Abstract Number: PB1669
Meeting: ISTH 2020 Congress
Background: Platelets are increasingly recognized as important players of inflammation, interacting with immune and endothelial cells. Ca2+ is an essential second messenger in virtually all cells, regulating a wide range of fundamental cellular processes. In platelets, the elevation in Ca2+ contributes to various steps of cellular activation, such as reorganization of the actin cytoskeleton necessary for shape change, degranulation or platelet aggregation. So far, measuring Ca2+ influx in platelets is mainly performed using fluorescent Ca2+ indicators like Fura-2. However, this approach is limited to in vitro studies.
Aims: We aim to overcome this limitation by generating a new mouse model that allows monitoring rises in intracellular Ca2+ levels in platelets in vivo.
Methods: We generated mice that express the fluorescent calcium indicator protein GCaMP5G, along with tdTomato (marks reporter-positive cells) exclusively in megakaryocytes and platelets using the Pf4-Cre system. The expression of the calcium sensor was investigated by fluorescence microscopy. Platelet function was assessed by in vitro assays, such as flow cytometry, aggregometry, spreading and flow adhesion assays. Intravital imaging was performed to assess the feasibility of the sensor in living animals.
Results: Expression of the sensor in platelets was confirmed and neither Ca2+ influx, nor basic platelet functions like platelet activation and aggregation were altered. Using a flow adhesion system to assess thrombus formation on a collagen-coated surface, we were able to visualize Ca2+ fluxes in platelets in a growing thrombus ex vivo. More importantly, calcium signals in platelets could be visualized in vivo, using different models of arterial thrombosis and thrombo-inflammation.
Conclusions: The presented mouse model allows imaging of Ca2+ rises in platelets in vivo. This provides a valuable tool not only to study platelet signaling in the context of thrombus formation but provides the possibility to visualize platelet responses upon interaction with other cells in the context of (patho-)physiological processes.
To cite this abstract in AMA style:Klaus V, Gross C, Beck S, Stegner D. A Novel Reporter Mouse Line Enables in vivo Imaging of Intracellular Calcium Fluxes in Platelets [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/a-novel-reporter-mouse-line-enables-in-vivo-imaging-of-intracellular-calcium-fluxes-in-platelets/. Accessed January 26, 2022.
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