Sildenafil citrate and sildenafil nitrate (NCX 911) are potent inhibitors of superoxide formation and gp91phox expression in porcine pulmonary artery endothelial cells

Br J Pharmacol. 2005 Sep;146(1):109-17. doi: 10.1038/sj.bjp.0706305.


Acute respiratory distress syndrome (ARDS) is associated with increased superoxide (O(2)(*-)) formation in the pulmonary vasculature and negation of the bioavailability of nitric oxide (NO). Since NO inhibits NADPH oxidase expression through a cyclic GMP-mediated mechanism, sildenafil, a type V phosphodiesterase inhibitor, may be therapeutically effective in ARDS through an augmentation of NO-mediated inhibition of NADPH oxidase. Therefore, the effect of sildenafil citrate and NO-donating sildenafil (NCX 911) on O(2)(*-) formation and gp91(phox) (active catalytic subunit of NADPH oxidase) expression was investigated in cultured porcine pulmonary artery endothelial cells (PAECs). PAECs were incubated with 10 nM TXA(2) analogue, 9,11-dideoxy-9alpha,11alpha-methanoepoxy-prostaglandin F(2alpha) (U46619) (+/-sildenafil or NCX 911), for 16 h and O(2)(*-) formation measured spectrophometrically and gp91(phox) using Western blotting. The role of the NO-cGMP axis was studied using morpholinosydnonimine hydrochloride (SIN-1), the diethylamine/NO complex (DETA-NONOate), the guanylyl cyclase inhibitor, 1H-{1,2,4}oxadiazolo{4,3-a}quinoxalin-1-one (ODQ), and the protein kinase G inhibitor, 8-bromoguanosine-3',5'-cyclic monophosphorothioate, Rp-isomer (Rp-8-Br-cGMPS). NO release was studied using a fluorescence assay and O(2)(*-)-NO interactions by measuring nitrites. After a 16-h incubation with 10 nM U46619, both NCX 911 and sildenafil elicited a concentration-dependent inhibition of O(2)(*-) formation and gp91(phox) expression, NCX 911 being more potent (IC(50); 0.26 nM) than sildenafil citrate (IC(50); 1.85 nM). These inhibitory effects were reversed by 1 microM ODQ and 10 microM Rp-8-Br-cGMPS. NCX 911 stimulated the formation of cGMP in PAECs and generated NO in a cell-free system to a greater degree than sildenafil citrate. The inhibitory effect of sildenafil was augmented by 1 muM SIN-1 and blocked partially by the eNOS inhibitor 10 microM N(5)-(1-iminoethyl)-ornithine (L-NIO). Acutely, sildenafil and NCX 911 also inhibited O(2)(*-) formation, again blocked by 1 microM ODQ. NCX 911 reacted with O(2)(*-) generated by xanthine oxidase, an effect that was inhibited by superoxide dismutase (500 U ml(-1)). Since O(2)(*-) formation plays contributory role in ARDS, both sildenafil citrate and NCX 911 may be indicated for treating ARDS through suppression of NADPH oxidase expression and therefore of O(2)(*-) formation and preservation of NO bioavailability.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid / pharmacology
  • Animals
  • Cells, Cultured
  • Cyclic GMP / analogs & derivatives
  • Cyclic GMP / metabolism
  • Cyclic GMP / pharmacology
  • Endothelial Cells / drug effects
  • Endothelial Cells / metabolism
  • Guanylate Cyclase / antagonists & inhibitors
  • Male
  • Membrane Glycoproteins / antagonists & inhibitors*
  • Membrane Glycoproteins / metabolism
  • NADPH Oxidases / antagonists & inhibitors*
  • NADPH Oxidases / metabolism
  • Nitric Oxide / metabolism
  • Nitric Oxide Synthase / antagonists & inhibitors
  • Ornithine / analogs & derivatives
  • Ornithine / pharmacology
  • Oxadiazoles / pharmacology
  • Piperazines / pharmacology*
  • Protein Kinase Inhibitors / pharmacology
  • Pulmonary Artery
  • Purines
  • Quinoxalines / pharmacology
  • Sildenafil Citrate
  • Sulfones
  • Superoxides / antagonists & inhibitors*
  • Superoxides / metabolism
  • Swine
  • Thionucleotides / pharmacology


  • 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one
  • Membrane Glycoproteins
  • Oxadiazoles
  • Piperazines
  • Protein Kinase Inhibitors
  • Purines
  • Quinoxalines
  • Sulfones
  • Thionucleotides
  • Superoxides
  • 8-bromoguanosino-3',5'-cyclic monophosphorothioate
  • Nitric Oxide
  • N(G)-iminoethylornithine
  • 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid
  • Sildenafil Citrate
  • Ornithine
  • Nitric Oxide Synthase
  • NADPH Oxidases
  • Guanylate Cyclase
  • Cyclic GMP