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BIN2 orchestrates platelet calcium signaling in thrombosis and thrombo-inflammation
Julia Volz, … , Markus Sauer, Bernhard Nieswandt
Julia Volz, … , Markus Sauer, Bernhard Nieswandt
Published August 4, 2020
Citation Information: J Clin Invest. 2020;130(11):6064-6079. https://doi.org/10.1172/JCI136457.
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Research Article Cell biology Hematology

BIN2 orchestrates platelet calcium signaling in thrombosis and thrombo-inflammation

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Abstract

Store-operated Ca2+ entry (SOCE) is the major route of Ca2+ influx in platelets. The Ca2+ sensor stromal interaction molecule 1 (STIM1) triggers SOCE by forming punctate structures with the Ca2+ channel Orai1 and the inositol trisphosphate receptor (IP3R), thereby linking the endo-/sarcoplasmic reticulum to the plasma membrane. Here, we identified the BAR domain superfamily member bridging integrator 2 (BIN2) as an interaction partner of STIM1 and IP3R in platelets. Deletion of platelet BIN2 (Bin2fl/fl,Pf4-Cre mice) resulted in reduced Ca2+ store release and Ca2+ influx in response to all tested platelet agonists. These defects were a consequence of impaired IP3R function in combination with defective STIM1-mediated SOC channel activation, while Ca2+ store content and agonist-induced IP3 production were unaltered. This severely defective Ca2+ signaling translated into impaired thrombus formation under flow and a protection of Bin2fl/fl,Pf4-Cre mice in models of arterial thrombosis and stroke. Our results establish BIN2 as a central regulator of platelet activation in thrombosis and thrombo-inflammatory disease settings.

Authors

Julia Volz, Charly Kusch, Sarah Beck, Michael Popp, Timo Vögtle, Mara Meub, Inga Scheller, Hannah S. Heil, Julia Preu, Michael K. Schuhmann, Katherina Hemmen, Thomas Premsler, Albert Sickmann, Katrin G. Heinze, David Stegner, Guido Stoll, Attila Braun, Markus Sauer, Bernhard Nieswandt

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Figure 6

Impaired hemostasis, defective arterial thrombus formation, and protection from ischemic brain infarction in Bin2fl/fl,Pf4Cre mice.

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Impaired hemostasis, defective arterial thrombus formation, and protecti...
(A) Adhesion and thrombus formation of platelets on collagen was assessed in a flow adhesion assay at a wall shear rate of 1700/s. Representative images and the quantification of the surface coverage and the relative thrombus volume are shown (n = 4). Representative example of 3 independent experiments. Values are depicted as mean ± SD, and P values were calculated using the Mann-Whitney U test. *P < 0.05. Scale bar: 50 μm. (B) Tail bleeding times of WT and Bin2fl/fl,Pf4Cre mice with or without acetylsalicylic acid (ASA) treatment (1 mg/kg i.v.) 15 minutes before the start of the experiment. Each symbol represents 1 animal. Kruskal-Wallis test (P value: 0.0022) followed by Dunn’s multiple-comparisons post hoc test; *P < 0.05, **P < 0.01. (C and D) Time to stable vessel occlusion of WT and Bin2fl/fl,Pf4Cre mice without (C) or with (D) ASA treatment (1 mg/kg i.v.) 15 minutes before the start of the experiment. The abdominal aorta was injured by firm compression with forceps, and blood flow was monitored for 30 minutes. Mann-Whitney U test was performed to compare mean occlusion times of occluded vessels. To compare frequency of occluded and nonoccluded vessels, Fisher’s exact test was used. *P < 0.05, ***P < 0.001. (E–I) WT and Bin2fl/fl Pf4-Cre mice were subjected to 60 minutes of transient middle cerebral artery occlusion (tMCAO) n = 10. (E) Representative images of 3 coronal brain sections stained with TTC. (F) Infarct volumes, (G) number of occluded ipsilateral vessels (2 slides per animal analyzed), (H) number of neutrophils, and (I) CD11b+ cells per section (2 slides per animal analyzed) were investigated after 24 hours. Values are depicted as mean ± SD, and P values were calculated using the Mann-Whitney U test. **P < 0.01, ***P < 0.001.

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