title: Regulation of mesangial cell apoptosis and proliferation by intracellular Ca(2+) signals.
abstract: BACKGROUND
  In inflammatory glomerular diseases, proliferation, as well as apoptosis of mesangial cells (MCs), has been shown histomorphologically. Both processes may regulate the cellular content of the mesangium by closely influencing each other. In the present study, we examined whether the cytoplasmic free Ca(2+) concentration [Ca(2+)](i) is involved as a key second messenger in the regulation of proliferative and apoptotic events.
  
  
  METHODS
  Thapsigargin, an inhibitor of the endoplasmic Ca(2+)-Mg(2+)-ATPase, was used as a test substance to investigate the role of [Ca(2+)](i) in signaling MC apoptosis and growth in vitro. Apoptosis was determined by nuclear chromatin staining with Hoechst 33258, by a [3H]-thymidine-based DNA fragmentation assay or by flow cytometry detecting binding of FITC-conjugated annexin V. Proliferation was measured by [3H]-thymidine incorporation into acid-precipitable material and corroborated by cell counting.
  
  
  RESULTS
  Thapsigargin significantly induced apoptosis and inhibited proliferation dose dependently in nanomolar concentrations without evoking necrotic damage when administered not longer than 12 hours. Significant apoptosis was measurable after a six-hour treatment of MCs with thapsigargin. Determination of [Ca(2+)](i) by fura-2-dependent spectrofluorometry showed that thapsigargin was able to induce prolonged [Ca(2+)](i) rises that could be prevented by preincubation with the intracellular Ca(2+) chelator 1, 2-bis(2-aminophenoxy)-ethane-N,N,N', N'-tetra-acetic acid (BAPTA) acetomethyl ester (AM). BAPTA had no influence on MC viability but reversed thapsigargin-induced apoptosis to control levels. After thapsigargin treatment (100 nmol/L, 12 hours), apoptotic MCs had a significantly higher [Ca(2+)](i) of 251 +/- 25 nmol/L (N = 41) as compared with MCs that were not or not yet apoptotic ([Ca(2+)](i) of 116 +/- 20 nmol/L, N = 26, P < 0,05). Platelet-derived growth factor (PDGF), a well-characterized growth factor for MCs, reversed the effects of thapsigargin on proliferation and apoptosis in a similar fashion as BAPTA. PDGF acutely stimulated increases of [Ca2+]i but abolished thapsigargin-dependent, but not angiotensin II- or ATP-induced Ca(2+) rises when administered during a 12-hour preincubation.
  
  
  CONCLUSIONS
  Our data suggest that a sustained increase of [Ca(2+)](i) may serve as a signal to trigger MC apoptosis. Growth factors such as PDGF can abolish apoptosis induced by elevations of [Ca(2+)](i) by altering intracellular Ca(2+) signaling.
authors.affiliation: Department of Medicine, University of Münster, Münster, Germany.
authors.forename: H
authors.initials: H
authors.lastname: Saleh
authors.forename: E
authors.initials: E
authors.lastname: Schlatter
authors.forename: D
authors.initials: D
authors.lastname: Lang
authors.forename: H G
authors.initials: HG
authors.lastname: Pauels
authors.forename: S
authors.initials: S
authors.lastname: Heidenreich
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002021:Buffers; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology*; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002614:Chelating Agents / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005920:Glomerular Mesangium / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D007425:Intracellular Membranes / Q000378:metabolism / Q000502:physiology*; D008297:Male; D009994:Osmolar Concentration; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D019284:Thapsigargin / Q000494:pharmacology
chemical_list: D002021:Buffers; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D010982:Platelet-Derived Growth Factor; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D019284:Thapsigargin; D002118:Calcium