Platelet aggregation of <i>quinonoid</i> dihydropteridine reductase knockout mice.

AbstractCadherins play a major role in mediating cell–cell adhesion, which shares many parallels with platelet–platelet interactions during aggregate formation and clot stabilization. Platelets express epithelial (E)-cadherin, but its contribution to platelet function and/or platelet production is currently unknown. To assess the role of E-cadherin in platelet production and function in vitro and in vivo, we utilized a megakaryocyte-specific E-cadherin knockout mouse model. Loss of E-cadherin in megakaryocytes does not affect megakaryocyte maturation, platelet number or size. However, platelet dysfunction in the absence of E-cadherin is revealed when conditional knockout mice are challenged with acute antibody-mediated platelet depletion. Unlike wild-type mice that recover fully, knockout mice die within 72 hours post-antibody administration, likely from haemorrhage. Furthermore, conditional knockout mice have prolonged tail bleeding times, unstable clot formation, reduced clot retraction and reduced fibrin deposition in in vivo injury models. Murine platelet aggregation in vitro in response to thrombin and thrombin receptor activating peptide is compromised in E-cadherin null platelets, while aggregation in response to adenosine diphosphate (ADP) is not significantly different. Consistent with this, in vitro aggregation of primary human platelets in response to thrombin is decreased by an inhibitory E-cadherin antibody. Integrin activation and granule secretion in response to ADP and thrombin are not affected in E-cadherin null platelets, but Akt and glycogen synthase kinase 3β (GSK3β) activation are attenuated, suggesting a that E-cadherin contributes to aggregation, clot stabilization and retraction that is mediated by phosphoinositide 3-kinase/Akt/GSK3β signalling. In summary, E-cadherin plays a salient role in platelet aggregation and clot stability.

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Sp1/Sp3 transcription factors regulate hallmarks of megakaryocyte maturation and platelet formation and function

Blood ◽

10.1182/blood-2014-08-593343 ◽

2015 ◽

Vol 125 (12) ◽

pp. 1957-1967 ◽

Cited By ~ 38

Author(s):

Marjolein Meinders ◽

Divine I. Kulu ◽

Harmen J. G. van de Werken ◽

Mark Hoogenboezem ◽

Hans Janssen ◽

...

Keyword(s):

Transcription Factors ◽

Platelet Aggregation ◽

Knockout Mice ◽

Platelet Dysfunction ◽

Double Knockout ◽

Key Points ◽

Proplatelet Formation ◽

And Function ◽

Platelet Formation

Key Points Megakaryocyte-specific Sp1/Sp3 double-knockout mice display thrombocytopenia, platelet dysfunction, and defects in megakaryocyte maturation. Selective Mylk inhibition by ML7 affects proplatelet formation and stabilization and ITAM receptor–mediated platelet aggregation.

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CalDAG-GEFI Is a Key Signal Integrator in Platelet Activation and Thrombus Formation.

Blood ◽

10.1182/blood.v104.11.326.326 ◽

2004 ◽

Vol 104 (11) ◽

pp. 326-326

Author(s):

Wolfgang Bergmeier ◽

Jill R. Crittenden ◽

Crystal L. Piffath ◽

Denisa D. Wagner ◽

David E. Housman ◽

...

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Knockout Mice ◽

Thrombus Formation ◽

Es Cells ◽

Embryonic Stem ◽

Premature Stop Codon ◽

Small Gtpase ◽

Integrin Αiibβ3

Abstract Inside-out activation of platelet integrin αIIbβ3 is a key step in agonist-induced platelet aggregation. Recent studies suggested the involvement of the small GTPase Rap1b in this process as it is highly expressed in platelets and becomes activated during platelet activation. In cell lines, overexpression of the Rap activator CalDAG-GEFI increased αIIbβ3-dependent adhesion, while overexpression of RapGAP, which inactivates Rap1, reduced αIIbβ3 activity. Here we provide evidence that CalDAG-GEFI is an essential component of this pathway in vivo. To generate CalDAG-GEFI knockout mice, we engineered mouse embryonic stem (ES) cells with a deletion that results in a frameshift mutation and a premature stop codon at the position encoding the 37th amino acid of CalDAG-GEFI. These ES cells were then used to derive chimeric mice that yielded germline transmission of the CalDAG-GEFI mutation. Deficiency of CalDAG-GEFI in mutant mice was confirmed by immunohistochemistry and western blot analysis. CalDAG-GEFI−/− platelets showed impaired Rap1b activation and aggregation in response to various agonists, with aggregation being completely blocked when platelets were activated with ADP, thromboxaneA2 analog, or calcium ionophore. Under physiological flow conditions in vitro and in vivo, CalDAG-GEFI-deficient platelets showed normal tethering to basement membrane components but failed to form thrombi. Mice deficient in CalDAG-GEFI were further characterized by a greatly increased bleeding time as well as by a strong protection against collagen-induced pulmonary thrombosis. In summary, we identified CalDAG-GEFI as a key signal integrator in the cascade leading through Rap1 and integrin αIIbβ3 to platelet aggregation and thrombus formation. The fact that CalDAG-GEFI knockout mice are resistant to collagen-induced thrombosis, and do not undergo spontaneous hemorrhaging, suggests that CalDAG-GEFI may be a promising new target for antithrombotic therapy.

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C57BL/6JOlaHsd Mice with Tandem Deletion of the Multimerin 1 and Alpha-Synuclein Genes Have Impaired Platelet Function in Vivo and in Vitro That Can Be Corrected by Multimerin 1

Blood ◽

10.1182/blood.v112.11.3926.3926 ◽

2008 ◽

Vol 112 (11) ◽

pp. 3926-3926 ◽

Cited By ~ 1

Author(s):

Subia Tasneem ◽

Adili Reheman ◽

Heyu Ni ◽

Catherine P.M. Hayward

Keyword(s):

Platelet Aggregation ◽

Platelet Function ◽

Knockout Mice ◽

Platelet Adhesion ◽

Thrombus Formation ◽

Alpha Synuclein ◽

Type I ◽

Significant Difference

Abstract Studies of mice with genetic deficiencies have provided important insights on the functions of many proteins in thrombosis and hemostasis. Recently, a strain of mice (C57BL/6JOlaHsd, an inbred strain of C57BL/6J) has been identified to have a spontaneous, tandem deletion of the multimerin 1 and α-synuclein genes, which are also adjacent genes on human chromosome 4q22. Multimerin 1 is an adhesive protein found in platelets and endothelial cells while α-synuclein is a protein found in the brain and in blood that is implicated in neurodegenerative diseases and exocytosis. In vitro, multimerin 1 supports platelet adhesion while α-synuclein inhibits α-granule release. We postulated that the loss of multimerin 1 and α-synuclein would alter platelet function and that recombinant human multimerin 1 might correct some of these abnormalities. We compared platelet adhesion, aggregation and thrombus formation in vitro and in vivo in C57BL/6JOlaHsd and C57BL/6 mice. Thrombus formation was studied by using the ferric-chloride injured mesenteric arteriole thrombosis model under intravital microscopy. We found that platelet adhesion, aggregation and thrombus formation in C57BL/6JOlaHsd were significantly impaired in comparison to control, C57BL/6 mice. The number of single platelets, deposited 3–5 minutes after injury, was significantly decreased in C57BL/6JOlaHsd mice (P <0.05, platelets/min: C57BL/6 = 157 ± 15, n=16; C57BL/6JOlaHsd = 77 ± 13, n=17). Moreover, thrombus formation in these mice was significantly delayed. Thrombi in C57BL/6JOlaHsd were unstable and easily dissolved, which resulted in significant delays (P<0.001) in vessel occlusion (mean occlusion times: C57BL/6 = 15.6 ± 1.2 min, n=16; C57BL/6JOlaHsd = 31.9 ± 2.1 min, n=17). We further tested platelet function in these mice by ADP and thrombin induced platelet aggregation using platelet rich plasma and gel-filtered platelets, respectively. Although no significant differences were seen with ADP aggregation, thrombin-induced platelet aggregation was significantly impaired in C57BL/6JOlaHsd mice. Platelet adhesion to type I collagen (evaluated using microcapillary chambers, perfused at 1500 s−1 with whole blood) was also impaired in C57BL/6JOlaHsd mice. However, platelets from C57BL/6JOlaHsd mice showed a normal pattern of agonist-induced release of α-granule P-selectin. Multimerin 1 corrected the in vitro aggregation and adhesion defects of C57BL/6JOlaHsd platelets. Furthermore, the transfusion of multimerin 1 into C57BL/6JOlaHsd mice corrected the impaired platelet deposition and thrombus formation in vivo. No significant difference was found in tail bleeding time between the two groups of mice. As α-synuclein knockout mice have a shortened time to thrombus formation (Circulation2007;116:II_76), the effects of multimerin 1 on impaired platelet function in C57BL/6JOlaHsd mice provide supportive evidence that multimerin 1 contributes to platelet adhesion and thrombus formation at the site of vessel injury. The findings suggest multimerin 1 knockout mice will be useful to explore platelet function. The first two authors and participating laboratories contributed equally to this study.

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P2 receptors, platelet function and pharmacological implications

Thrombosis and Haemostasis ◽

10.1160/th07-11-0673 ◽

2008 ◽

Vol 99 (03) ◽

pp. 466-472 ◽

Cited By ~ 186

Author(s):

Christian Gachet

Keyword(s):

Platelet Aggregation ◽

G Protein ◽

Platelet Activation ◽

Knockout Mice ◽

P2 Receptors ◽

The Other ◽

Antithrombotic Drugs ◽

Adp Receptors ◽

Inflammatory Processes ◽

G Protein Coupled

SummaryADP and ATP play a crucial role in platelet activation and their receptors are potential targets for antithrombotic drugs. The ATP-gated cation channel P2X1 and the two G protein-coupled ADP receptors, P2Y1 and P2Y12, selectively contribute to platelet aggregation and formation of a thrombus.Owing to its central role in the growth and stabilization of a thrombus, the P2Y12 receptor is an established target of antithrombotic drugs like the thienopyridines clopidogrel or prasugrel, or competitive antag-onists such as cangrelor or AZD6140.The optimal inhibition of this receptor to reach clinical efficacy while preserving patients from unacceptable bleeding is a matter of debate. On the other hand, studies in P2Y1 and P2X1 knockout mice and using selective P2Y1 and P2X1 antagonists have shown that these receptors are also attractive targets for new antithrombotic compounds. Finally, the regulation by the P2 receptors of the platelet involvement in inflammatory processes is also briefly discussed.

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Effect of methotrexate on pteridine-contents and relaxation response of the aorta from quinonoid-dihydropteridine reductase (QDPR) knockout mice

Proceedings for Annual Meeting of The Japanese Pharmacological Society ◽

10.1254/jpssuppl.wcp2018.0_po3-3-47 ◽

2018 ◽

Vol WCP2018 (0) ◽

pp. PO3-3-47

Author(s):

Chiho Sumi-Ichinose ◽

Yui Suganuma ◽

Taiki Kano ◽

Noriko Ihira ◽

Kazuhisa Ikemoto ◽

...

Keyword(s):

Knockout Mice ◽

Relaxation Response ◽

Dihydropteridine Reductase

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A new mouse model mimicking thrombotic thrombocytopenic purpura: correction of symptoms by recombinant human ADAMTS13

Blood ◽

10.1182/blood-2011-09-380535 ◽

2012 ◽

Vol 119 (25) ◽

pp. 6128-6135 ◽

Cited By ~ 71

Author(s):

Alexandra Schiviz ◽

Kuno Wuersch ◽

Christina Piskernik ◽

Barbara Dietrich ◽

Werner Hoellriegl ◽

...

Keyword(s):

Platelet Aggregation ◽

Mouse Model ◽

Thrombotic Thrombocytopenic Purpura ◽

Thrombotic Microangiopathy ◽

Knockout Mice ◽

Serum Lactate Dehydrogenase ◽

Severe Thrombocytopenia ◽

Dependent Manner ◽

Thrombocytopenic Purpura ◽

Induce Platelet Aggregation

Abstract Deficiency of a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13), a VWF-cleaving protease, is the key factor in the pathogenesis of thrombotic thrombocytopenic purpura (TTP), a life-threatening thrombotic microangiopathy. It is well established that ADAMTS13 deficiency results in elevated plasma levels of ultra-large VWF multimers (ULVWF), which are prone to induce platelet aggregation; however, the actual trigger of TTP development remains uncertain. Here we describe a new animal model in which some TTP-like symptoms can be triggered in ADAMTS13 knockout mice by challenge with 2000 units/kg body weight of recombinant human VWF containing ULVWF multimers. Animals rapidly showed clinical symptoms and developed severe thrombocytopenia. Schistocytosis, a decrease in hematocrit, and elevated serum lactate dehydrogenase levels were observed. The heart was identified as the most sensitive target organ with rapid onset of extensive platelet aggregation in the ventricles and myocardial necrosis. Prophylactic administration of 200 units/kg recombinant human ADAMTS13 protected ADAMTS13 knockout mice from developing TTP. Therapeutic administration of 320 units/kg rhADAMTS13 reduced the incidence and severity of TTP findings in a treatment interval-dependent manner. We therefore consider this newly established mouse model of thrombotic microangiopathy highly predictive for investigating the efficacy of new treatments for TTP.

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5-hydroxytryptophan supplement recovers impaired platelet aggregation in Quinonoid dihydropteridine reductase deficient mice

Proceedings for Annual Meeting of The Japanese Pharmacological Society ◽

10.1254/jpssuppl.94.0_3-o-f4-1 ◽

2021 ◽

Vol 94 (0) ◽

pp. 3-O-F4-1

Author(s):

Yui Suganuma ◽

Taiki Kano ◽

Kazuhisa Ikemoto ◽

Chiho Sumi-Ichinose ◽

Hiroshi Ichinose ◽

...

Keyword(s):

Platelet Aggregation ◽

Dihydropteridine Reductase ◽

Deficient Mice

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Genetic strain differences in platelet aggregation of laboratory mice

Thrombosis and Haemostasis ◽

10.1160/th05-05-0322 ◽

2006 ◽

Vol 95 (01) ◽

pp. 159-165 ◽

Cited By ~ 11

Author(s):

Hideki Ito ◽

Yukio Kimura ◽

Toshiki Sudo

Keyword(s):

Platelet Aggregation ◽

Knockout Mice ◽

Strain Difference ◽

Inbred Mouse ◽

Laboratory Mouse ◽

Physiological Role ◽

Strain Differences ◽

Mouse Strains ◽

Laboratory Mice

SummaryTo investigate the physiological role of novel genes and proteins in platelet activation, various knockout mice have been produced. A number of standard inbred mouse strains each possessing genetically unique characters such as high tumor generation, hyperglycemia or hyperlipidemia, have been bred. In breeding knockout mice for investigation of specific physiological functions, appropriate selection of parental or backcross strains is necessary. Thus, examination of strain-specific platelet characteristics is important. In the present study, platelet aggregation responses of 13 laboratory mouse strains, 129/Sv, A, AKR, BALB/c, C3H/He, C57BL/6J, CBA, DBA/1, DBA/2, ddY, FVB, ICR, and NZW, and the diabetic strain C57BL/KsJ db/db, were compared. Marked strain differences were observed inADP- and collagen-induced platelet aggregation. The highest responses with both were seen in AKR/J and NZW/N, whereas the lowest were seen in DBA/2 and DBA/1.There was a 5-fold difference in the platelet aggregation threshold index (PATI) for ADP-induced PRP aggregation between AKR/J (0.6 µM) and DBA/2 (3.0 µM). With whole blood aggregation, the highest response was seen in AKR, whereas the lowest was seen in DBA/2 and DBA/1. The present study demonstrated that there is considerable strain difference in platelet aggregation among laboratory mice, which should be taken into account in backcrossing knockout strains.

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Fine Structure of Platelet Interaction with a New Microcrystalline Collagen Preparation

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100050305 ◽

1974 ◽

Vol 32 ◽

pp. 58-59

Author(s):

W. H. Zucker ◽

R. G. Mason

Keyword(s):

Fine Structure ◽

Platelet Aggregation ◽

Hydrochloric Acid ◽

Whole Blood ◽

Platelet Adhesion ◽

Hemostatic Agent ◽

Native Collagen ◽

Fibrillar Morphology ◽

Clinical Interest ◽

New Form

Platelet adhesion initiates platelet aggregation and is an important component of the hemostatic process. Since the development of a new form of collagen as a topical hemostatic agent is of both basic and clinical interest, an ultrastructural and hematologic study of the interaction of platelets with the microcrystalline collagen preparation was undertaken.In this study, whole blood anticoagulated with EDTA was used in order to inhibit aggregation and permit study of platelet adhesion to collagen as an isolated event. The microcrystalline collagen was prepared from bovine dermal corium; milling was with sharp blades. The preparation consists of partial hydrochloric acid amine collagen salts and retains much of the fibrillar morphology of native collagen.

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