In vitro and in vivo evaluation of a dextran-graft-polybutylmethacrylate copolymer coated on CoCr metallic stent

International audienceIntroduction: The major complications of stent implantation are restenosis and late stent thrombosis. PBMA polymers are used for stent coating because of their mechanical properties. We previously synthesized and characterized Dextrangraft-polybutylmethacrylate copolymer (Dex-PBMA) as a potential stent coating. In this study, we evaluated the haemocompatibility and biocompatibility properties of Dex-PBMA in vitro and in vivo.Methods: Here, we investigated: (1) the effectiveness of polymer coating under physiological conditions and its ability to release Tacrolimus®, (2) the capacity of Dex-PBMA to inhibit Staphylococcus aureus adhesion, (3) the thrombin generation and the human platelet adhesion in static and dynamic conditions, (4) thebiocompatibility properties in vitro on human endothelial colony forming cells (ECFC) and on mesenchymal stem cells (MSC) and in vivo in rat models, and (5) we implanted Dex-PBMA and Dex-PBMA TAC coated stents in neointimal hyperplasia restenosis rabbit model. Results: Dex-PBMA coating efficiently prevented bacterial adhesion and release Tacrolimus®. Dex-PBMA exhibit haemocompatibility properties under flow and ECFC and MSC compatibility. In vivo, no pathological foreign body reaction was observed neither after intramuscular nor intravascular aortic implantation. After Dex-PBMA and Dex-PBMATAC coated stents 30 days implantation in a restenosis rabbit model, an endothelial cell coverage was observed and the lumenpatency was preserved.Conclusion: Based on our findings, Dex-PBMA exhibited vascular compatibility and can potentially be used as a coating for metallic coronary stents

The mouse dorsal skinfold chamber as a model for the study of thrombolysis by intravital microscopy.: Thrombolysis imaging in the mouse skinfold chamber

International audienceAlthough intravital microscopy models of thrombosis in mice have contributed to dissect the mechanisms of thrombus formation and stability, they have not been well adapted to study long-term evolution of occlusive thrombi. Here, we assessed the suitability of the dorsal skinfold chamber (DSC) for the study of thrombolysis and testing of thrombolytic agents by intravital microscopy. We show that induction of FeCl3-induced occlusive thrombosis is achievable in microvessels of DSCs, and that thrombi formed in DSCs can be visualised by intravital microscopy using brightfield transmitted light, or fluorescent staining of thrombus components such as fibrinogen, platelets, leukocytes, and von Willebrand factor. Direct application of control saline or recombinant tissue-plasminogen activator (rtPA) to FeCl3-produced thrombi in DSCs did not affect thrombus size or induce recanalisation. However, in the presence of hirudin, rtPA treatment caused a rapid dose-dependent lysis of occlusive thrombi, resulting in recanalisation within 1 hour after treatment. Skin haemorrhage originating from vessels located inside and outside the FeCl3-injured area was also observed in DSCs of rtPA-treated mice. We further show that rtPA-induced thrombolysis was enhanced in plasminogen activator inhibitor-1-deficient (PAI-1-/-) mice, and dropped considerably as the time between occlusion and treatment application increased. Together, our results show that by allowing visualization and measurement of thrombus lysis and potential bleeding complications of thrombolytic treatments, the DSC provides a model for studying endogenous fibrinolysis and for first-line screening of thrombolytic agents. Furthermore, using this system, we found that thrombin and clot aging impair the thrombolytic action of rtPA towards FeCl3-produced thrombi

USPIO–PEG nanoparticles functionalized with a highly specific collagen-binding peptide: a step towards MRI diagnosis of fibrosis

International audienceFibrosis is characterized by a pathologic deposition of collagen I, leading to impaired function of organs. Tissue biopsy is the gold standard method for the diagnosis of fibrosis but this is an invasive procedure, subject to sampling errors. Several non-invasive techniques such as magnetic resonance imaging (MRI) using non-specific probes have been developed but they are not fully satisfying as they allow diagnosis at a late stage. In this study, collagelin, a collagen-binding peptide has been covalently linked using click chemistry to pegylated Ultra Small Super Paramagnetic Iron Oxide Nanoparticles (USPIO–PO–PEG–collagelin NPs) with the aim of diagnosing fibrosis at an early stage by MRI. USPIO–PO–PEG–collagelin NPs showed a high affinity for collagen I, two times higher than that of free collagelin whereas not peptide labeled USPIO NPs (USPIO–PO–PEG-yne) did not present any affinity. NPs were not toxic for macrophages and fibroblasts. Diffusion through collagen hydrogels concentrated at 3 and 10 mg mL−1 revealed a large accumulation of USPIO–PO–PEG–collagelin NPs within the collagen network after 72 hours, ca. 3 times larger than that of unlabeled USPIO, thereby evidencing the specific targeting of collagen I. Moreover, the quantity of USPIO–PO–PEG–collagelin NPs accumulated within hydrogels was proportional to the collagen concentration. Subsequently, the NPs diffusion through collagen hydrogels was monitored by MRI. The MRI T2 time relaxation decreased much more significantly with depth for USPIO–PO–PEG–collagelin NPs compared to unlabeled ones. Taken together, these results show that USPIO–PEG–collagelin NPs are promising as effective MRI nanotracers for molecular imaging of fibrosis at an early stage

Acute ischemic stroke thrombi have an outer shell that impairs fibrinolysis

Erratum inAcute ischemic stroke thrombi have an outer shell that impairs fibrinolysis. [Neurology. 2019]International audienceOBJECTIVES:Thrombi responsible for large vessel occlusion (LVO) in the setting of acute ischemic stroke (AIS) are characterized by a low recanalization rate after IV thrombolysis. To test whether AIS thrombi have inherent common features that limit their susceptibility to thrombolysis, we analyzed the composition and ultrastructural organization of AIS thrombi causing LVO.METHODS:A total of 199 endovascular thrombectomy-retrieved thrombi were analyzed by immunohistology and scanning electron microscopy (SEM) and subjected to ex vivo thrombolysis assay. The relationship between thrombus organization and thrombolysis resistance was further investigated in vitro using thrombus produced by recalcification of citrated whole blood.RESULTS:SEM and immunohistology analyses revealed that, although AIS thrombus composition and organization was highly heterogeneous, AIS thrombi shared a common remarkable structural feature in the form of an outer shell made of densely compacted thrombus components including fibrin, von Willebrand factor, and aggregated platelets. In vitro thrombosis experiments using human blood indicated that platelets were essential to the formation of the thrombus outer shell. Finally, in both AIS and in vitro thrombi, the thrombus outer shell showed a decreased susceptibility to tissue plasminogen activator-mediated thrombolysis as compared to the thrombus inner core.INTERPRETATION:Irrespective of their etiology and despite their heterogeneity, intracranial thrombi causing LVO have a core shell structure that influences their susceptibility to thrombolysis

Validation of the ISTH/SSC bleeding assessment tool for inherited platelet disorders: a communication from the Platelet Physiology SSC

Background: Careful assessment of bleeding history is the first step in the evaluation of patients with mild/moderate bleeding disorders, and the use of a bleeding assessment tool (BAT) is strongly encouraged. Although a few studies have assessed the utility of the ISTH-BAT in patients with inherited platelet function disorders (IPFD) none of them was sufficiently large to draw conclusions and/or included appropriate control groups. Objectives: The aim of the present study was to test the utility of the ISTH-BAT in a large cohort of patients with a well-defined diagnosis of inherited platelets disorder in comparison with two parallel cohorts, one of patients with type-1 von Willebrand disease (VWD-1) and one of healthy controls (HC). Patients/Methods: We enrolled 1098 subjects, 482 of whom had inherited platelet disorders (196 IPFD and 286 inherited platelet number disorders [IT]) from 17 countries. Results: IPFD patients had significantly higher bleeding score (BS; median 9) than VWD-1 patients (median 5), a higher number of hemorrhagic symptoms (4 versus 3), and higher percentage of patients with clinically relevant symptoms (score > 2). The ISTH-BAT showed excellent discrimination power between IPFD and HC (0.9 < area under the curve [AUC] < 1), moderate (0.7 < AUC < 0.9) between IPFD and VWD-1 and between IPFD and inherited thrombocytopenia (IT), while it was inaccurate (AUC ≤ 0.7) in discriminating IT from HC. Conclusions: The ISTH-BAT allows to efficiently discriminate IPFD from HC, while it has lower accuracy in distinguishing IPFD from VWD-1. Therefore, the ISTH-BAT appears useful for identifying subjects requiring laboratory evaluation for a suspected IPFD once VWD is preliminarily excluded.Fil: Gresele, Paolo. Università di Perugia; ItaliaFil: Orsini, Sara. Università di Perugia; ItaliaFil: Noris, Patrizia. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; Italia. Universita Degli Studi Di Pavia; ItaliaFil: Falcinelli, Emanuela. Università di Perugia; ItaliaFil: Alessi, Marie Christine. Centre For Cardiovascular And Nutrition Research; FranciaFil: Bury, Loredana. Università di Perugia; ItaliaFil: Borhany, Munira. No especifíca;Fil: Santoro, Cristina. Azienda Ospedaliera Universitaria Policlinico Umberto I; ItaliaFil: Glembotsky, Ana Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Cid, Ana Rosa. Hospital Universitario y Politecnico La Fe; EspañaFil: Tosetto, Alberto. No especifíca;Fil: De Candia, Erica. Fondazione Policlinico Agostino Gemelli; Italia. Università Cattolica del Sacro Cuore; ItaliaFil: Fontana, Pierre. University Hospitals of Geneva; SuizaFil: Guglielmini, Giuseppe. Università di Perugia; ItaliaFil: Pecci, Alessandro. Universita Degli Studi Di Pavia; ItaliaFil: Heller, Paula Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Médicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Médicas; ArgentinaFil: Rodorigo, Giuseppina. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Lammle, Bernhard. Università di Perugia; ItaliaFil: Trinchero, Alice. Università di Perugia; ItaliaFil: Paolo, Radossi. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Ferrari, Silvia. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Rancitelli, Davide. Università di Perugia; ItaliaFil: Stolinski, Amy. Università di Perugia; ItaliaFil: Arulselvan, Abinaya. Università di Perugia; ItaliaFil: Lassandro, Giuseppe. Università di Perugia; ItaliaFil: Sánchez Luceros, Analía Gabriela. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Medicina Experimental. Academia Nacional de Medicina de Buenos Aires. Instituto de Medicina Experimental; ArgentinaFil: Jandrot Perrus, Martine. Università di Perugia; ItaliaFil: Kunishima, Shinji. Università di Perugia; ItaliaFil: Rivera Pozo, José. Universidad de Murcia; EspañaFil: Lordkipanidzé, Marie. Università di Perugia; ItaliaFil: Melazzini, Federica. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Falaise, Céline. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Casonato, Alessandra. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Podda, Gianmarco. Università di Perugia; ItaliaFil: Kannan, Meganathan. Università di Perugia; ItaliaFil: Jurk, Kerstin. Università di Perugia; ItaliaFil: Sevivas, Teresa. Università di Perugia; ItaliaFil: Castaman, Giancarlo. Universidad de Bologna; ItaliaFil: Grandone, Elvira. Università di Perugia; ItaliaFil: Fiore, Mathieu. Università di Perugia; ItaliaFil: Zuniga, Pamela. Università di Perugia; ItaliaFil: Henskens, Yvonne. Università di Perugia; ItaliaFil: Miyazaki, Koji. Università di Perugia; ItaliaFil: Dupuis, Arnaud. Università di Perugia; ItaliaFil: Hayward, Catherine. Università di Perugia; ItaliaFil: Zaninetti, Carlo. Istituto Nazionale di Ricovero e Cura a Carattere Scientifico "Saverio de Bellis"; ItaliaFil: Abid, Madiha. Università di Perugia; ItaliaFil: Ferrara, Grazia. Università di Perugia; ItaliaFil: Mazzucconi, Maria Gabriella. Università di Perugia; ItaliaFil: Tagariello, Giuseppe. Università di Perugia; ItaliaFil: James, Paula. Università di Perugia; ItaliaFil: Fabris, Fabrizio. Universidad de Bologna; ItaliaFil: Russo, Alexandra. Università di Perugia; ItaliaFil: Bermejo, Nuria. Hospital de San Pedro de Alcantara, Cáceres; EspañaFil: Napolitano, Mariasanta. Università di Perugia; ItaliaFil: Curnow, Jennifer. Università di Perugia; ItaliaFil: Vasiliki, Gkalea. Università di Perugia; ItaliaFil: Zieger, Barbara. Università di Perugia; ItaliaFil: Fedor, Marian. Università di Perugia; ItaliaFil: Chitlur, Meera. Università di Perugia; ItaliaFil: Lambert, Michele. Università di Perugia; ItaliaFil: Barcella, Luca. Università di Perugia; ItaliaFil: Cosmi, Benilde. Università di Perugia; ItaliaFil: Giordano, Paola. Università di Perugia; ItaliaFil: Porri, Claudia. Università di Perugia; ItaliaFil: Eker, Ibrahim. Università di Perugia; ItaliaFil: Morel Kopp, Marie Christine. Università di Perugia; ItaliaFil: Deckmyn, Hans. Università di Perugia; ItaliaFil: Frelinger, Andrew L.. Università di Perugia; ItaliaFil: Harrison, Paul. Università di Perugia; ItaliaFil: Mezzano, Diego. Pontificia Universidad Católica de Chile; ChileFil: Mumford, Andrew D.. University of Bristol; Reino Unid