title: CalDAG-GEFI deficiency protects mice in a novel model of Fcγ RIIA-mediated thrombosis and thrombocytopenia.
abstract: Platelet activation via Fcγ receptor IIA (FcγRIIA) is a critical event in immune-mediated thrombocytopenia and thrombosis syndromes (ITT). We recently identified signaling by the guanine nucleotide exchange factor CalDAG-GEFI and the adenosine diphosphate receptor P2Y12 as independent pathways leading to Rap1 small GTPase activation and platelet aggregation. Here, we evaluated the contribution of CalDAG-GEFI and P2Y12 signaling to platelet activation in ITT. Mice transgenic for the human FcγRIIA (hFcR) and deficient in CalDAG-GEFI(-/-) (hFcR/CDGI(-/-)) were generated. Compared with controls, aggregation of hFcR/CDGI(-/-) platelets or P2Y12 inhibitor-treated hFcR platelets required more than 5-fold and approximately 2-fold higher concentrations of a FcγRIIA stimulating antibody against CD9, respectively. Aggregation and Rap1 activation were abolished in P2Y12 inhibitor-treated hFcR/CDGI(-/-) platelets. For in vivo studies, a novel model for antibody-induced thrombocytopenia and thrombosis was established. FcγRIIA-dependent platelet thrombosis was induced by infusion of Alexa750-labeled antibodies to glycoprotein IX (CD42a), and pulmonary thrombi were detected by near-infrared imaging technology. Anti-GPIX antibodies dose-dependently caused thrombocytopenia and pulmonary thrombosis in hFcR-transgenic but not wild-type mice. CalDAG-GEFI-deficient but not clopidogrel-treated hFcR-transgenic mice were completely protected from ITT. In summary, we established a novel mouse model for ITT, which was used to identify CalDAG-GEFI as a potential new target in the treatment of ITT.
authors.affiliation: Cardeza Foundation and Department of Medicine, Thomas Jefferson University, Philadelphia, PA, USA.
authors.forename: Moritz
authors.initials: M
authors.lastname: Stolla
authors.forename: Lucia
authors.initials: L
authors.lastname: Stefanini
authors.forename: Pierrette
authors.initials: P
authors.lastname: André
authors.forename: Timothy D
authors.initials: TD
authors.lastname: Ouellette
authors.forename: Michael P
authors.initials: MP
authors.lastname: Reilly
authors.forename: Steven E
authors.initials: SE
authors.lastname: McKenzie
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Bergmeier
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002469:Cell Separation; D004195:Disease Models, Animal; D005434:Flow Cytometry; D020662:Guanine Nucleotide Exchange Factors / Q000172:deficiency*; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D008822:Mice, Transgenic; D015539:Platelet Activation / Q000502:physiology*; D010974:Platelet Aggregation / Q000502:physiology; D016553:Purpura, Thrombocytopenic, Idiopathic / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D017452:Receptors, IgG / Q000235:genetics / Q000276:immunology; D058925:Receptors, Purinergic P2Y12 / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D013927:Thrombosis / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: C080568:Fc gamma receptor IIA; D020662:Guanine Nucleotide Exchange Factors; C548577:P2ry12 protein, mouse; C485927:Rasgrp2 protein, mouse; D017452:Receptors, IgG; D058925:Receptors, Purinergic P2Y12