title: 1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice.
abstract: Platelets are activated by increased cytosolic Ca(2+) concentration ([Ca(2+)]i) following store-operated calcium entry (SOCE) accomplished by calcium-release-activated calcium (CRAC) channel moiety Orai1 and its regulator STIM1. In other cells, Ca(2+) transport is regulated by 1,25(OH)2 vitamin D3 [1,25(OH)2D3]. 1,25(OH)2D3 formation is inhibited by klotho and excessive in klotho-deficient mice (kl/kl). The present study explored the effect of klotho deficiency on platelet Ca(2+) signaling and activation. Platelets and megakaryocytes isolated from WT and kl/kl-mice were analyzed by RT-PCR, Western blotting, confocal microscopy, Fura-2-fluorescence, patch clamp, flow cytometry, aggregometry, and flow chamber. STIM1/Orai1 transcript and protein levels, SOCE, agonist-induced [Ca(2+)]i increase, activation-dependent degranulation, integrin αIIbβ3 activation and aggregation, and thrombus formation were significantly blunted in kl/kl platelets (by 27-90%). STIM1/Orai1 transcript and protein levels, as well as CRAC currents, were significantly reduced in kl/kl megakaryocytes (by 38-73%) and 1,25(OH)2D3-treated WT megakaryocytes. Nuclear NF-κB subunit p50/p65 abundance was significantly reduced in kl/kl-megakaryocytes (by 51-76%). Transfection with p50/p65 significantly increased STIM1/Orai1 transcript and protein levels in megakaryocytic MEG-01 cells (by 46-97%). Low-vitamin D diet (LVD) of kl/kl mice normalized plasma 1,25(OH)2D3 concentration and function of platelets and megakaryocytes. Klotho deficiency inhibits platelet Ca(2+) signaling and activation, an effect at least partially due to 1,25(OH)2D3-dependent down-regulation of NF-κB activity and STIM1/Orai1 expression in megakaryocytes.
authors: 2Department of Physiology, University of Tübingen, Gmelinstr. 5, 72076 Tübingen, Germany. florian.lang@uni-tuebingen.de.
authors: Oliver
authors: O
authors: Borst
authors: Patrick
authors: P
authors: Münzer
authors: Evi
authors: E
authors: Schmid
authors: Eva-Maria
authors: EM
authors: Schmidt
authors: Antonella
authors: A
authors: Russo
authors: Britta
authors: B
authors: Walker
authors: Wenting
authors: W
authors: Yang
authors: Christina
authors: C
authors: Leibrock
authors: Kalina
authors: K
authors: Szteyn
authors: Sebastian
authors: S
authors: Schmidt
authors: Margitta
authors: M
authors: Elvers
authors: Caterina
authors: C
authors: Faggio
authors: Ekaterina
authors: E
authors: Shumilina
authors: Makoto
authors: M
authors: Kuro-o
authors: Meinrad
authors: M
authors: Gawaz
authors: Florian
authors: F
authors: Lang
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000378:metabolism*; D002117:Calcitriol / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D020013:Calcium Signaling; D002467:Cell Nucleus / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D015536:Down-Regulation; D005966:Glucuronidase / Q000235:genetics*; D000090265:Klotho Proteins; D008533:Megakaryocytes / Q000166:cytology; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D016328:NF-kappa B / Q000378:metabolism; D000071740:ORAI1 Protein; D018408:Patch-Clamp Techniques; D010974:Platelet Aggregation; D015398:Signal Transduction; D000071737:Stromal Interaction Molecule 1; D013927:Thrombosis / Q000378:metabolism*; D014162:Transfection
chemical_list: D015220:Calcium Channels; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D000071740:ORAI1 Protein; C520430:Orai1 protein, mouse; C100757:Stim1 protein, mouse; D000071737:Stromal Interaction Molecule 1; D005966:Glucuronidase; D000090265:Klotho Proteins; D002117:Calcitriol; D002118:Calcium
keywords: NF-κB; Orai1; SOCE; STIM1; megakaryocytes