Superoxide from NADPH oxidase upregulates type 5 phosphodiesterase in human vascular smooth muscle cells: inhibition with iloprost and NONOate

Br J Pharmacol. 2008 Nov;155(6):847-56. doi: 10.1038/bjp.2008.300. Epub 2008 Jul 28.

Abstract

Background and purpose: To determine whether there is an association between vascular NADPH oxidase (NOX), superoxide, the small GTPase Rac(1) and PDE type 5 (PDE5) in human vascular smooth muscle cell (hVSMCs).

Experimental approach: hVSMCs were incubated with xanthine-xanthine oxidase (X-XO; a superoxide generating system) or the thromboxane A(2) analogue, U46619 (+/-superoxide dismutase (SOD) or apocynin) for 16 h. The expression of PDE5 and NOX-1 was assessed using Western blotting and superoxide measured. The role of Rac(1) in superoxide generation was assessed by overexpressing either the dominant-negative or constitutively active Rac isoforms. The effects of iloprost, DETA-NONOate and the Rho-kinase inhibitor, Y27632, on PDE5 and NOX-1 expression were also studied.

Key results: Following 16 h incubation, U46619 and X-XO promoted the expression of PDE5 and NOX-1, an effect blocked by SOD or apocynin when co-incubated over the same time course. X-XO and U46619 both promoted the formation of superoxide. Overexpression of dominant-negative Rac(1) or addition of iloprost, DETA-NONOate or Y27632 completely blocked both superoxide release and PDE5 protein expression and activity.

Conclusions and implications: These data demonstrate that superoxide derived from NOX upregulates the expression of PDE5 in human VSMCs. As PDE5 hydrolyses cyclic GMP, this effect may blunt the vasculoprotective actions of NO.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cells, Cultured
  • Cyclic Nucleotide Phosphodiesterases, Type 5 / metabolism
  • Humans
  • Iloprost / pharmacology*
  • Muscle, Smooth, Vascular* / drug effects
  • Muscle, Smooth, Vascular* / enzymology
  • Muscle, Smooth, Vascular* / metabolism
  • NADPH Oxidases / metabolism*
  • Nitric Oxide Donors / pharmacology
  • Nitroso Compounds / pharmacology*
  • Phosphodiesterase 5 Inhibitors*
  • Saphenous Vein / cytology
  • Superoxides / metabolism*
  • Superoxides / pharmacology
  • Up-Regulation
  • Vasodilator Agents / pharmacology

Substances

  • Nitric Oxide Donors
  • Nitroso Compounds
  • Phosphodiesterase 5 Inhibitors
  • Vasodilator Agents
  • Superoxides
  • 2,2'-(hydroxynitrosohydrazono)bis-ethanamine
  • NADPH Oxidases
  • Cyclic Nucleotide Phosphodiesterases, Type 5
  • Iloprost