Madeleine Austinat

Publications of Madeleine Austinat

• Inhibition of bradykinin receptor B1 protects mice from focal brain injury by reducing blood-brain barrier leakage and inflammation.

  Authors: Furat Raslan, Tobias Schwarz, Sven G Meuth, Madeleine Austinat, Michael Bader, Thomas Renné, Klaus Roosen, Guido Stoll, Anna-Leena Sirén, Christoph Kleinschnitz

  Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism. 03/2010; 30(8):1477-86.

  Kinins are proinflammatory and vasoactive peptides that are released during tissue damage and may contribute to neuronal degeneration, inflammation, and edema formation after brain injury by actingKinins are proinflammatory and vasoactive peptides that are released during tissue damage and may contribute to neuronal degeneration, inflammation, and edema formation after brain injury by acting on discrete bradykinin receptors, B1R and B2R. We studied the expression of B1R and B2R and the effect of their inhibition on lesion size, blood-brain barrier (BBB) disruption, and inflammatory processes after a focal cryolesion of the right parietal cortex in mice. B1R and B2R gene transcripts were significantly induced in the lesioned hemispheres of wild-type mice (P<0.05). The volume of the cortical lesions and neuronal damage at 24 h after injury in B1R(-/-) mice were significantly smaller than in wild-type controls (2.5+/-2.6 versus 11.5+/-3.9 mm(3), P<0.001). Treatment with the B1R antagonist R-715 1 h after lesion induction likewise reduced lesion volume in wild-type mice (2.6+/-1.4 versus 12.2+/-6.1 mm(3), P<0.001). This was accompanied by a remarkable reduction of BBB disruption and tissue inflammation. In contrast, genetic deletion or pharmacological inhibition of B2R had no significant impact on lesion formation or the development of brain edema. We conclude that B1R inhibition may offer a novel therapeutic strategy after acute brain injuries.

• Early detrimental T-cell effects in experimental cerebral ischemia are neither related to adaptive immunity nor thrombus formation.

  Authors: Christoph Kleinschnitz, Nicholas Schwab, Peter Kraft, Ina Hagedorn, Angela Dreykluft, Tobias Schwarz, Madeleine Austinat, Bernhard Nieswandt, Heinz Wiendl, Guido Stoll

  Blood. 03/2010; 115(18):3835-42.

  T cells contribute to the pathophysiology of ischemic stroke by yet unknown mechanisms. Mice with transgenic T-cell receptors (TCRs) and mutations in costimulatory molecules were used to define theT cells contribute to the pathophysiology of ischemic stroke by yet unknown mechanisms. Mice with transgenic T-cell receptors (TCRs) and mutations in costimulatory molecules were used to define the minimal immunologic requirements for T cell-mediated ischemic brain damage. Stroke was induced in recombination activating gene 1-deficient (RAG1(-/-)) mice devoid of T and B cells, RAG1(-/-) mice reconstituted with B cells or T cells, TCR-transgenic mice bearing 1 single CD8(+) (2C/RAG2, OTI/RAG1 mice) or CD4(+) (OTII/RAG1, 2D2/RAG1 mice) TCR, mice lacking accessory molecules of TCR stimulation (CD28(-/-), PD1(-/-), B7-H1(-/-) mice), or mice deficient in nonclassical T cells (natural killer T [NKT] and gammadelta T cells) by transient middle cerebral artery occlusion (tMCAO). Stroke outcome was assessed at day 1. RAG1(-/-) mice and RAG1(-/-) mice reconstituted with B cells developed significantly smaller brain infarctions compared with controls, but thrombus formation after FeCl(3)-induced vessel injury was unimpaired. In contrast, TCR-transgenic mice and mice lacking costimulatory TCR signals were fully susceptible to tMCAO similar to mice lacking NKT and gammadelta T cells. These findings were corroborated by adoptive transfer experiments. Our data demonstrate that T cells critically contribute to cerebral ischemia, but their detrimental effect neither depends on antigen recognition nor TCR costimulation or thrombus formation.

• Deficiency of vasodilator-stimulated phosphoprotein (VASP) increases blood-brain-barrier damage and edema formation after ischemic stroke in mice.

  Authors: Peter Kraft, Peter Michael Benz, Madeleine Austinat, Marc Elmar Brede, Kai Schuh, Ulrich Walter, Guido Stoll, Christoph Kleinschnitz

  PloS one. 01/2010; 5(12):e15106.

  Stroke-induced brain edema formation is a frequent cause of secondary infarct growth and deterioration of neurological function. The molecular mechanisms underlying edema formation after stroke areStroke-induced brain edema formation is a frequent cause of secondary infarct growth and deterioration of neurological function. The molecular mechanisms underlying edema formation after stroke are largely unknown. Vasodilator-stimulated phosphoprotein (VASP) is an important regulator of actin dynamics and stabilizes endothelial barriers through interaction with cell-cell contacts and focal adhesion sites. Hypoxia has been shown to foster vascular leakage by downregulation of VASP in vitro but the significance of VASP for regulating vascular permeability in the hypoxic brain in vivo awaits clarification. Focal cerebral ischemia was induced in Vasp(-/-) mice and wild-type (WT) littermates by transient middle cerebral artery occlusion (tMCAO). Evan's Blue tracer was applied to visualize the extent of blood-brain-barrier (BBB) damage. Brain edema formation and infarct volumes were calculated from 2,3,5-triphenyltetrazolium chloride (TTC)-stained brain slices. Both mouse groups were carefully controlled for anatomical and physiological parameters relevant for edema formation and stroke outcome. BBB damage (p<0.05) and edema volumes (1.7 mm(3)±0.5 mm(3) versus 0.8 mm(3)±0.4 mm(3); p<0.0001) were significantly enhanced in Vasp(-/-) mice compared to controls on day 1 after tMCAO. This was accompanied by a significant increase in infarct size (56.1 mm(3)±17.3 mm(3) versus 39.3 mm(3)±10.7 mm(3), respectively; p<0.01) and a non significant trend (p>0.05) towards worse neurological outcomes. Our study identifies VASP as critical regulator of BBB maintenance during acute ischemic stroke. Therapeutic modulation of VASP or VASP-dependent signalling pathways could become a novel strategy to combat excessive edema formation in ischemic brain damage.

• Deficiency of von Willebrand factor protects mice from ischemic stroke.

  Authors: Christoph Kleinschnitz, Simon F De Meyer, Tobias Schwarz, Madeleine Austinat, Karen Vanhoorelbeke, Bernhard Nieswandt, Hans Deckmyn, Guido Stoll

  Blood. 01/2009;

  We recently demonstrated that blockade of the platelet adhesion receptor glycoprotein(GP)Ibalpha protects mice from ischemic stroke. Although von Willebrand factor (VWF) is the major ligand forWe recently demonstrated that blockade of the platelet adhesion receptor glycoprotein(GP)Ibalpha protects mice from ischemic stroke. Although von Willebrand factor (VWF) is the major ligand for GPIbalpha, GPIbalpha can engage other counterreceptors on endothelial cells, platelets and leukocytes (e.g. Mac-1 or P-selectin) potentially involved in stroke outcome. To further analyze whether VWF is of particular relevance for stroke development, VWF(-/-) mice underwent 60 min of middle cerebral artery occlusion. After 24 h, VWF(-/-) mice had significantly smaller infarctions (p<0.05) and less severe neurological deficits (p<0.01) compared to controls. This effect was sustained after 1 week and intracranial bleeding was absent in VWF(-/-) mice as revealed by serial magnetic resonance imaging. Hydrodynamic injection of a VWF-encoding plasmid restored the susceptibility for stroke in VWF(-/-) mice. This study indicates that VWF is critically involved in cerebral ischemia. Hence, targeted inhibition of the GPIbalpha-VWF pathway might become a promising therapeutic option.

• Blockade of Bradykinin Receptor B1 but Not Bradykinin Receptor B2 Provides Protection From Cerebral Infarction and Brain Edema.

  Authors: Madeleine Austinat, Stefan Braeuninger, João B Pesquero, Marc Brede, Michael Bader, Guido Stoll, Thomas Renné, Christoph Kleinschnitz

  Stroke; a journal of cerebral circulation. 12/2008;

  BACKGROUND AND PURPOSE: Brain edema is detrimental in ischemic stroke and its treatment options are limited. Kinins are proinflammatory peptides that are released during tissue injury. The effects ofBACKGROUND AND PURPOSE: Brain edema is detrimental in ischemic stroke and its treatment options are limited. Kinins are proinflammatory peptides that are released during tissue injury. The effects of kinins are mediated by 2 different receptors (B1 and B2 receptor [B1R and B2R]) and comprise induction of edema formation and release of proinflammatory mediators. METHODS: Focal cerebral ischemia was induced in B1R knockout, B2R knockout, and wild-type mice by transient middle cerebral artery occlusion. Infarct volumes were measured by planimetry. Evan's blue tracer was applied to determine the extent of brain edema. Postischemic inflammation was assessed by real-time reverse-transcriptase polymerase chain reaction and immunohistochemistry. To analyze the effect of a pharmacological kinin receptor blockade, B1R and B2R inhibitors were injected. RESULTS: B1R knockout mice developed significantly smaller brain infarctions and less neurological deficits compared to wild-type controls (16.8+/-4.7 mm(3) vs 50.1+/-9.1 mm(3), respectively; P<0.0001). This was accompanied by a dramatic reduction of brain edema and endothelin-1 expression, as well as less postischemic inflammation. Pharmacological blockade of B1R likewise salvaged ischemic tissue (15.0+/-9.5 mm(3) vs 50.1+/-9.1 mm(3), respectively; P<0.01) in a dose-dependent manner, even when B1R inhibitor was applied 1 hour after transient middle cerebral artery occlusion. In contrast, B2R deficiency did not confer neuroprotection and had no effect on the development of tissue edema. CONCLUSIONS: These data demonstrate that blocking of B1R can diminish brain infarction and edema formation in mice and may open new avenues for acute stroke treatment in humans.

• Orai1 (CRACM1) is the platelet SOC channel and essential for pathological thrombus formation.

  Authors: Attila Braun, David Varga-Szabo, Christoph Kleinschnitz, Irina Pleines, Markus Bender, Madeleine Austinat, Michael Bosl, Guido Stoll, Bernhard Nieswandt

  Blood. 11/2008;

  Platelet activation and aggregation at sites of vascular injury is essential for primary hemostasis, but is also a major pathomechanism underlying myocardial infarction and stroke. Changes inPlatelet activation and aggregation at sites of vascular injury is essential for primary hemostasis, but is also a major pathomechanism underlying myocardial infarction and stroke. Changes in [Ca(2+)]i are a central step in platelet activation. In non-excitable cells receptor-mediated depletion of intracellular Ca(2+) stores triggers Ca(2+) entry through store-operated calcium (SOC) channels. STIM1 has been identified as an endoplasmic reticulum (ER)-resident Ca(2+) sensor that regulates store-operated calcium entry (SOCE) in immune cells and platelets, but the identity of the platelet SOC channel has remained elusive. Orai1 (CRACM1) is the recently discovered SOC (CRAC) channel in T-cells and mast cells but its role in mammalian physiology is unknown. Here we report that Orai1 is strongly expressed in human and mouse platelets. To test its role in blood clotting, we generated Orai1-deficient mice and found that their platelets display severely defective SOCE, agonist-induced Ca(2+) responses, and impaired activation and thrombus formation under flow in vitro. As a direct consequence, Orai1-deficiency in mice results in resistance to pulmonary thromboembolism, arterial thrombosis and ischemic brain infarction, but only a mild bleeding time prolongation. These results establish Orai1 as the long-sought platelet SOC channel and a crucial mediator of ischemic cardio- and cerebrovascular events.

• The neuroprotective impact of the leak potassium channel TASK1 on stroke development in mice.

  Authors: Sven G Meuth, Christoph Kleinschnitz, Tilman Broicher, Madeleine Austinat, Stefan Bräuninger, Stefan Bittner, Stephan Fischer, Douglas A Bayliss, Thomas Budde, Guido Stoll, Heinz Wiendl

  Neurobiology of disease. 10/2008;

  Oxygen depletion (O(2)) and a decrease in pH are initial pathophysiological events in stroke development, but secondary mechanisms of ischemic cell death are incompletely understood. By patch-clampOxygen depletion (O(2)) and a decrease in pH are initial pathophysiological events in stroke development, but secondary mechanisms of ischemic cell death are incompletely understood. By patch-clamp recordings of brain slice preparations we show that TASK1 and TASK3 channels are inhibited by pH-reduction (42+/-2%) and O(2) deprivation (36+/-5%) leading to membrane depolarization, increased input resistance and a switch in action potential generation under ischemic conditions. In vivo TASK blockade by anandamide significantly increased infarct volumes at 24 h in mice undergoing 30 min of transient middle cerebral artery occlusion (tMCAO). Moreover, blockade of TASK channels accelerated stroke development. Supporting these findings TASK1(-/-) mice developed significantly larger infarct volumes after tMCAO accompanied by worse outcome in functional neurological tests compared to wild type mice. In conclusion, our data provide evidence for an important role of functional TASK channels in limiting tissue damage during cerebral ischemia.

• Deficiency of bradykinin receptor B2 is not detrimental in experimental stroke.

  Authors: Christoph Kleinschnitz, Madeleine Austinat, Michael Bader, Thomas Renné, Guido Stoll

  Hypertension. 06/2008; 51(5):e41; author reply e42-3.

• Blocking of platelets or intrinsic coagulation pathway-driven thrombosis does not prevent cerebral infarctions induced by photothrombosis.

  Authors: Christoph Kleinschnitz, Stefan Braeuninger, Mirko Pham, Madeleine Austinat, Ingo Nölte, Thomas Renné, Bernhard Nieswandt, Martin Bendszus, Guido Stoll

  Stroke; a journal of cerebral circulation. 05/2008; 39(4):1262-8.

  BACKGROUND AND PURPOSE: Models of photochemically-induced thrombosis are widely used in cerebrovascular research. Photothrombotic brain infarctions can be induced by systemic application ofBACKGROUND AND PURPOSE: Models of photochemically-induced thrombosis are widely used in cerebrovascular research. Photothrombotic brain infarctions can be induced by systemic application of photosensitizing dyes followed by focal illumination of the cerebral cortex. Although the ensuing activation of platelets is well established, their contribution for thrombosis and tissue damage has not formally been proved. METHODS: Infarction to the cerebral cortex was induced in mice by Rose Bengal and a cold light source. To assess the functional role of platelets, animals were platelet-depleted by anti-GPIbalpha antibodies or treated with GPIIb/IIIa-blocking F(ab)(2) fragments. The significance of the plasmatic coagulation cascade was determined by using blood coagulation factor XII (FXII)-deficient mice or heparin. Infarct development and infarct volumes were determined by serial MRI and conventional and electron microscopy. RESULTS: There was no difference in development and final size of photothrombotic infarctions in mice with impaired platelet function. Moreover, deficiency of FXII, which initiates the intrinsic pathway of coagulation and is essential for thrombus formation, or blockade of FXa, the key protease during the waterfall cascade of plasmatic coagulation, by heparin likewise did not affect lesion development. CONCLUSIONS: Our data demonstrate that platelet activation, factor XII-driven thrombus formation, and plasmatic coagulation pathways downstream of FX are not a prerequisite for ensuing tissue damage in models of photothrombotic vessel injury indicating that other pathomechanisms are involved. We suggest that this widely used model does not depend on platelet- or plasmatic coagulation-derived thrombosis.

• The neuroprotective impact of the leak potassium channel TASK1 on stroke development in mice

  Authors: Sven G. Meuth, Christoph Kleinschnitz, Tilman Broicher, Madeleine Austinat, Stefan Braeuninger, Stefan Bittner, Stephan Fischer, Douglas A. Bayliss, Thomas Budde, Guido Stoll, Heinz Wiendl

  Neurobiology of Disease.

  Oxygen depletion (O2) and a decrease in pH are initial pathophysiological events in stroke development, but secondary mechanisms of ischemic cell death are incompletely understood. By patch-clampOxygen depletion (O2) and a decrease in pH are initial pathophysiological events in stroke development, but secondary mechanisms of ischemic cell death are incompletely understood. By patch-clamp recordings of brain slice preparations we show that TASK1 and TASK3 channels are inhibited by pH-reduction (42 ± 2%) and O2 deprivation (36 ± 5%) leading to membrane depolarization, increased input resistance and a switch in action potential generation under ischemic conditions. In vivo TASK blockade by anandamide significantly increased infarct volumes at 24 h in mice undergoing 30 min of transient middle cerebral artery occlusion (tMCAO). Moreover, blockade of TASK channels accelerated stroke development. Supporting these findings TASK1−/− mice developed significantly larger infarct volumes after tMCAO accompanied by worse outcome in functional neurological tests compared to wild type mice. In conclusion, our data provide evidence for an important role of functional TASK channels in limiting tissue damage during cerebral ischemia.