Effect of silencing hypoxia-inducible factor-1 alpha by RNA interference on human breast carcinoma cell line / 中国医学科学院学报
Acta Academiae Medicinae Sinicae
;
(6): 670-674, 2006.
Article
in Chinese
| WPRIM
| ID: wpr-313709
ABSTRACT
<p><b>OBJECTIVE</b>To investigate the effect of short hairpin RNA (shRNA) targeting hypoxia-inducible factor-1 alpha (HIF-1 alpha) on the human breast carcinoma MCF-7 cell line.</p><p><b>METHODS</b>The hypoxia environment was achieved by treating cells with cobalt chloride. The shRNA eukaryotic expression vector targeting HIF-1 alpha was constructed, and transfected into MCF-7 cells through lipofectamine 2000. Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to study the expression of vascular endothelial growth factor (VEGF). The mRNA and protein level of HIF-1 alpha were detected by real-time PCR and Western blot. Sub-G1 apoptotic population analysis, Annexin V/PI binding assay, and DNA ladder analysis were applied to investigate the cell apoptosis. The cell cycle was detected by flow cytometry.</p><p><b>RESULTS</b>The mRNA and protein level of HIF-1 alpha increased after exposure of MCF-7 cells to hypoxia (P < 0.01). However, apoptosis was lower in hypoxia compared with normoxia (P < 0.05). The HIF-1 level of MCF-7 transfected with HIF-1 alpha shRNA decreased approximately 91.63% (P < 0.01). When the cells were treated with or without apoptosis inducer Ara-C, the apoptosis of MCF-7 cells transfected with HIF-1 alpha shRNA increased by 1.75 times (P < 0.01) and 61. 31 times (P < 0.01), respectively. The expression of VEGF in MCF-7 cells transfected with HIF-1 alpha shRNA decreased 66.8% compared with untransfected cells (P < 0.05). Cell cycle progression was inhibited when the MCF-7 cells were transfected with HIF-1 alpha shRNA.</p><p><b>CONCLUSIONS</b>HIF-1 alpha plays an anti-apoptotic role in human breast carcinoma MCF-7 cell line. The shRNA we designed targeting HIF-1 alpha in MCF-7 can promote cell apoptosis, inhibit the expression of VEGF, and delay cell cycle progression.</p>
Full text:
Available
Index:
WPRIM (Western Pacific)
Main subject:
Pathology
/
Breast Neoplasms
/
RNA, Messenger
/
Transfection
/
Down-Regulation
/
Cell Cycle
/
Apoptosis
/
RNA, Small Interfering
/
RNA Interference
/
Cell Line, Tumor
Limits:
Female
/
Humans
Language:
Chinese
Journal:
Acta Academiae Medicinae Sinicae
Year:
2006
Type:
Article
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