title: Complement-mediated injury reversibly disrupts glomerular epithelial cell actin microfilaments and focal adhesions.
abstract: BACKGROUND
  Foot process effacement and condensation of the glomerular epithelial cell (GEC) cytoskeleton are manifestations of passive Heymann nephritis, a model of complement-mediated membranous nephropathy.
  
  
  METHODS
  To study the effects of complement on the actin cytoskeleton in this model, we have used an in vitro system in which GECs are sublethally injured using a combination of complement-fixing anti-Fx1A IgG and human serum as a source of complement. We examined the effects of this injury on the organization of the cytoskeleton and focal contacts using immunohistology and immunochemistry.
  
  
  RESULTS
  By immunofluorescence, sublethal complement-mediated injury was accompanied by a loss of actin stress fibers and focal contacts but retention of matrix-associated integrins. Full recovery was seen after 18 hours. Western blot analysis showed no change in the cellular content of the focal contact proteins. Inhibition of the calcium-dependent protease calpain did not prevent injury. In addition, cycloheximide during recovery did not inhibit the reassembly of stress fibers or focal contacts. Injury was associated with a reduction in tyrosine phosphorylation of paxillin and a currently unidentified 200 kDa protein, but inhibition of tyrosine phosphatase activity with sodium vanadate did not prevent injury. Cellular adenosine triphosphate content was significantly reduced in injured cells.
  
  
  CONCLUSIONS
  These results document reversible, complement-dependent disruption of actin microfilaments and focal contacts leading to the dissociation of the cytoskeleton from matrix-attached integrins. This may explain the altered cell-matrix relationship accompanying podocyte effacement in membranous nephropathy.
authors.affiliation: Evans Department of Medicine and Clinical Research, Boston University Medical Center, Massachusetts, USA.
authors.forename: P S
authors.initials: PS
authors.lastname: Topham
authors.forename: S A
authors.initials: SA
authors.lastname: Haydar
authors.forename: R
authors.initials: R
authors.lastname: Kuphal
authors.forename: J D
authors.initials: JD
authors.lastname: Lightfoot
authors.forename: D J
authors.initials: DJ
authors.lastname: Salant
mesh_terms: D008841:Actin Cytoskeleton / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D000199:Actins / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D002448:Cell Adhesion / Q000276:immunology; D015815:Cell Adhesion Molecules / Q000032:analysis; D002460:Cell Line; D015938:Complement Membrane Attack Complex / Q000737:chemistry / Q000276:immunology; D003165:Complement System Proteins / Q000276:immunology*; D003513:Cycloheximide / Q000494:pharmacology; D003598:Cytoskeletal Proteins / Q000032:analysis; D004847:Epithelial Cells / Q000737:chemistry / Q000166:cytology / Q000201:enzymology; D005455:Fluorescent Antibody Technique; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D005921:Glomerulonephritis / Q000276:immunology* / Q000378:metabolism; D007074:Immunoglobulin G / Q000494:pharmacology; D039201:Integrin alpha3beta1; D016023:Integrins / Q000032:analysis; D007678:Kidney Glomerulus / Q000166:cytology* / Q000276:immunology*; D051419:Paxillin; D010750:Phosphoproteins / Q000032:analysis; D019000:Phosphotyrosine / Q000032:analysis / Q000276:immunology; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011505:Protein-Tyrosine Kinases / Q000032:analysis; D051381:Rats; D017474:Receptors, Laminin / Q000032:analysis; D016608:Talin / Q000032:analysis; D014638:Vanadates / Q000494:pharmacology; D016596:Vinculin / Q000032:analysis
chemical_list: D000199:Actins; D015815:Cell Adhesion Molecules; D015938:Complement Membrane Attack Complex; D003598:Cytoskeletal Proteins; D007074:Immunoglobulin G; D039201:Integrin alpha3beta1; D016023:Integrins; D051419:Paxillin; D010750:Phosphoproteins; D011500:Protein Synthesis Inhibitors; C496063:Pxn protein, rat; D017474:Receptors, Laminin; D016608:Talin; D016596:Vinculin; D019000:Phosphotyrosine; D014638:Vanadates; D003165:Complement System Proteins; D003513:Cycloheximide; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496052:Ptk2 protein, rat