title: Upregulation of store operated Ca channel Orai1, stimulation of Ca(2+) entry and triggering of cell membrane scrambling in platelets by mineralocorticoid DOCA.
abstract: OBJECTIVE
  Mineralocorticoid excess leads to vascular injury, which is partially due to hypertension but in addition involves increased concentration of cytosolic Ca(2+) concentration in platelets, key players in the pathophysiology of occlusive vascular disease. Mineralocorticoids are in part effective by rapid nongenomic mechanisms including phosphatidylinositide-3-kinase (PI3K) signaling, which involves activation of the serum & glucocorticoid inducible kinase (SGK) isoforms. SGK1 has in turn been shown to participate in the regulation of the pore forming Ca(2+) channel protein Orai1 in platelets. Orai1 accomplishes entry of Ca(2+), which is in turn known to trigger cell membrane scrambling. Platelets lack nuclei but are able to express protein by translation, which is stimulated by PI3K signaling. The present study explored whether the mineralocorticoid desoxycorticosterone acetate (DOCA) influences platelet Orai1 protein abundance, cytosolic Ca(2+)-activity ([Ca(2+)]i), phosphatidylserine abundance at the cell surface and/or cell volume.
  
  
  METHODS
  Orai1 protein abundance was estimated utilizing CF™488A conjugated antibodies, [Ca(2+)]i utilizing Fluo3-fluorescence, phosphatidylserine abundance utilizing FITC-labelled annexin V, and cell volume utilizing forward scatter in flow cytometry.
  
  
  RESULTS
  DOCA (10 µg/ml) treatment of murine platelets was followed by a significant increase of Orai1 protein abundance, [Ca(2+)]i, percentage of phosphatidylserine exposing platelets and platelet swelling. The effect on [Ca(2+)]i, phosphatidylserine abundance and cell volume were completely abrogated by addition of the specific SGK inhibitor EMD638683 (50 µM) CONCLUSIONS: The mineralocorticoid DOCA upregulates Orai1 protein abundance in the cell membrane, thus increasing [Ca(2+)]i and triggering phosphatidylserine abundance, effects paralleled by platelet swelling.
authors: Department of Physiology, University of Tübingen, Tübingen, Germany.
authors: Guoxing
authors: G
authors: Liu
authors: Guilai
authors: G
authors: Liu
authors: Kousi
authors: K
authors: Alzoubi
authors: Anja T
authors: AT
authors: Umbach
authors: Lisann
authors: L
authors: Pelzl
authors: Oliver
authors: O
authors: Borst
authors: Meinrad
authors: M
authors: Gawaz
authors: Florian
authors: F
authors: Lang
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000096:biosynthesis* / Q000235:genetics; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D048429:Cell Size / Q000187:drug effects; D064791:Desoxycorticosterone Acetate / Q000494:pharmacology*; D005260:Female; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D008901:Mineralocorticoids / Q000494:pharmacology*; D000071740:ORAI1 Protein; D010718:Phosphatidylserines / Q000494:pharmacology; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D015220:Calcium Channels; D008901:Mineralocorticoids; D000071740:ORAI1 Protein; C520430:Orai1 protein, mouse; D010718:Phosphatidylserines; D064791:Desoxycorticosterone Acetate; D002118:Calcium