Abstract
NADPH oxidase is the major source of superoxide production in cardiovascular tissues. We reported previously that PG (prostaglandin) F2alpha caused hypertrophy of vascular smooth muscle cells by induction of NOX1, a catalytic subunit of NADPH oxidase. PGF2alpha-induced NOX1 expression was mediated by transactivation of the EGF (epidermal growth factor) receptor and subsequent activation of ERK (extracellular-signal-regulated kinase) 1/2, PI3K (phosphoinositide 3-kinase) and ATF-1 (activating transcription factor-1), a member of the CREB (cAMP-response-element-binding protein)/ATF family. As the receptor for PGF2alpha is known to activate PKC (protein kinase C), involvement of PKC in up-regulation of NOX1 expression was investigated in A7r5 cells. GF109203x, a non-selective inhibitor of PKC, dose-dependently suppressed the induction of NOX1 mRNA by PGF2alpha. Whereas an inhibitor of the conventional PKC, Gö 6976, and a PKCeta translocation-inhibitor peptide had no effect, an inhibitor of PKCdelta, rottlerin, significantly attenuated the PGF2alpha-induced increase in NOX1 mRNA. Gene silencing of PKCdelta by RNA interference significantly suppressed the PGF2alpha-induced increase in NOX1 mRNA, as well as phosphorylation of the EGF receptor, ERK1/2 and ATF-1. Silencing of the PKCdelta gene also attenuated the PDGF (platelet-derived growth factor)- induced increase in NOX1 mRNA and transactivation of the EGF receptor. Moreover, the augmented synthesis of the protein induced by PGF2alpha or PDGF was abolished by gene silencing of PKCdelta. These results suggest that PKCdelta-mediated transactivation of the EGF receptor is elicited not only by PGF2alpha, but also by PDGF, and that the subsequent activation of ERK1/2 and ATF-1 leads to up-regulation of NOX1 gene expression and ensuing hypertrophy in the vascular cell lineage.