TY  - JOUR
A1  - Mammadova-Bach, Elmina
A1  - Braun, Attila
T1  - Zinc homeostasis in platelet-related diseases
JF  - International Journal of Molecular Sciences
N2  - Zn\(^{2+}\) deficiency in the human population is frequent in underdeveloped countries. Worldwide, approximatively 2 billion people consume Zn\(^{2+}\)-deficient diets, accounting for 1–4% of deaths each year, mainly in infants with a compromised immune system. Depending on the severity of Zn\(^{2+}\) deficiency, clinical symptoms are associated with impaired wound healing, alopecia, diarrhea, poor growth, dysfunction of the immune and nervous system with congenital abnormalities and bleeding disorders. Poor nutritional Zn\(^{2+}\) status in patients with metastatic squamous cell carcinoma or with advanced non-Hodgkin lymphoma, was accompanied by cutaneous bleeding and platelet dysfunction. Forcing Zn\(^{2+}\) uptake in the gut using different nutritional supplementation of Zn\(^{2+}\) could ameliorate many of these pathological symptoms in humans. Feeding adult rodents with a low Zn\(^{2+}\) diet caused poor platelet aggregation and increased bleeding tendency, thereby attracting great scientific interest in investigating the role of Zn\(^{2+}\) in hemostasis. Storage protein metallothionein maintains or releases Zn\(^{2+}\) in the cytoplasm, and the dynamic change of this cytoplasmic Zn\(^{2+}\) pool is regulated by the redox status of the cell. An increase of labile Zn\(^{2+}\) pool can be toxic for the cells, and therefore cytoplasmic Zn\(^{2+}\) levels are tightly regulated by several Zn\(^{2+}\) transporters located on the cell surface and also on the intracellular membrane of Zn\(^{2+}\) storage organelles, such as secretory vesicles, endoplasmic reticulum or Golgi apparatus. Although Zn\(^{2+}\) is a critical cofactor for more than 2000 transcription factors and 300 enzymes, regulating cell differentiation, proliferation, and basic metabolic functions of the cells, the molecular mechanisms of Zn\(^{2+}\) transport and the physiological role of Zn\(^{2+}\) store in megakaryocyte and platelet function remain elusive. In this review, we summarize the contribution of extracellular or intracellular Zn\(^{2+}\) to megakaryocyte and platelet function and discuss the consequences of dysregulated Zn\(^{2+}\) homeostasis in platelet-related diseases by focusing on thrombosis, ischemic stroke and storage pool diseases.
KW  - Zinc
KW  - platelets
KW  - hemostasis
KW  - thrombosis
KW  - ischemic stroke
KW  - storage-pool diseases
Y1  - 2019
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-285554
SN  - 1422-0067
VL  - 20
IS  - 21
ER  - 
TY  - JOUR
A1  - Göb, Vanessa
A1  - Voll, Maximilian G.
A1  - Zimmermann, Lena
A1  - Hemmen, Katharina
A1  - Stoll, Guido
A1  - Nieswandt, Bernhard
A1  - Schuhmann, Michael K.
A1  - Heinze, Katrin G.
A1  - Stegner, David
T1  - Infarct growth precedes cerebral thrombosis following experimental stroke in mice
JF  - Scientific Reports
N2  - Ischemic stroke is among the leading causes of disability and death worldwide. In acute ischemic stroke, successful recanalization of occluded vessels is the primary therapeutic aim, but even if it is achieved, not all patients benefit. Although blockade of platelet aggregation did not prevent infarct progression, cerebral thrombosis as cause of secondary infarct growth has remained a matter of debate. As cerebral thrombi are frequently observed after experimental stroke, a thrombus-induced impairment of the brain microcirculation is considered to contribute to tissue damage. Here, we combine the model of transient middle cerebral artery occlusion (tMCAO) with light sheet fluorescence microscopy and immunohistochemistry of brain slices to investigate the kinetics of thrombus formation and infarct progression. Our data reveal that tissue damage already peaks after 8 h of reperfusion following 60 min MCAO, while cerebral thrombi are only observed at later time points. Thus, cerebral thrombosis is not causative for secondary infarct growth during ischemic stroke.
KW  - cerebrovascular disorders
KW  - thrombosis
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-265791
VL  - 11
IS  - 1
ER  - 
TY  - JOUR
A1  - Beck, Sarah
A1  - Stegner, David
A1  - Loroch, Stefan
A1  - Baig, Ayesha A.
A1  - Göb, Vanessa
A1  - Schumbutzki, Cornelia
A1  - Eilers, Eva
A1  - Sickmann, Albert
A1  - May, Frauke
A1  - Nolte, Marc W.
A1  - Panousis, Con
A1  - Nieswandt, Bernhard
T1  - Generation of a humanized FXII knock-in mouse-A powerful model system to test novel anti-thrombotic agents
JF  - Journal of Thrombosis and Haemostasis
N2  - Background
Effective inhibition of thrombosis without generating bleeding risks is a major challenge in medicine. Accumulating evidence suggests that this can be achieved by inhibition of coagulation factor XII (FXII), as either its knock-out or inhibition in animal models efficiently reduced thrombosis without affecting normal hemostasis. Based on these findings, highly specific inhibitors for human FXII(a) are under development. However, currently, in vivo studies on their efficacy and safety are impeded by the lack of an optimized animal model expressing the specific target, that is, human FXII.

Objective
The primary objective of this study is to develop and functionally characterize a humanized FXII mouse model.

Methods
A humanized FXII mouse model was generated by replacing the murine with the human F12 gene (genetic knock-in) and tested it in in vitro coagulation assays and in in vivo thrombosis models.

Results
These hF12\(^{KI}\) mice were indistinguishable from wild-type mice in all tested assays of coagulation and platelet function in vitro and in vivo, except for reduced expression levels of hFXII compared to human plasma. Targeting FXII by the anti-human FXIIa antibody 3F7 increased activated partial thromboplastin time dose-dependently and protected hF12\(^{KI}\) mice in an arterial thrombosis model without affecting bleeding times.

Conclusion
These data establish the newly generated hF12\(^{KI}\) mouse as a powerful and unique model system for in vivo studies on anti-FXII(a) biologics, supporting the development of efficient and safe human FXII(a) inhibitors.
KW  - hemostasis,
KW  - blood coagulation
KW  - factor XII
KW  - animal models
KW  - thrombosis
Y1  - 2021
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-259567
VL  - 19
IS  - 11
ER  - 
TY  - THES
A1  - Rabie, Tamer
T1  - Cellular regulation of platelet glycoprotein VI : in vivo and in vitro studies in mice
T1  - Zelluläre Regulation von Plättchen Glykoprotein VI : in vivo und in vitro Studien in der Maus
N2  - Platelet interaction with the subendothelium is essential to limit blood loss after tissue injury. However, upon rupture of atherosclerotic plaques, this interaction may result in blood vessel occlusion leading to life threatening diseases such as myocardial infarction or stroke. Among the subendothelial matrix proteins, collagen is considered to be the most thrombogenic component as it directly activates platelets. Platelets interact with collagen, either indirectly through glycoprotein (GP) Ib-V-IX receptor complex, or directly through the major collagen receptor on the platelet surface, GPVI. The work presented here focused on studying the cellular regulation of GPVI. In addition, a possible role for GPVI in thrombus formation induced by atherosclerotic plaque material was investigated and it was found that GPVI plays an important role in this process. Using a recently published mitochondrial injury model, it was found that GPVI contains a cleavage site for a platelet-expressed metalloproteinase. Further studies showed that platelet activation by CRP, or thrombin induced down-regulation of GPIb, but not GPVI. In parallel, cellular regulation of GPV was studied and it was found that GPV is cleaved in vitro by the metalloproteinase ADAM17. In previous studies it was shown that injection of mice with the anti-GPVI mAb, JAQ1, induces GPVI down-regulation, which is associated with a strong, but transient, thrombocytopenia. Using new anti-GPVI mAbs, which bind different epitopes on the receptor, it is shown in this study that GPVI down-regulation occurs in an epitope-independent manner. Further experiments showed that antibody treatment induces a transient, but significant increase in bleeding time. Using different genetically modified mice, it is shown that, upon antibody injection, GPVI is both, shed from the platelet surface and internalized into the platelet. Signaling through the immunoreceptor tyrosine-based activation motif (ITAM) of the FcR chain is essential for both processes, while LAT and PLC2 are essential for the shedding process only. Antibody-induced increase in bleeding time and thrombocytopenia were absent in LAT deficient mice, showing that it is possible to uncouple the associated side effects from the down-regulation process. As antibody-induced GPVI internalization still occurs in LAT and PLC2 deficient mice, this suggests a novel signaling pathway downstream of GPVI that has not been described so far.
N2  - Plättchen Interaktion mit dem Subendothel ist für die Blutstillung essentiell. Dies kann jedoch nach dem Aufbrechen atherosklerotischer Plaques zu lebensbedrohlicher Erkrankungen wie Infarkt oder Schlaganfall führen. Kollagen, welches die Plättchen dirket aktiviert, ist der thrombogenste Bestandteil der Extrazellularmatrix (EZM). Die Bindung zwischen Plättchen und Kollagen wird sowohl indirekt durch den Glykoprotein (GP) Ib-V-IX Rezeptorkomplex, als auch direkt durch den Kollagenrezeptor GPVI, auf der Plättchenoberfläche vermittelt. In der vorliegenden Arbeit wurde die zelluläre Regulation von GPVI untersucht. Des Weiteren wurde die Rolle von GPVI in durch atheroklerotisches Plaquematerial induzierter Thrombusbildung studiert. Hierbei wurde festgestellt, dass GPVI eine wichtige Funktion in diesem Prozess spielt. Mittels eines jüngst publizierten mitochondrialen Verletzungsmodels, konnte gezeigt werden, dass GPVI eine Erkennungsstelle für eine in den Plättchen exprimierte Metalloproteinase besitzt. Mehrere Versuche haben gezeigt, dass Plättchenaktivierung durch CRP, und Thrombin zur Runterregulierung von GPIb aber nicht von GPVI führt. Parallellaufende Untersuchungen zeigten, dass GPV durch die Metalloproteinase ADAM17 in vitro abgespalten wird. Vorherige Studien ergaben, dass die in vivo Behandlung von Mäusen mit dem anti-GPVI Antikörper, JAQ1, zur Runterregulierung des Rezeptors führt. Dieses ist mit einer starken, transienten Thrombozytopenie assoziiert. Mittels neu generierte anti-GPVI Antikörper (JAQ2, 3), die unterschiedliche Bindungsstellen auf GPVI erkennen, konnte demonstriert werden, dass die Antikörper vermittele GPVI Runterregulierung Epitop unabhängig ist. Weitere Untersuchungen ergaben, dass Anitkörperinjektion eine transiente Erhöhung der Blutungszeit verursacht. Mittels genetisch modifizierter Mäuse konnte dargestellt werden, dass die Antikörpergabe GPVI sowohl von der Plättchenoberfläche abgespalten, als auch internalisiert wird. Während die Signaltransduktion durch das ITAM Motif der FcR Kette essentiell für beide Prozesse ist, sind LAT und PLC2 nur für das Abspalten wichtig. Antikörper induzierte Erhöhung der Blutungszeit und Thrombozytopenie sind abwesend in LAT-defizienten Mäuse, was zeigt, dass möglicherweise die GPVI Runterregulierung von den assoziierten Nebenwirkungen zu trennen ist. Da die GPVI Runterregulierung in LAT und –PLC2 defizienten Mäusen weiterhin stattfindet, zeigt dies einen neuen GPVI Signalweg, der bisher noch nicht beschrieben wurde.
KW  - Maus
KW  - Thrombozyt
KW  - Glykoproteine
KW  - Regulation
KW  - Biologie
KW  - Plättchen
KW  - Maus
KW  - Thrombose
KW  - Kardiovaskulär
KW  - maus
KW  - platelets
KW  - thrombosis
KW  - cardiovascular
Y1  - 2005
U6  - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-14267
ER  -