title: Vardenafil modulates bladder contractility through cGMP-mediated inhibition of RhoA/Rho kinase signaling pathway in spontaneously hypertensive rats.
abstract: BACKGROUND
  Phosphodiesterase type 5 inhibitors (PDE5i), the most widely used drugs for erectile dysfunction, could also improve lower urinary tract symptoms, essentially due to overactive bladder (OAB), a condition hypothesized to be a result of an increased RhoA/Rho-kinase (ROCK) signaling. Phosphorylation/inactivation of RhoA by cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG) activity has been described in vascular smooth muscle.
  
  
  OBJECTIVE
  The aim of this paper was to investigate whether vardenafil-induced cGMP accumulation reduces RhoA/ROCK signaling in bladder.
  
  
  METHODS
  Spontaneously hypertensive rats (SHRs), a strain genetically prone to develop OAB, were treated with vardenafil (10 mg/kg/day) for 2 weeks. Wistar-Kyoto rats (WKY) were used as control. In vitro experiments were performed in human bladder smooth muscle cells (hBCs).
  
  
  METHODS
  Urodynamic parameters were registered in vivo in anesthetized WKY and SHRs. RhoA/ROCK activity in bladder was evaluated by molecular and functional studies in tissues and cells.
  
  
  RESULTS
  The intercontraction interval and bladder capacity, and were decreased in SHRs and restored by vardenafil. The in vitro relaxant effect of the ROCK inhibitor Y-27632 was higher in bladder strips from SHR than from WKY and reduced by vardenafil. Nomega-nitro-L-arginine-methyl-ester (a NO-synthase inhibitor, 40 mg/kg/day during the last week of the 2-week treatment with vardenafil) partially antagonized vardenafil effect on Y-27632 responsiveness. Vardenafil prevented RhoA membrane translocation/activation, decreased ROCK activity, and increased cGMP levels in vivo (rat) and in vitro (hBCs). Exposing hBCs to vardenafil increased Ser(188) RhoA phosphorylation, to the same extent as the PDE5-insensitive PKG agonist Sp-8-Br-PET-cGMP. Moreover, vardenafil inhibited several RhoA-dependent functions in hBCs, including smooth muscle gene transcription and endothelin-1-induced migration. These effects were reverted by the PKG inhibitor KT 5823, further suggesting a cGMP/PKG-dependency. In hBCs, vardenafil was active in the low nanomolar range.
  
  
  CONCLUSIONS
  This is the first study demonstrating that the effect of vardenafil on OAB could be partially determined by a cGMP-dependent RhoA/ROCK signaling inhibition.
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Annamaria
authors.initials: A
authors.lastname: Morelli
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Sandra
authors.initials: S
authors.lastname: Filippi
authors.affiliation: Bayer Healthcare, Wuppertal, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Sandner
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Benedetta
authors.initials: B
authors.lastname: Fibbi
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Aravinda K
authors.initials: AK
authors.lastname: Chavalmane
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Enrico
authors.initials: E
authors.lastname: Silvestrini
authors.affiliation: Department of Anatomy, Histology and Forensic Medicine, University of Florence, Florence, Italy.
authors.forename: Erica
authors.initials: E
authors.lastname: Sarchielli
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy.
authors.forename: Linda
authors.initials: L
authors.lastname: Vignozzi
authors.affiliation: Department of Urology, University of Florence, Florence, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Gacci
authors.affiliation: Department of Urology, University of Florence, Florence, Italy.
authors.forename: Marco
authors.initials: M
authors.lastname: Carini
authors.affiliation: Department of Anatomy, Histology and Forensic Medicine, University of Florence, Florence, Italy.
authors.forename: Gabriella B
authors.initials: GB
authors.lastname: Vannelli
authors.affiliation: Department of Clinical Physiopathology, University of Florence, Florence, Italy;. Electronic address: m.maggi@DFC.UNIFI.IT.
authors.forename: Mario
authors.initials: M
authors.lastname: Maggi
mesh_terms: D000818:Animals; D015153:Blotting, Western; D006152:Cyclic GMP / Q000378:metabolism*; D006973:Hypertension / Q000201:enzymology* / Q000503:physiopathology*; D007093:Imidazoles / Q000494:pharmacology*; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects / Q000201:enzymology; D058986:Phosphodiesterase 5 Inhibitors; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology*; D010879:Piperazines / Q000494:pharmacology*; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects*; D013450:Sulfones / Q000494:pharmacology; D014227:Triazines / Q000494:pharmacology; D053201:Urinary Bladder, Overactive / Q000188:drug therapy*; D014563:Urodynamics / Q000187:drug effects; D000069058:Vardenafil Dihydrochloride; D054460:rho-Associated Kinases / Q000187:drug effects*; D020742:rhoA GTP-Binding Protein / Q000187:drug effects*
chemical_list: D007093:Imidazoles; D058986:Phosphodiesterase 5 Inhibitors; D010726:Phosphodiesterase Inhibitors; D010879:Piperazines; D013450:Sulfones; D014227:Triazines; D000069058:Vardenafil Dihydrochloride; D054460:rho-Associated Kinases; D020742:rhoA GTP-Binding Protein; D006152:Cyclic GMP