro could provide a new idea for the clinical chemotherapy based on the combination of traditional drugs with the specific-siRNA targeted on the protective response gene.

• Authors: Jun Zhang, Chao Cheng, Chuang-Long He, Yu-Jian Zhou, Yang Cao
• Journal: Cancer investigation (Cancer Invest) Vol. 26 Issue 5 Pg. 456-63 (Jun 2008) ISSN: 1532-4192 [Electronic] England
• PMID: 18568767 (Publication Type: Journal Article)

Chemical References

• Antineoplastic Agents
• BCL2L1 protein, human
• CDKN1B protein, human
• Enzyme Inhibitors
• Intracellular Signaling Peptides and Proteins
• Topoisomerase I Inhibitors
• bcl-X Protein
• Cyclin-Dependent Kinase Inhibitor p27
• Topotecan
• DNA Topoisomerases, Type I

Topics

• Antineoplastic Agents (pharmacology)
• Apoptosis (drug effects)
• Cell Cycle (drug effects)
• Cell Line
• Cell Proliferation (drug effects)
• Cell Survival (drug effects)
• Cyclin-Dependent Kinase Inhibitor p27
• DNA Topoisomerases, Type I (metabolism)
• Dose-Response Relationship, Drug
• Enzyme Inhibitors (pharmacology)
• Flow Cytometry
• Hepatoblastoma (enzymology, metabolism, ultrastructure)
• Humans
• Immunohistochemistry
• Inhibitory Concentration 50
• Intracellular Signaling Peptides and Proteins (genetics, metabolism)
• Liver Neoplasms (enzymology, metabolism, ultrastructure)
• Microscopy, Electron, Transmission
• RNA Interference
• Signal Transduction (drug effects)
• Time Factors
• Topoisomerase I Inhibitors
• Topotecan (pharmacology)
• Transfection
• bcl-X Protein (metabolism)