Copyright © 2007, European Society of Cardiology
NADPH oxidase mediates angiotensin II-induced endothelin-1 expression in vascular adventitial fibroblasts
aDepartment of Community Health Sciences, Brock University, St. Catharines, ON, Canada L2S 3Y6
bDepartment of Pharmacology, University of Saskatchewan, Saskatoon, SK, Canada
cMyocardial Biology Unit, Boston University Medical Center, Boston, Massachusetts, USA
* Corresponding author. Tel.: +1 905 688 4819x4819; fax: +1 905 688 8954. huidi.wang{at}brocku.ca
Objective We have recently reported that adventitial fibroblasts are able to express endothelin-1 (ET-1) in response to angiotensin II (Ang II) stimulation. However, the mechanism by which this occurs in the adventitia remains unclear. As Ang II has been reported to increase oxidant production by NADPH oxidase, we examined the role of this complex in Ang II stimulated ET-1 expression in vascular adventitial fibroblasts.
Methods and results Adventitial fibroblasts were isolated and cultured from mouse aorta. Cells were treated with Ang II (100 nmol/L) in the presence or absence of NADPH oxidase inhibitors, apocynin (100 µmol/L) and diphenyleneiodonium (10 µmol/L), superoxide scavengers, SOD (350 units/mL), tempol (100 µmol/L), tiron (100 µmol/L), and ET-receptor antagonists (10 µM), BQ123 (for ETA-) and BQ788 (for ETB-). PreproET-1 mRNA and ET-1 level were determined by relative RT-PCR and ELISA, respectively. Type I procollagen-
-I (collagen) level was detected by Western blot. Superoxide anion (superoxide) production was determined by coelenterazine or lucigenin chemiluminescence. Ang II-induced collagen expression was inhibited by BQ123, suggesting that adventitial ET-1 plays a significant role in regulating the extracellular matrix. NADPH oxidase inhibitors and superoxide scavengers significantly decreased Ang II-induced ET-1 mRNA and peptide expression, superoxide production as well as collagen expression. Furthermore, deletion of gp91phox (a key subunit of NADPH oxidase) and overexpression of SOD1 attenuated Ang II-induced responses.
Conclusion Ang II-evoked expression of ET-1 in adventitial fibroblasts appears to be mediated, at least in part, by NADPH oxidase. Functionally, this mechanism stimulates collagen expression thereby implicating the adventitia as a potential contributor to the vascular pathophysiology associated with oxidative stress and vascular remodeling.
KEYWORDS Adventitial fibroblast; Superoxide anion; NADPH oxidase; Ang II; ET-1
1 Ryan Boyd and Min Zhu contribute equally.
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