Effects of Smac gene over-expression on the radiotherapeutic sensitivities of cervical cancer cell line HeLa.
- Author:
Li-Duan ZHENG
1
;
Zhou-Fang XIONG
;
Jian-Wen ZHU
;
Ze-Hua WANG
Author Information
- Publication Type:Journal Article
- MeSH: Apoptosis; radiation effects; Carrier Proteins; genetics; physiology; Caspase 3; Caspases; metabolism; Female; Gene Transfer, Horizontal; HeLa Cells; Humans; Intracellular Signaling Peptides and Proteins; Mitochondrial Proteins; genetics; physiology; Radiation Tolerance; Uterine Cervical Neoplasms; drug therapy; enzymology; pathology
- From: Chinese Medical Journal 2005;118(3):226-230
- CountryChina
- Language:English
-
Abstract:
BACKGROUNDThe second mitochondria-derived activator of caspases (Smac) is a novel proapoptotic gene, which plays an important role in the apoptosis-inducing effects of irradiation on tumor cells. The purpose of this study was to investigate the effects of extrinsic Smac gene transfer and its over-expression in radiotherapeutic sensitivities of cervical cancer cells.
METHODSAfter the Smac gene was transferred into the cervical cancer cell line HeLa, subcloned cells were obtained by persistent G418 selection. Cellular Smac gene expression was detected by RT-PCR and Western blot, while in vitro cell viabilities were detected by trypan blue staining assay. After treatment with X-ray irradiation, cellular radiotherapeutic sensitivities were investigated by tetrazolium bromide colorimetry. Cellular apoptosis and its rate were determined by electronic microscopy, annexin V-FITC and propidium iodide staining flow cytometry. The expression and activities of cellular caspase-3 were assayed by Western blot and colorimetry.
RESULTSSmac mRNA and protein levels in HeLa/Smac cells and the selected subclone cell line of cervical cancer were significantly higher than those of HeLa (P < 0.01). There was no significant difference in cellular viabilities between them (P > 0.05). However, after irradiation with 8 Gy X-ray, growth activities of HeLa/Smac were reduced by 22.42% (P < 0.01). When compared with those of HeLa, partial HeLa/Smac cells presented characteristic morphological changes of apoptosis under electronic microscope, with higher apoptosis rates (16.4% vs. 6.2%, P < 0.01); the caspase-3 expression levels in HeLa/Smac cells were improved significantly (P < 0.01), while its activities were increased by 3.42 times (P < 0.01).
CONCLUSIONSStable transfer of the extrinsic Smac gene and its over-expression in cervical cancer cell line could significantly enhance the expression and activities of cellular caspase-3 and ameliorate apoptosis-inducing effects of irradiation on cancer cells, which was a novel strategy to improve radiotherapeutic effects on cervical cancer.
