Oxidative stress and expression of p22phox are involved in the up-regulation of tissue factor in vascular smooth muscle cells in response to activated platelets

Vascular injury after balloon angioplasty results in the rapid activation o f platelets leading to the release of growth factors and vasoactive substan ces, In addition, up-regulation of tissue factor (TF) and an increased prod uction of reactive oxygen species (ROS) have been detected at sites of vasc ular injury. We investigated whether platelet-derived products (PDP) releas ed from activated human platelets increase ROS production, resulting in the induction of TF expression in vascular smooth muscle cells (SMC). PDP indu ced a time- and concentration-dependent increase in ROS generation in cultu red SMC that was mediated mainly by PDGF-AB and TGF-beta 1 and impaired by the flavin inhibitor diphenylene iodonium. Increased ROS formation was asso ciated with enhanced mRNA levels of the small NAD(P)H oxidase subunit p22ph ox or its smooth muscle isoform. Transient transfection with a p22phox anti sense vector decreased PDP-induced ROS generation. PDP up-regulated TF mRNA expression, which was redox sensitive and reduced by transfection of the p 22phox antisense vector. In addition, PDP-stimulated reporter gene activity of two TF promoter constructs was decreased by coexpression of the p22phox antisense vector. These results indicate that activated platelets up-regul ate TF expression and that this response involves ROS generation and a p22p hox-containing NAD(P)H oxidase in SMC.