DHA, RvD1, RvD5, and MaR1 reduce human coronary arteries contractions induced by PGE2.

In patients with coronary artery disease (CAD), plasma levels of pro-inflammatory lipid mediators such as PGE2 and TxA2 are increased. They could increase vascular contraction while EPA and DHA could reduce it. Studies have been mostly conducted on animal vessels. Therefore, the aim of the study was to investigate if EPA, DHA, and DHA-derived metabolites: RvD1, RvD5 and MaR1 can modulate contraction of human coronary arteries (HCA) induced by PGE2 or TxA2 stable analogue (U46619). DHA and EPA relaxed HCA pre-contracted with PGE2. 18 h-incubation with DHA but not EPA reduced the PGE2-induced contractions. Pre-incubation with RvD1, RvD5 and MaR1 reduced the PGE2-induced contractions. Indomethacin did not significantly modify the PGE2 responses. L-NOARG (inhibitor of nitric oxide synthase), reduced only the PGE2-induced contractions in RvD1-treated rings. Finally, FPR2/ALX, GPR32 and LGR6 receptors are detected in HCA by immunofluorescence. Our results indicate that DHA and its metabolites could be beneficial for HCA blood flow and could be a therapeutic perspective for patients with CAD.

Intravenous abciximab as a rescue therapy for immediate reocclusion after successful mechanical thrombectomy in acute ischemic stroke patients.

Immediate reocclusion after mechanical thrombectomy (MT) for acute ischemic stroke (AIS) is a rare but devastating condition associated with poor functional outcome. The aim of this study was to gain insights into the mechanisms underlying immediate reocclusion, and to evaluate the efficacy and safety of the glycoprotein IIb/IIIa antagonist abciximab, for its treatment. Clinical data were collected from April 2015 to April 2019 in a monocentric prospective registry of AIS patients treated by MT. All patients with immediate reocclusion were retrospectively selected and subdivided into 2 groups according to abciximab treatment status. In vitro, the separate and combined effects of abciximab and alteplase on clot formation in whole blood under flow conditions were further investigated in microfluidic chambers. From 929 MT-treated patients, 21 had post-MT immediate reocclusion. Abciximab treatment in reocclusion patients (n = 10) led to higher rate of final recanalization (p < .001) while it did not increase bleeding complications. Flow chamber experiments revealed that, in contrast to alteplase, abciximab efficiently limits thrombus accretion from flowing blood by blocking platelet aggregation. Our results underscore a key role for platelet aggregation and the potential of Glycoprotein IIb/IIIa antagonists as a rescue therapy in post-MT immediate reocclusion.

Pathogenic variants in THSD4, encoding the ADAMTS-like 6 protein, predispose to inherited thoracic aortic aneurysm.

PURPOSE: Thoracic aortic aneurysm and dissection (TAAD) is a life-threatening disease with often unrecognized inherited forms. We sought to identify novel pathogenic variants associated with autosomal dominant inheritance of TAAD. METHODS: We analyzed exome sequencing data from 35 French TAAD families and performed next-generation sequencing capture panel of genes in 1114 unrelated TAAD patients. Functional effects of pathogenic variants identified were validated in cell, tissue, and mouse models. RESULTS: We identified five functional variants in THSD4 of which two heterozygous variants lead to a premature termination codon. THSD4 encodes ADAMTSL6 (member of the ADAMTS/L superfamily), a microfibril-associated protein that promotes fibrillin-1 matrix assembly. The THSD4 variants studied lead to haploinsufficiency or impaired assembly of fibrillin-1 microfibrils. Thsd4+/- mice showed progressive dilation of the thoracic aorta. Histologic examination of aortic samples from a patient carrying a THSD4 variant and from Thsd4+/- mice, revealed typical medial degeneration and diffuse disruption of extracellular matrix. CONCLUSION: These findings highlight the role of ADAMTSL6 in aortic physiology and TAAD pathogenesis. They will improve TAAD management and help develop new targeted therapies.

Glenzocimab does not impact glycoprotein VI-dependent inflammatory haemostasis.

Not available.

Lymphatic blood filling in CLEC-2-deficient mouse models.

C-type lectin-like receptor 2 (CLEC-2) is considered as a potential drug target in settings of wound healing, inflammation, and infection. A potential barrier to this is evidence that CLEC-2 and its ligand podoplanin play a critical role in preventing lymphatic vessel blood filling in mice throughout life. In this study, this aspect of CLEC-2/podoplanin function is investigated in more detail using new and established mouse models of CLEC-2 and podoplanin deficiency, and models of acute and chronic vascular remodeling. We report that CLEC-2 expression on platelets is not required to maintain a barrier between the blood and lymphatic systems in unchallenged mice, post-development. However, under certain conditions of chronic vascular remodeling, such as during tumorigenesis, deficiency in CLEC-2 can lead to lymphatic vessel blood filling. These data provide a new understanding of the function of CLEC-2 in adult mice and confirm the essential nature of CLEC-2-driven platelet activation in vascular developmental programs. This work expands our understanding of how lymphatic blood filling is prevented by CLEC-2-dependent platelet function and provides a context for the development of safe targeting strategies for CLEC-2 and podoplanin.

Platelet Serotonin Aggravates Myocardial Ischemia/Reperfusion Injury via Neutrophil Degranulation.

BACKGROUND: Platelets store large amounts of serotonin that they release during thrombus formation or acute inflammation. This facilitates hemostasis and modulates the inflammatory response. METHODS: Infarct size, heart function, and inflammatory cell composition were analyzed in mouse models of myocardial reperfusion injury with genetic and pharmacological depletion of platelet serotonin. These studies were complemented by in vitro serotonin stimulation assays of platelets and leukocytes in mice and men, and by measuring plasma serotonin levels and leukocyte activation in patients with acute coronary syndrome. RESULTS: Platelet-derived serotonin induced neutrophil degranulation with release of myeloperoxidase and hydrogen peroxide (H2O2) and increased expression of membrane-bound leukocyte adhesion molecule CD11b, leading to enhanced inflammation in the infarct area and reduced myocardial salvage. In patients hospitalized with acute coronary syndrome, plasmatic serotonin levels correlated with CD11b expression on neutrophils and myeloperoxidase plasma levels. Long-term serotonin reuptake inhibition-reported to protect patients with depression from cardiovascular events-resulted in the depletion of platelet serotonin stores in mice. These mice displayed a reduction in neutrophil degranulation and preserved cardiac function. In line, patients with depression using serotonin reuptake inhibition, presented with suppressed levels of CD11b surface expression on neutrophils and lower myeloperoxidase levels in blood. CONCLUSIONS: Taken together, we identify serotonin as a potent therapeutic target in neutrophil-dependent thromboinflammation during myocardial reperfusion injury.

Protective Effect of ApoA1 (Apolipoprotein A1)-Milano in a Rat Model of Large Vessel Occlusion Stroke.

Background and Purpose- Previous experimental studies found that the infusion of human purified nascent HDL (high-density lipoprotein) significantly reduced infarct volume and hemorrhagic transformation rate by decreasing neutrophil recruitment. ApoA1-M (apolipoprotein A1-Milano) is a natural variant of human ApoA1 that confers protection against atherosclerosis. Recombinant ApoA1-M has been formulated as a complex with phospholipids to mimic the properties of nascent HDL. The aim of this study was to assess the impact of intravenous ApoA1-M in a transient middle cerebral artery occlusion stroke model in rats. Methods- In a first experiment, rats were subjected to 120-minute transient middle cerebral artery occlusion and intravenous ApoA1-M was infused immediately or 4 hours after occlusion. In a second experiment, rats were subjected to 240-minute transient middle cerebral artery occlusion and intravenous ApoA1-M was infused with or without recombinant tPA (tissue-type plasminogen activator) immediately after recanalization. Primary outcome criteria were the infarct volume and hemorrhagic transformation rate measured at 24 hours. Platelets, coagulation, and neutrophil activation biomarkers were measured in brain homogenates and plasma. Additional in vitro experiments studied the effects of ApoA1-M on platelet aggregation and platelet-neutrophil interactions. Results- The infusion of ApoA1-M immediately or 4 hours after 120-minute transient middle cerebral artery occlusion significantly reduced the infarct volume compared with saline (P=0.034 and P=0.036, respectively). Compared with tPA alone, co-administration of ApoA1-M and tPA showed similar rates of hemorrhagic transformation. ApoA1-M had no significant inhibition effect on neutrophil activation biomarkers. Platelet activation was slightly decreased in rats treated with ApoA1-M compared with saline. In vitro, the incubation of human and rat platelet-rich plasma with ApoA1-M significantly reduced ADP-induced platelet aggregation (P=0.001 and P=0.02, respectively). Conclusions- ApoA1-Milano significantly decreased the infarct volume through an inhibition of platelet aggregation but did not reduce hemorrhagic transformation and neutrophils activation as expected after previous experimental studies with nascent HDL. Visual Overview- An online visual overview is available for this article.

DNA Content in Ischemic Stroke Thrombi Can Help Identify Cardioembolic Strokes Among Strokes of Undetermined Cause.

BACKGROUND AND PURPOSE: Identification of acute ischemic stroke (AIS) cause is crucial for guidance of secondary prevention. Previous studies have yielded inconsistent results regarding possible correlations between AIS cause and thrombus composition, as assessed by semiquantitative histological analysis. Here, we performed a correlation analysis between AIS cause and AIS thrombus cellular composition and content, as assessed using quantitative biochemical assays. METHODS: Homogenates of 250 patients with AIS thrombi were prepared by mechanical grinding. Platelet, red blood cell, and leukocyte content of AIS thrombi were estimated by quantification of GP (glycoprotein) VI, heme, and DNA in thrombus homogenates. AIS cause was defined as cardioembolic, noncardioembolic, or embolic stroke of undetermined source, according to the TOAST classification (Trial of ORG 10172 in Acute Stroke Treatment). RESULTS: Cardioembolic thrombi were richer in DNA (35.8 versus 13.8 ng/mg, P<0.001) and poorer in GPVI (0.104 versus 0.117 ng/mg, P=0.045) than noncardioembolic ones. The area under the receiver operating characteristic curve of DNA content to discriminate cardioembolic thrombi from noncardioembolic was 0.72 (95% CI, 0.63-0.81). With a threshold of 44.7 ng DNA/mg thrombus, 47% of thrombi from undetermined cause would be classified as cardioembolic with a specificity of 90%. CONCLUSIONS: Thrombus DNA content may provide an accurate biomarker for identification of cardioembolic thrombi in patients with AIS with embolic stroke of undetermined source. Registration: URL: https://www.clinicaltrials.gov. Unique identifier: NCT03268668.

Inhibition of macrophage proliferation dominates plaque regression in response to cholesterol lowering.

Statins induce plaque regression characterized by reduced macrophage content in humans, but the underlying mechanisms remain speculative. Studying the translational APOE*3-Leiden.CETP mouse model with a humanized lipoprotein metabolism, we find that systemic cholesterol lowering by oral atorvastatin or dietary restriction inhibits monocyte infiltration, and reverses macrophage accumulation in atherosclerotic plaques. Contrary to current believes, none of (1) reduced monocyte influx (studied by cell fate mapping in thorax-shielded irradiation bone marrow chimeras), (2) enhanced macrophage egress (studied by fluorescent bead labeling and transfer), or (3) atorvastatin accumulation in murine or human plaque (assessed by mass spectrometry) could adequately account for the observed loss in macrophage content in plaques that undergo phenotypic regression. Instead, suppression of local proliferation of macrophages dominates phenotypic plaque regression in response to cholesterol lowering: the lower the levels of serum LDL-cholesterol and lipid contents in murine aortic and human carotid artery plaques, the lower the rates of in situ macrophage proliferation. Our study identifies macrophage proliferation as the predominant turnover determinant and an attractive target for inducing plaque regression.

Platelets and vascular integrity: how platelets prevent bleeding in inflammation.

Platelets play a central role in primary hemostasis by forming aggregates that plug holes in injured vessels. Half a century ago, detailed studies of the microvasculature by electron microscopy revealed that under inflammatory conditions that do not induce major disruption to vascular structure, individual platelets are mobilized to the vessel wall, where they interact with leukocytes and appear to seal gaps that arise between endothelial cells. Recent developments in genetic engineering and intravital microscopy have allowed further molecular and temporal characterization of these events. Surprisingly, it turns out that platelets support the recruitment of leukocytes to sites of inflammation. In parallel, however, they exercise their hemostatic function by securing the integrity of inflamed blood vessels to prevent bleeding from sites of leukocyte infiltration. It thus appears that platelets not only serve in concert as building blocks of the hemostatic plug but also act individually as gatekeepers of the vascular wall to help preserve vascular integrity while coordinating host defense. Variants of this recently appreciated hemostatic function of platelets that we refer to as "inflammation-associated hemostasis" are engaged in different contexts in which the endothelium is challenged or dysfunctional. Although the distinguishing characteristics of these variants and the underlying mechanisms of inflammation-associated hemostasis remain to be fully elucidated, they can differ notably from those supporting thrombosis, thus presenting therapeutic opportunities.

The reversed passive Arthus reaction as a model for investigating the mechanisms of inflammation-associated hemostasis.

In recent years, accumulating evidence has indicated that platelets continuously repair vascular damage at sites of inflammation and/or infection. Studies in mouse models of inflammation have highlighted the fact that the mechanisms underlying bleeding prevention by platelets in inflamed organs can substantially differ from those supporting primary hemostasis following tail tip transection or thrombus formation in models of thrombosis. As a consequence, exploration of the hemostatic function of platelets in inflammation, as well as assessment of the risk of inflammation-induced bleeding associated with a platelet deficit and/or the use of anti-thrombotic drugs, require the use of dedicated experimental models. In the present review, we present the pros and cons of the cutaneous reversed passive Arthus reaction, a model of inflammation which has been instrumental in studying how inflammation causes vascular injury and how platelets continuously intervene to repair it. The limitations and common issues encountered when working with mouse models of inflammation for investigating platelet functions in inflammation are also discussed.

Neurologic Complications of Infective Endocarditis: A Joint Model for a Septic Thromboembolism and Inflammatory Small Vessel Disease.

OBJECTIVES: Embolic events from vegetations are commonly accepted as the main mechanism involved in neurologic complications of infective endocarditis. The pathophysiology may imply other phenomena, including vasculitis. We aimed to define the cerebral lesion spectrum in an infective endocarditis rat model. DESIGN: Experimental model of Staphylococcus aureus or Enterococcus faecalis infective endocarditis. Neurologic lesions observed in the infective endocarditis model were compared with three other conditions, namely bacteremia, nonbacterial thrombotic endocarditis, and healthy controls. SETTING: Research laboratory of a university hospital. SUBJECTS: Male Wistar rats. INTERVENTIONS: Brain MRI, neuropathology, immunohistochemistry for astrocyte and microglia, and bacterial studies on brain tissue were used to characterize neurologic lesions. MEASUREMENTS AND MAIN RESULTS: In the infective endocarditis group, MRI revealed at least one cerebral lesion in 12 of 23 rats (52%), including brain infarctions (n = 9/23, 39%) and cerebral microbleeds (n = 8/23, 35%). In the infective endocarditis group, neuropathology revealed brain infarctions (n = 12/23, 52%), microhemorrhages (n = 10/23, 44%), and inflammatory processes (i.e., cell infiltrates including abscesses, vasculitis, meningoencephalitis, and/or ependymitis; n = 11/23, 48%). In the bacteremia group, MRI studies were normal and neuropathology revealed only hemorrhages (n = 2/11, 18%). Neuropathologic patterns observed in the nonbacterial thrombotic endocarditis group were similar to those observed in the infective endocarditis group. Immunochemistry revealed higher microglial activation in the infective endocarditis group (n = 11/23, 48%), when compared with the bacteremia (n = 1/11, 9%; p = 0.03) and nonbacterial thrombotic endocarditis groups (n = 0/7, 0%; p = 0.02). CONCLUSIONS: This original model of infective endocarditis recapitulates the neurologic lesion spectrum observed in humans and suggests synergistic mechanisms involved, including thromboembolism and cerebral vasculitis, promoted by a systemic bacteremia-mediated inflammation.

Microfluidic Modeling of Thrombolysis.

Objective- Despite the high clinical relevance of thrombolysis, models for its study in human flowing blood are lacking. Our objective was to develop a microfluidic model for comparative evaluation of thrombolytic therapeutic strategies. Approach and Results- Citrated human blood was supplemented with 3,3'-dihexyloxacarbocyanine iodide and Alexa Fluor 647 fibrinogen conjugate, recalcified, and perfused for 3 to 4 minutes at venous or arterial wall shear rate in microfluidic flow chambers coated with collagen and tissue factor to generate nonocclusive fluorescent thrombi. A second perfusion was performed for 10 minutes with rhodamine-6G-labeled citrated whole blood, supplemented or not with r-tPA (recombinant tissue-type plasminogen activator), fluorescein isothiocyanate-conjugated r-tPA, and Alexa Fluor 568 plasminogen conjugate. Plasminogen and r-tPA bound to preformed thrombi and r-tPA caused a concentration-dependent decrease in thrombus fibrin content (up to 50% reduction at 15 µg/mL r-tPA) as assessed by fluorescence microscopy. Fibrinolysis was confirmed by measurement of D-dimers in the output flow. Remarkably, despite ongoing fibrinolysis, new platelets continued to be recruited to the thrombus under lysis. Under the arterial condition, combining r-tPA with hirudin enhanced fibrinolysis but did not prevent the recruitment of new platelets, which was, however, prevented by antiplatelet agents (ticagrelor or the GPVI [glycoprotein VI]-blocking antigen-binding fragment 9O12). Conclusions- Our microfluidic thrombolysis model is suitable for studying thrombolysis and testing the efficacy of drugs used in combination with r-tPA. Real-time analysis of fibrin and platelets during r-tPA-mediated fibrinolysis at arterial or venous flow conditions showed that platelets continue to accumulate during fibrinolysis. Such platelet accumulation may impair r-tPA-mediated recanalization.

Acoustophoretic purification of platelets: feasibility and impact on platelet activation and function.

Purity, limited platelet activation, and preservation of platelet function are important stakes of preparation of platelet concentrates (PC) for clinical use. In fact, contaminating red blood cells and leukocytes, as well as activated and/or poorly functional platelets in PC, represents a risk of poor efficiency and adverse side effects during platelet transfusion. Therefore, optimization of preparation and storage of PC is still an active field of research. Shear-induced platelet activation is an unwanted side effect of the hard-spin (up to 5000g) step of centrifugation-based methods currently used in blood banks to prepare PC from whole blood samples. Here, we evaluated the effectiveness of an acoustic-based fractionation device for the isolation of human platelets from whole blood bags. The purity, activation status, and functionality of platelets isolated by acoustopheresis were compared with those of platelets isolated using a reference protocol known to produce limited platelet activation and consisting of two consecutive soft-spin centrifugations (120g and 1200g). Platelet concentration and purity were determined using an automated hematology analyzer. Platelet activation status and platelet reactivity to collagen and thrombin were assessed in flow cytometry by measurement of surface expression of P-selectin and activated integrin αIIbß3. The ability of isolated platelets to incorporate into a thrombus when transfused to NOD/SCID mice was investigated by intravital microscopy using the ferric chloride-induced thrombosis model. Blood fractionation by acoustophoresis led to the elimination of more than 80% of red blood cells and leukocytes from the platelet fraction, whose mean purity was of 92.8 ± 12.8%. The activation status and reactivity to collagen and thrombin of acoustophoresis-isolated platelets were similar to those of platelets isolated by soft-spin centrifugation. Finally, acoustophoresis-isolated platelets were tethered, adhered to the vessel wall, and incorporated into a growing thrombus following ferric chloride-induced vascular injury. Together, our results indicate that acoustophoresis is a suitable method for the isolation of human platelets with minimal platelet activation and preservation of platelet function.

Thrombus Neutrophil Extracellular Traps Content Impair tPA-Induced Thrombolysis in Acute Ischemic Stroke.

BACKGROUND AND PURPOSE: Neutrophil Extracellular Traps (NETs) are DNA extracellular networks decorated with histones and granular proteins produced by activated neutrophils. NETs have been identified as major triggers and structural factors of thrombosis. A recent study designated extracellular DNA threads from NETs as a potential therapeutic target for improving tissue-type plasminogen activator (tPA)-induced thrombolysis in acute coronary syndrome. The aim of this study was to assess the presence of NETs in thrombi retrieved during endovascular therapy in patients with acute ischemic stroke (AIS) and their impact on tPA-induced thrombolysis. METHODS: We analyzed thrombi from 108 AIS patients treated with endovascular therapy. Thrombi were characterized by hematoxylin/eosin staining, immunostaining, and ex vivo enzymatic assay. Additionally, we assessed ex vivo the impact of deoxyribonuclease 1 (DNAse 1) on thrombolysis of AIS thrombi. RESULTS: Histological analysis revealed that NETs contributed to the composition of all AIS thrombi especially in their outer layers. Quantitative measurement of thrombus NETs content was not associated with clinical outcome or AIS pathogenesis but correlated significantly with endovascular therapy procedure length and device number of passes. Ex vivo, recombinant DNAse 1 accelerated tPA-induced thrombolysis, whereas DNAse 1 alone was ineffective. CONCLUSIONS: This study suggests that thrombus NETs content may be responsible for reperfusion resistance, including mechanical or pharmacological approaches with intravenous tPA, irrespectively of their etiology. The efficacy of a strategy involving an administration of DNAse 1 in addition to tPA should be explored in the setting of AIS. CLINICAL TRIAL REGISTRATION: URL: http://www.clinicaltrials.gov. Unique identifier: NCT02907736.

Platelet and Erythrocyte Sources of S1P Are Redundant for Vascular Development and Homeostasis, but Both Rendered Essential After Plasma S1P Depletion in Anaphylactic Shock.

RATIONALE: Sphingosine-1-phosphate (S1P) signaling is essential for vascular development and postnatal vascular homeostasis. The relative importance of S1P sources sustaining these processes remains unclear. OBJECTIVE: To address the level of redundancy in bioactive S1P provision to the developing and mature vasculature. METHODS AND RESULTS: S1P production was selectively impaired in mouse platelets, erythrocytes, endothelium, or smooth muscle cells by targeted deletion of genes encoding sphingosine kinases -1 and -2. S1P deficiency impaired aggregation and spreading of washed platelets and profoundly reduced their capacity to promote endothelial barrier function ex vivo. However, and in contrast to recent reports, neither platelets nor any other source of S1P was essential for vascular development, vascular integrity, or hemostasis/thrombosis. Yet rapid and profound depletion of plasma S1P during systemic anaphylaxis rendered both platelet- and erythrocyte-derived S1P essential for survival, with a contribution from blood endothelium observed only in the absence of circulating sources. Recovery was sensitive to aspirin in mice with but not without platelet S1P, suggesting that platelet activation and stimulus-response coupling is needed. S1P deficiency aggravated vasoplegia in this model, arguing a vital role for S1P in maintaining vascular resistance during recovery from circulatory shock. Accordingly, the S1P2 receptor mediated most of the survival benefit of S1P, whereas the endothelial S1P1 receptor was dispensable for survival despite its importance for maintaining vascular integrity. CONCLUSIONS: Although source redundancy normally secures essential S1P signaling in developing and mature blood vessels, profound depletion of plasma S1P renders both erythrocyte and platelet S1P pools necessary for recovery and high basal plasma S1P levels protective during anaphylactic shock.

Cholesterol crystallization in human atherosclerosis is triggered in smooth muscle cells during the transition from fatty streak to fibroatheroma.

Recent studies have shown that in addition to being major constituents of the atheromatous core, solid cholesterol crystals (CCs) promote atherosclerotic lesion development and rupture by causing mechanical damage and exerting cytotoxic and pro-inflammatory effects. These findings suggest that targeting CCs might represent a therapeutic strategy for plaque stabilization. However, little is known about how cholesterol crystallization is initiated in human atherothrombotic disease. Here, we investigated these mechanisms. We performed a thorough immunohistological analysis of non-embedded, minimally processed human aortic tissues, combining polarized light and fluorescence microscopy. We found that CC formation was initiated during the fatty streak to fibroatheroma transition in tight association with the death of intralesional smooth muscle cells (SMCs). Cholesterol-loaded human SMCs were capable of producing CCs in vitro, a process that was enhanced by type I collagen and by inhibition of autophagy and cholesterol esterification. The fibrous transition, which was characterized by increased type I collagen expression, was associated with changes in the expression of autophagy and cholesterol flux-related genes, including a decrease in the autophagic adapter p62 and an increase in the cholesterol intracellular transporter Niemann-Pick C1. Collagen was identified as a potent inducer of these changes in SMCs. Collagen-induced changes in cholesterol metabolism and autophagy flux in smooth muscle foam cells at the fibrolipid transition likely contribute to initiate cholesterol crystallization in human atherosclerosis. Also, our data are in support of a protective role of autophagy against CC formation. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Exacerbation of Thromboinflammation by Hyperglycemia Precipitates Cerebral Infarct Growth and Hemorrhagic Transformation.

BACKGROUND AND PURPOSE: Admission hyperglycemia is associated with a poor outcome in acute ischemic stroke. How hyperglycemia impacts the pathophysiology of acute ischemic stroke remains largely unknown. We investigated how preexisting hyperglycemia increases ischemia/reperfusion cerebral injury. METHODS: Normoglycemic and streptozotocin-treated hyperglycemic rats were subjected to transient middle cerebral artery occlusion. Infarct growth and brain perfusion were assessed by magnetic resonance imaging. Markers of platelet, coagulation, and neutrophil activation were measured in brain homogenates and plasma. Downstream microvascular thromboinflammation (DMT) was investigated by intravital microscopy. RESULTS: Hyperglycemic rats had an increased infarct volume with an increased blood-brain barrier disruption and hemorrhagic transformation rate compared with normoglycemic rats. Magnetic resonance imaging scans revealed that hyperglycemia enhanced and accelerated lesion growth and was associated with hemorrhagic transformation originating from territories that were still not completely reperfused at 1 hour after middle cerebral artery recanalization. Intravital microscopy and analysis of brain homogenates showed that DMT began immediately after middle cerebral artery occlusion and was exacerbated by hyperglycemia. Measurement of plasma serotonin and matrix metalloproteinase-9 indicated that platelets and neutrophils were preactivated in hyperglycemic rats. Neutrophils from hyperglycemic diabetic patients showed increased adhesion to endothelial cells as compared with neutrophils from normoglycemic donors in flow chamber experiments. CONCLUSIONS: We show that hyperglycemia primes the thromboinflammatory cascade, thus, amplifying middle cerebral artery occlusion-induced DMT. DMT exacerbation in hyperglycemic rats impaired reperfusion and precipitated neurovascular damage, blood-brain barrier disruption, and hemorrhagic transformation. Our results designate DMT as a possible target for reduction of the deleterious impact of hyperglycemia in acute ischemic stroke.

MFAP5 loss-of-function mutations underscore the involvement of matrix alteration in the pathogenesis of familial thoracic aortic aneurysms and dissections.

Thoracic aortic aneurysm and dissection (TAAD) is an autosomal-dominant disorder with major life-threatening complications. The disease displays great genetic heterogeneity with some forms allelic to Marfan and Loeys-Dietz syndrome, and an important number of cases still remain unexplained at the molecular level. Through whole-exome sequencing of affected members in a large TAAD-affected family, we identified the c.472C>T (p.Arg158(∗)) nonsense mutation in MFAP5 encoding the extracellular matrix component MAGP-2. This protein interacts with elastin fibers and the microfibrillar network. Mutation screening of 403 additional probands identified an additional missense mutation of MFAP5 (c.62G>T [p.Trp21Leu]) segregating with the disease in a second family. Functional analyses performed on both affected individual's cells and in vitro models showed that these two mutations caused pure or partial haploinsufficiency. Thus, alteration of MAGP-2, a component of microfibrils and elastic fibers, appears as an initiating mechanism of inherited TAAD.