Involvement of transcription factor activator protein-2alpha in doxazosin-induced HeLa cell apoptosis.
OBJECTIVE
To investigate the pro-apoptotic effects of alpha1-adrenergic inhibitor doxazosin in HeLa cells and the potential involvement of transcription factor activator protein-2alpha (AP-2alpha) in doxazosin-induced apoptosis.


METHODS
The HeLa cells were exposed to various concentrations of doxazosin for 16 h. Apoptosis was detected using a DNA fragmentation assay, Hoechst 33258 staining, and flow cytometric analysis. The expression of AP-2alpha and caspase-3 was detected by relative quantitative RT-PCR and Western blot assays, respectively. After the respective transfections of the HeLa cells with AP-2alpha overexpressing constructs and an antisense oligonucleotide against AP-2alpha, apoptosis was assessed by flow cytometric analysis, and the expression of AP-2alpha and caspase-3 was detected by relative quantitative RT-PCR and Western blot assays. The colorimetric assay was performed to detect the caspase-3 activity.


RESULTS
Treatment with various concentrations of doxazosin for 16 h increased the apoptotic rate and total cell death rate of the HeLa cells in a dose-dependent manner and upregulated the expression of AP-2alpha and caspase-3 in a dose-dependent manner. A dose-dependent increase was observed in the caspase-3 activity. Overexpressing AP-2alpha led to the increased rate of doxazosin-induced apoptosis and the total cell death, whereas doxazosin-induced apoptosis and the total cell death in HeLa cells decreased by antisense AP-2alpha. Furthermore, overexpressing AP-2alpha increased the expression and activity of caspase-3, whereas antisense AP-2alpha in part abolished the increased effects of doxazosin on caspase-3 expression and activity.


CONCLUSIONS
Doxazosin induces apoptosis in HeLa cells in a dose-dependent manner, and transcription factor AP-2alpha is functionally involved in doxazosin-induced HeLa cell apoptosis.
Gan
Lu
L
The Key Laboratory of Protein Chemistry and Developmental Biology, Ministry of Education of China, College of Life Sciences, Hu-nan Normal University, Changsha 410081, China.
Zhu
Dong-xing
DX
Yang
Li-ping
LP
Liu
Ru-shi
RS
Yan
Feng
F
Zhang
Jian
J
D000317:Adrenergic alpha-Antagonists / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D006690:Bisbenzimidazole / Q000378:metabolism; D053148:Caspase 3 / Q000235:genetics / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D017292:Doxazosin / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic; D006367:HeLa Cells; D006801:Humans; D013997:Time Factors; D050656:Transcription Factor AP-2 / Q000235:genetics / Q000378:metabolism*
D000317:Adrenergic alpha-Antagonists; D005456:Fluorescent Dyes; D050656:Transcription Factor AP-2; D053148:Caspase 3; D006690:Bisbenzimidazole; D017292:Doxazosin