title: Peroxynitrite-induced alpha-actinin nitration and contractile alterations in isolated human myocardial cells.
abstract: OBJECTIVE
  Peroxynitrite-mediated myocardial protein nitration has been associated with a depressed cardiac pump function. In the present study, an attempt was made to elucidate the molecular background of peroxynitrite-evoked alterations in the human myocardium.
  
  
  METHODS
  Isometric force generation was measured in permeabilized human ventricular myocytes and biochemical methods were employed to identify the proteins affected by peroxynitrite-induced nitrotyrosine formation.
  
  
  RESULTS
  The maximal Ca(2+)-activated isometric force (pCa=4.75) decreased to zero with increasing concentrations of peroxynitrite in a concentration-dependent manner (IC50: 55+/-4 microM; based on a total of 75 myocytes). However, there were no differences before and after the application of 50 microM peroxynitrite in the Ca(2+)-sensitivity of force production (pCa50: 5.89+/-0.02 and 5.86+/-0.04), in the steepness of the Ca(2+)-force relationship (nHill: 2.22+/-0.11 and 2.42+/-0.25), and in the actin-myosin turnover kinetics (k(tr) at saturating [Ca2+]: 1.14+/-0.03 1/s and 1.05+/-0.07 1/s) (P>0.05). Nevertheless, 50 muM peroxynitrite greatly deteriorated the cross-striation pattern and induced a slight, but significant, increase in the passive force component (from 2.1+/-0.1 to 2.5+/-0.2 kN/m2; n=57 cells), reflecting ultrastructural alterations. Western immunoblots revealed that 50 microM peroxynitrite selectively induced the nitration of a protein with an apparent molecular mass of about 100 kDa. Subsequent immunoprecipitation assays identified this nitrated protein as alpha-actinin, a major Z-line protein.
  
  
  CONCLUSIONS
  These results suggest alpha-actinin as a novel target for peroxynitrite in the human myocardium; its nitration induces a contractile dysfunction, presumably by decreasing the longitudinal transmission of force between adjacent sarcomeres.
authors: Division of Clinical Physiology, Institute of Cardiology, UD MHSC, P.O. Box 1, H-4004 Debrecen, Hungary.
authors: Attila
authors: A
authors: Borbély
authors: Attila
authors: A
authors: Tóth
authors: István
authors: I
authors: Edes
authors: László
authors: L
authors: Virág
authors: Julius Gy
authors: JG
authors: Papp
authors: András
authors: A
authors: Varró
authors: Walter J
authors: WJ
authors: Paulus
authors: Jolanda
authors: J
authors: van der Velden
authors: Ger J M
authors: GJ
authors: Stienen
authors: Zoltán
authors: Z
authors: Papp
mesh_terms: D000185:Actinin / Q000378:metabolism*; D015153:Blotting, Western / Q000379:methods; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D003864:Depression, Chemical; D004305:Dose-Response Relationship, Drug; D006352:Heart Ventricles; D006801:Humans; D015151:Immunoblotting / Q000379:methods; D009200:Myocardial Contraction / Q000187:drug effects*; D032383:Myocytes, Cardiac / Q000378:metabolism*; D030421:Peroxynitrous Acid / Q000378:metabolism / Q000494:pharmacology*; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
chemical_list: D000185:Actinin; D030421:Peroxynitrous Acid; C002744:3-nitrotyrosine; D014443:Tyrosine; D002118:Calcium