1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice

Oliver Borst, Patrick Münzer, Evi Schmid, Eva Maria Schmidt, Antonella Russo, Britta Walker, Wenting Yang, Christina Leibrock, Kalina Szteyn, Sebastian Schmidt, Margitta Elvers, Caterina Faggio, Ekaterina Shumilina, Makoto Kuro-O, Meinrad Gawaz, Florian Lang

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

Platelets are activated by increased cytosolic Ca2+ concentration ([Ca2+]i) following store-operated calcium entry (SOCE) accomplished by calciumrelease-activated calcium (CRAC) channel moiety Orai1 and its regulator STIM1. In other cells, Ca2+ 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 Ca2+ 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 [Ca2+]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-kB 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 Ca2+ signaling and activation, an effect at least partially due to 1,25(OH)2D3-dependent down-regulation of NF-kB activity and STIM1/Orai1 expression in megakaryocytes.

Original languageEnglish (US)
Pages (from-to)2108-2119
Number of pages12
JournalFASEB Journal
Volume28
Issue number5
DOIs
StatePublished - 2014

Fingerprint

Megakaryocytes
Cholecalciferol
Platelets
Thrombosis
Blood Platelets
Chemical activation
NF-kappa B
Calcium
Proteins
Flow cytometry
Fura-2
Confocal microscopy
Clamping devices
Calcium Channels
Nutrition
Confocal Microscopy
Integrins
Vitamin D
Transfection
Flow Cytometry

Keywords

  • Megakaryocytes
  • NF-kB
  • Orai1
  • SOCE
  • STIM1

ASJC Scopus subject areas

  • Biochemistry
  • Biotechnology
  • Genetics
  • Molecular Biology

Cite this

Borst, O., Münzer, P., Schmid, E., Schmidt, E. M., Russo, A., Walker, B., ... Lang, F. (2014). 1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice. FASEB Journal, 28(5), 2108-2119. https://doi.org/10.1096/fj.13-239277

1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice. / Borst, Oliver; Münzer, Patrick; Schmid, Evi; Schmidt, Eva Maria; Russo, Antonella; Walker, Britta; Yang, Wenting; Leibrock, Christina; Szteyn, Kalina; Schmidt, Sebastian; Elvers, Margitta; Faggio, Caterina; Shumilina, Ekaterina; Kuro-O, Makoto; Gawaz, Meinrad; Lang, Florian.

In: FASEB Journal, Vol. 28, No. 5, 2014, p. 2108-2119.

Research output: Contribution to journalArticle

Borst, O, Münzer, P, Schmid, E, Schmidt, EM, Russo, A, Walker, B, Yang, W, Leibrock, C, Szteyn, K, Schmidt, S, Elvers, M, Faggio, C, Shumilina, E, Kuro-O, M, Gawaz, M & Lang, F 2014, '1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice', FASEB Journal, vol. 28, no. 5, pp. 2108-2119. https://doi.org/10.1096/fj.13-239277
Borst, Oliver ; Münzer, Patrick ; Schmid, Evi ; Schmidt, Eva Maria ; Russo, Antonella ; Walker, Britta ; Yang, Wenting ; Leibrock, Christina ; Szteyn, Kalina ; Schmidt, Sebastian ; Elvers, Margitta ; Faggio, Caterina ; Shumilina, Ekaterina ; Kuro-O, Makoto ; Gawaz, Meinrad ; Lang, Florian. / 1,25(OH)2 vitamin D3-dependent inhibition of platelet Ca2+ signaling and thrombus formation in klotho-deficient mice. In: FASEB Journal. 2014 ; Vol. 28, No. 5. pp. 2108-2119.
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AU - Russo, Antonella

AU - Walker, Britta

AU - Yang, Wenting

AU - Leibrock, Christina

AU - Szteyn, Kalina

AU - Schmidt, Sebastian

AU - Elvers, Margitta

AU - Faggio, Caterina

AU - Shumilina, Ekaterina

AU - Kuro-O, Makoto

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AU - Lang, Florian

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N2 - Platelets are activated by increased cytosolic Ca2+ concentration ([Ca2+]i) following store-operated calcium entry (SOCE) accomplished by calciumrelease-activated calcium (CRAC) channel moiety Orai1 and its regulator STIM1. In other cells, Ca2+ 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 Ca2+ 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 [Ca2+]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-kB 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 Ca2+ signaling and activation, an effect at least partially due to 1,25(OH)2D3-dependent down-regulation of NF-kB activity and STIM1/Orai1 expression in megakaryocytes.

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