University of Leicester
Evidence for shear-mediated Ca2+ entry through mechanosensitive cation channels in human platelets and a megakaryocytic cell line.pdf (2.82 MB)

Evidence for shear-mediated Ca(2+) entry through mechanosensitive cation channels in human platelets and a megakaryocytic cell line.

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journal contribution
posted on 2017-07-10, 14:36 authored by Zeki Ilkan, Joy R. Wright, Alison H. Goodall, Jonathan M. Gibbins, Chris I. Jones, Martyn P. Mahaut-Smith
The role of mechanosensitive (MS) Ca(2+)-permeable ion channels in platelets is unclear, despite the importance of shear stress in platelet function and life-threatening thrombus formation. We therefore sought to investigate the expression and functional relevance of MS channels in human platelets. The effect of shear stress on Ca(2+) entry in human platelets and Meg-01 megakaryocytic cells loaded with Fluo-3 was examined by confocal microscopy. Cells were attached to glass coverslips within flow chambers that allowed applications of physiological and pathological shear stress. Arterial shear (1002.6 s(-1)) induced a sustained increase in [Ca(2+)] i in Meg-01 cells and enhanced the frequency of repetitive Ca(2+) transients by 80% in platelets. These Ca(2+) increases were abrogated by the MS channel inhibitor Grammostola spatulata mechanotoxin 4 (GsMTx-4) or by chelation of extracellular Ca(2+) Thrombus formation was studied on collagen-coated surfaces using DiOC6-stained platelets. In addition, [Ca(2+)] i and functional responses of washed platelet suspensions were studied with Fura-2 and light transmission aggregometry, respectively. Thrombus size was reduced 50% by GsMTx-4, independently of P2X1 receptors. In contrast, GsMTx-4 had no effect on collagen-induced aggregation or on Ca(2+) influx via TRPC6 or Orai1 channels and caused only a minor inhibition of P2X1-dependent Ca(2+) entry. The Piezo1 agonist, Yoda1, potentiated shear-dependent platelet Ca(2+) transients by 170%. Piezo1 mRNA transcripts and protein were detected with quantitative RT-PCR and Western blotting, respectively, in both platelets and Meg-01 cells. We conclude that platelets and Meg-01 cells express the MS cation channel Piezo1, which may contribute to Ca(2+) entry and thrombus formation under arterial shear.



Journal of Biological Chemistry, 2017, 292 (22), pp. 9204-9217

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/Organisation/COLLEGE OF MEDICINE, BIOLOGICAL SCIENCES AND PSYCHOLOGY/MBSP Non-Medical Departments/Molecular & Cell Biology


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