Enhanced levels of reactive oxygen species (ROS) and a prothrombotic state have been associated with remodelling processes in the systemic and pulmonary vasculature. However, the exact links connecting thrombin signalling with ROS production are not completely resolved.

Stimulation of pulmonary artery smooth muscle cells (PASMC) with thrombin resulted in a rapid increase in ROS levels within 5 minutes. This response was mimicked by the thrombin receptor PAR1 activating peptide TRAP6. In line, depletion of PAR1 by siRNA decreased ROS production. Subsequently, treatment with pertussis toxin, an inhibitor of Gai-proteins, prevented thrombin- stimulated ROS generation. Furthermore, thrombin activated the GTPase Rac1, involving PAR1 and Gai-proteins. In constrast a dominant negative Rac1 mutant or depletion of Rac1 by siRNA decreased thrombin-induced ROS generation. Since Rac1 has been shown to be required for activation of NOX1, ROS production was determined in NOX1-depleted PASMC. In contrast to controls, Rac1 and thrombin-induced ROS generation was decreased. In addition, NOX1 colocalized with PAR1 upon thrombin treatment. Furthermore, migration of PASMC was decreased upon thrombin stimulation after NOX1 depletion.

Taken together, these findings show that thrombin rapidly activated a NOX1-dependent oxidase involving PAR1, Ga_i and Rac1. Since thrombin, ROS and NOX1 stimulate migration and proliferation of PASMC, this mechanism might be involved in promoting pulmonary and possibly systemic vascular remodelling.