[Effect of NADPH oxidase on hypoxia-inducible factor-1alpha and endothelin-1 expression in human umbilical endothelial cells and its possible mechanism].
OBJECTIVE
To study the effect of NADPH oxidase on hypoxia-inducible factor (HIF)-1alpha and endothelin (ET)-1 expression in human umbilical endothelia cells (HUVECs) and its possible mechanism.


METHODS
Twenty-five bottles of HUVECs culture fluid were randomly assigned into five groups: group A (normoxic control), group B (hypoxic), group C (NADPH oxidase inhibitor apocynin + normoxic), group D (H2O2 which can degrade HIF-1alpha rapidly+hypoxic) and group E (H2O2+apocynin+normoxic), with five bottles in each group. The culture supernates were collected and the total protein was extracted 3 hrs after treatment. Western Blot and ELISA were used to detect the HIF-1alpha protein expression in HUVECs and the ET-1 level in the culture supernates respectively.


RESULTS
There was a lower expression of HIF-1alpha protein (0.336 +/- 0.012) and lower ET-1 levels (5.87 +/- 2.22 pg/mL) in group A. The HIF-1alpha protein expression in groups B and C (0.773 +/- 0.018 and 0.888 +/- 0.022) and ET-1 levels (95.38 +/- 8.06 and 33.67 +/- 4.21 pg/mL) were noticeably higher than in group A (P < 0.05). The groups D and E had the HIF-1alpha protein expression levels similar to group A, but the ET-1 levels in group D (108.43 +/- 8.38 pg/mL) and group E (109.66 +/- 5.80 pg/mL) were significantly higher than in group A (P < 0.05).


CONCLUSIONS
Hypoxia or apocynin can increase the HIF-1alpha and ET-1 expression in HUVECs. H2O2 can inhibit the HIF-1alpha expression but increase the ET-1levels. It is speculated that NADPH oxidase as an oxygen sensor regulates the HIF-1alpha expression by changing the intracellular redox reaction and that except HIF-1, H2O2 might contribute to ET-1 synthesis and release.
Dong
Lan
L
Department of Neonatology, Guangzhou Children's Hospital, Guangzhou 510120, China. kezchen@163.com
Chen
Ke-Zheng
KZ
He
Liu-Fang
LF
D015153:Blotting, Western; D015687:Cell Hypoxia; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D019332:Endothelin-1 / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000032:analysis / Q000378:metabolism*; D019255:NADPH Oxidases / Q000502:physiology*
D019332:Endothelin-1; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019255:NADPH Oxidases