Mice lacking the signaling molecule CalDAG-GEFI represent a model for leukocyte adhesion deficiency type III. J Clin Invest

CBR Institute for Biomedical Research and Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Journal of Clinical Investigation (Impact Factor: 13.22). 07/2007; 117(6):1699-707. DOI: 10.1172/JCI30575
Source: PubMed

ABSTRACT Single gene mutations in beta integrins can account for functional defects of individual cells of the hematopoietic system. In humans, mutations in beta(2) integrin lead to leukocyte adhesion deficiency (LAD) syndrome and mutations in beta(3) integrin cause the bleeding disorder Glanzmann thrombasthenia. However, multiple defects in blood cells involving various beta integrins (beta(1), beta(2), and beta(3)) occur simultaneously in patients with the recently described LAD type III (LAD-III). Here we show that the product of a single gene, Ca(2+) and diacylglycerol-regulated guanine nucleotide exchange factor I (CalDAG-GEFI), controlled the activation of all 3 integrins in the hematopoietic system. Neutrophils from CalDAG-GEFI(-/-) mice exhibited strong defects in Rap1 and beta(1) and beta(2) integrin activation while maintaining normal calcium flux, degranulation, and ROS generation. Neutrophils from CalDAG-GEFI-deficient mice failed to adhere firmly to stimulated venules and to migrate into sites of inflammation. Furthermore, CalDAG-GEFI regulated the activation of beta(1) and beta(3) integrins in platelets, and CalDAG-GEFI deficiency caused complete inhibition of arterial thrombus formation in mice. Thus, mice engineered to lack CalDAG-GEFI have a combination of defects in leukocyte and platelet functions similar to that of LAD-III patients.

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Available from: Ann M Graybiel, Sep 29, 2015
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    • "The injection of fluoxetine also more than doubled the number of firmly adhered leukocytes. In comparison to earlier studies without intraperitoneal injection, more leukocyte adhesion was observed, even without fluoxetine [10], [23], [24]. Most likely, this difference can be explained by endothelial stimulation induced by mechanical irritation provoked by the process of injecting a liquid into the abdominal cavity that cannot be avoided. "
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    ABSTRACT: Activated platelets release serotonin at sites of inflammation where it acts as inflammatory mediator and enhances recruitment of neutrophils. Chronic treatment with selective serotonin reuptake inhibitors (SSRI) depletes the serotonin storage pool in platelets, leading to reduced leukocyte recruitment in murine experiments. Here, we examined the direct and acute effects of SSRI on leukocyte recruitment in murine peritonitis. C57Bl/6 and Tph1-/- (Tryptophan hydroxylase1) mice underwent acute treatment with the SSRI fluoxetine or vehicle. Serotonin concentrations were measured by ELISA. Leukocyte rolling and adhesion on endothelium was analyzed by intravital microscopy in mesentery venules with and without lipopolysaccharide challenge. Leukocyte extravasation in sterile peritonitis was measured by flow cytometry of abdominal lavage fluid. Plasma serotonin levels were elevated 2 hours after fluoxetine treatment (0.70±0.1 µg/ml versus 0.27±0.1, p = 0.03, n = 14), while serum serotonin did not change. Without further stimulation, acute fluoxetine treatment increased the number of rolling leukocytes (63±8 versus 165±17/0.04 mm(2)min(-1)) and decreased their velocity (61±6 versus 28±1 µm/s, both p<0.0001, n = 10). In Tph1-/- mice leukocyte rolling was not significantly influenced by acute fluoxetine treatment. Stimulation with lipopolysaccharide decreased rolling velocity and induced leukocyte adhesion, which was enhanced after fluoxetine pretreatment (27±3 versus 36±2/0.04 mm(2), p = 0.008, n = 10). Leukocyte extravasation in sterile peritonitis, however, was not affected by acute fluoxetine treatment. Acute fluoxetine treatment increased plasma serotonin concentrations and promoted leukocyte-endothelial interactions in-vivo, suggesting that serotonin is a promoter of acute inflammation. E-selectin was upregulated on endothelial cells in the presence of serotonin, possibly explaining the observed increase in leukocyte-endothelial interactions. However transmigration of neutrophils in sterile peritonitis was not affected by higher serotonin concentrations, indicating that the effect of fluoxetine was restricted to early steps in the leukocyte recruitment. Whether SSRI use in humans alters leukocyte recruitment remains to be investigated.
    PLoS ONE 02/2014; 9(2):e88316. DOI:10.1371/journal.pone.0088316 · 3.23 Impact Factor
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    • "The IP3R, designated as calciumselective cation channel, allows efflux of Ca2+ from the DTS, thus increasing the cytoplasmic Ca2+ level [80]. Increased Ca2+ level activates “Ca2+ and diacylglycerol regulated guanine nucleotide exchange factor I” (CalDAG-GEFI) [81,82], which in turn promotes the activation of Rap1b, a small GTP binding protein. Rap1b is critical for GpIIb/IIIa activation and platelet function [83] by regulating cytoskeletal rearrangements through interactions with Rap1-interacting adaptor molecule (RIAM) [84]. "
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    ABSTRACT: von Willebrand factor (vWF) is a multimeric glycoprotein essential for hemostasis after vascular injury, which modulates platelet-surface and platelet-platelet interactions by linking platelet receptors to the extracellular matrix and to each other. The crucial role of vWF in platelet function is particularly apparent when hemodynamic conditions create blood flow with high shear stress. Through multiple functional domains, vWF mediates the attachment of platelets to exposed tissues, where immobilized vWF is able to support a homotypic and/or heterotypic self-association. The self-association of vWF is also supported by a rapidly expanding reservoir of novel evidences that the thiol/disulfide exchange regulates vWF multimer size in the blood circulation. Moreover, in addition to proteolysis and reduction of ADAMTS13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13), the regulation of vWF multimer size and self-association may depend on a disulfide bond reductase activity ascribed to thrombospondin-1 (TSP-1). Along with the classical signaling pathways in activated platelets, evidence is emerging that lipid rafts also play important roles in various phases of hemostasis and thrombosis and facilitate the interaction between the key signaling molecules. Developments in these areas will refine our understanding of the role played by vWF self-association in physiological hemostasis and pathological thrombosis.
    Journal of Hematology & Oncology 10/2012; 5(1):65. DOI:10.1186/1756-8722-5-65 · 4.81 Impact Factor
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    • "Interestingly, the addition of ADP and TxA 2 increases the adhesion of CalDAG-GEFI knockout platelets to a collagen surface at low shear rates, without restoring thrombus growth [38]. In addition to the key role played in platelet adhesion to collagen, the expression of CalDAG-GEFI is also required for an efficient platelet interaction with other β 1 integrin ligands as laminin and fibronectin [52]. The observed differences between Rap1b and CalDAG- GEFI knockout mice, which are particularly evident in terms of phenotype severity, may be related to possible additional functions of CalDAG-GEFI, independent of Rap1b activation , that can be required for efficient integrin-mediated adhesion. "
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    ABSTRACT: Integrins and other families of cell adhesion receptors are responsible for platelet adhesion and aggregation, which are essential steps for physiological haemostasis, as well as for the development of thrombosis. The modulation of platelet adhesive properties is the result of a complex pattern of inside-out and outside-in signaling pathways, in which the members of the Rap family of small GTPases are bidirectionally involved. This paper focuses on the regulation of the main Rap GTPase expressed in circulating platelets, Rap1b, downstream of adhesion receptors, and summarizes the most recent achievements in the investigation of the function of this protein as regulator of platelet adhesion and thrombus formation.
    06/2012; 2012:412089. DOI:10.1155/2012/412089
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