[Effects of a triple mutant hypoxia-inducible factor-1α on proliferation and vascular endothelial growth factor expression in human microvascular endothelial cells].
OBJECTIVE
To investigate the effects of a recombinant adenovirus-mediated triple mutant hypoxia-inducible factor-1α (HIF-1α) on the proliferation and vascular endothelial growth factor (VEGF) expression in human microvascular endothelial cells (hMVECs).


METHODS
The adenovirus vector of the triple mutant HIF-1α (Ad-HIF-1α(564/402/803)), adenovirus vector of wild-type HIF-1α (Ad-HIF-1α(nature)), Ad-lacZ and Ad-Null were amplified in HEK293A cells, and the adenoviruses were purified and titrated. Dual luciferase reporter assay system was employed to detect the transcriptional activities of wild-type and triple mutant HIF-1α. After infection of the hMVECs with the adenoviruses, the cellular protein expressions of HIF-1α and VEGF were detected using Western blotting, and the cell proliferation was assessed by MTS assay.


RESULTS
The transcriptional activity of the triple mutant HIF-1α was significantly higher than that of wildtype HIF-1α in the infected hMVECs (P<0.001). The protein levels of HIF-1α and VEGF in cells infected with Ad-HIF-1α(564/402/803) were significantly higher than those in cells infected with other adenoviruses, and HIF-1α dose-dependently up-regulated VEGF protein expression. The absorbance was significantly higher in Ad-HIF-1α(564/402/803) group than in the other groups (P<0.01) on the third and fifth days after infection.


CONCLUSIONS
The recombinant adenovirus-mediated triple mutant HIF-1α expression is stable under normoxic condition. The triple mutant HIF-1α can up-regulate the expression of VEGF protein in hMVECs to promote the cell proliferation.
Department of Cardiology, Southern Medical University, Guangzhou, China. xiaopei1586@yahoo.com.cn
Jingxian
J
Pei
Yuegang
Y
Wang
Cheng
C
Liu
Xuan
X
Wei
Mingyan
M
Li
Jianwei
J
Chen
Pingsheng
P
Wu
D000256:Adenoviridae / Q000235:genetics; D049109:Cell Proliferation; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology; D005822:Genetic Vectors; D057809:HEK293 Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000494:pharmacology*; D055806:Microvessels / Q000166:cytology*; D011994:Recombinant Proteins / Q000235:genetics; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D011994:Recombinant Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A