Conference: 2006 International PHA Conference and Scientific Sessions
Release Date: 06.23.2006
Presentation Type: Abstracts
Grenett H.E., Olave N.C., Pasten M.C., Benza R.L., Wolkowicz P.E.
Department of Medicine, Division of Cardiovascular Disease, University of Alabama at Birmingham, Birmingham, Alabama USA
BACKGROUND: Reactive oxygen species (ROS) like superoxide anion (O2. -) play an important role as signaling molecules in vascular cells, and NADPH oxidases are important contributors to ROS production within the vasculature. NADPH oxidase consists of two membrane-bound subunits (p22phox and gp91phox) and two cytosolic subunits (p47phox and p67phox). Elevated expression of p22phox in pulmornary arteries has been implicated in the pathogenesis and pathophysiology of pulmonary artery hypertension (PAH).. Quercetin, is a polyphenol compund that occurs naturally in vegetables, fruits and tea. Intake of quercetin in dietary polyphenols has been suggested to play an important role in the human cardiovascular system although the molecular mechanism of its cardioprotective benefit remains unknown. Here we report the effect of quercetin on the regulation of human pulmonary endotheila cells (HPAECs)
METHODS: HPAECs were incubated in the presence or absence of 10 µM quercetin for 0- 24 hr. Total cytoplasmic protein and RNA were extracted and p22phox expression was analyzed by Western blot analysis and by reverse transcription followed by real-time PCR. HPAECs also were transiently transfected with a 1,010-bp promoter fragment of the p22phox gene ligated into the pGL3 basic luciferase reporter vector. Transfected HPAECs were incubated with 10µM quercetin for 4 hr and endothelial cell extracts were prepared and assayed for luciferase activity.
RESULTS: Real time PCR analyses demonstrated that quercetin repressed HPAECs p22phox gene expression in a time-dependent manner. A two hour exposure to 10µM quercetin produced a 60% decrease in endothelial cell p22phox mRNA. Western blot analysis showed decreased p22phox protein expression of 3- to 4-fold at 16 to 24 hr. Transiently transfected HPAECs with the p22/luc showed a ~ 3-fold decrease in promoter activity in the presence of quercetin.
CONCLUSIONS: These results indicate that quercetin downregulates p22phox gene expression in cultured HPAECs and that the 1,010-bp p22phox promoter construct contains regulatory elements that respond to quercetin. Quercetin-induced decreased in EC p22phox expression might reduce vascular ROS production and contribute, in part, to the cardioprotective benefit associated with a dietary polyphenols consumption.