NADPH oxidase 1 mediates upregulation of thromboxane A2 synthase in human vascular smooth muscle cells: Inhibition with iloprost

Thromboxane A2 (TXA2) upregulates and activates NADPH oxidase (Nox) both of which are associated with cardiovascular disease. The aim of this study, therefore, was to investigate the relationship between thromboxane A2 synthase (TXAS) status and Nox in human vascular smooth muscle cells (hVSMCs), in particular, whether superoxide (O2▪−) derived from Nox influences TXAS expression and activity. hVSMCs were incubated with TNFα: (10 ng/ml), TXA2 mimetic U46619 (100 nM), 8-isoprostane F2α (8-IP; 100 nM) and hypoxia. Expression of TXAS was assessed using western blotting and quantitative PCR. The role of Nox1 and Nox4 was studied using apocynin and mRNA silencing. The effect of the thromboxane receptor antagonist picotamide and of iloprost, a prostacyclin (PGI2) analogue was also studied. TNF-α, U46619 and 8-IP and hypoxia all augmented TXAS expression as well as TXA2 formation, effects inhibited by apocynin. Nox-1 (but not Nox4) gene silencing inhibited the increase in TXAS expression and activity. Both picotamide and iloprost inhibited the upregulation of TXAS as well as TXA2 formation induced by TNF-α, U46619 and 8-isoprostane F2α and hypoxia. It is concluded that upregulation of TXA2 synthase expression and activity in human VSMCs is mediated by an a priori upregulation of Nox1 and represents a self amplifying cascade. The inhibition of this effect with iloprost consolidates that PGI2 plays a protective anti-oxidative role in the vasculature and that picotamide and like drugs may be effective in reducing the incidence of cardiovascular disease associated with an oxidative aetiology.

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