title: Modulation of the ASK1-MKK3/6-p38/MAPK signalling pathway mediates sildenafil protection against chemical hypoxia caused by malonate.
abstract: OBJECTIVE
  PD5 inhibitors have recently been reported to exert beneficial effects against ischaemia-reperfusion injury in several organs. However, there are few studies regarding their neuroprotective effects in brain ischaemia. The present study was designed to assess the effects of sildenafil against chemical hypoxia induced by malonate. Intrastriatal injection of malonate produces energy depletion and striatal lesions similar to that seen in cerebral ischaemia through mechanisms that involve generation of reactive oxygen species (ROS).
  
  
  METHODS
  Volume lesion was analysed by cytochrome oxidase histochemistry. Generation of reactive species was determined by in situ visualization of superoxide production and nitrotyrosine measurement. Protein levels were determined by Western blot after subcellular fractionation.
  
  
  RESULTS
  Sildenafil, given 30 min before malonate, significantly decreased the lesion volume in the rat. This protective effect cannot be attributed to any effect on ROS production but to the inhibition of downstream pathways. Thus, malonate induced the activation of apoptosis signal-regulating kinase-1 (ASK1) and two MAPK kinases, MKK3/6 and MKK7, which lead to an increased phosphorylation of JNK and p38 MAPK, effects that were blocked by sildenafil. Selective inhibitors of p38 and JNK (SB203580 or SP600125, respectively) were used in combination with malonate in order to evaluate the plausible implication of these pathways in the protection afforded by sildenafil. While inhibition of p38 provided a significant protection against malonate-induced neurotoxicity, inhibition of JNK did not.
  
  
  CONCLUSIONS
  Sildenafil protects against the chemical hypoxia induced by malonate through the regulation of the ASK1-MKK3/6-p38/MAPK signalling pathway.
authors.affiliation: Department of Pharmacology, School of Pharmacy, University of Navarra, Pamplona, Spain.
authors.forename: L
authors.initials: L
authors.lastname: Barros-Miñones
authors.forename: L
authors.initials: L
authors.lastname: Orejana
authors.forename: B
authors.initials: B
authors.lastname: Goñi-Allo
authors.forename: V
authors.initials: V
authors.lastname: Suquía
authors.forename: I
authors.initials: I
authors.lastname: Hervías
authors.forename: N
authors.initials: N
authors.lastname: Aguirre
authors.forename: E
authors.initials: E
authors.lastname: Puerta
mesh_terms: D000284:Administration, Oral; D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D002534:Hypoxia, Brain / Q000139:chemically induced* / Q000517:prevention & control*; D007093:Imidazoles / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D008297:Male; D008314:Malonates / Q000494:pharmacology*; D018696:Neuroprotective Agents / Q000494:pharmacology*; D058986:Phosphodiesterase 5 Inhibitors / Q000494:pharmacology*; D010879:Piperazines / Q000494:pharmacology*; D011687:Purines / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species / Q000378:metabolism*; D000068677:Sildenafil Citrate; D013449:Sulfonamides / Q000494:pharmacology*
chemical_list: D000873:Anthracenes; D007093:Imidazoles; D008314:Malonates; D018696:Neuroprotective Agents; D058986:Phosphodiesterase 5 Inhibitors; D010879:Piperazines; D011687:Purines; D011725:Pyridines; D017382:Reactive Oxygen Species; D013449:Sulfonamides; C432165:pyrazolanthrone; C030290:malonic acid; D000068677:Sildenafil Citrate; C093642:SB 203580