Sizing nanomatter in biological fluids by fluorescence single particle tracking

Accurate sizing of nanoparticles in biological media is important for drug delivery and biomedical imaging applications since size directly influences the nanoparticle processing and nanotoxicity in vivo. Using fluorescence single particle cracking we have succeeded for the first time in following the aggregation of drug delivery nanoparticles in real time in undiluted whole blood. We demonstrate that, by using a suitable surface functionalization, nanoparticle aggregation in the blood circulation is prevented to a large extent

Fluorescence single particle tracking for sizing of nanoparticles in undiluted biological fluids

While extremely relevant to many life science fields, such as biomedical diagnostics and drug delivery, studies on the size of nanoparticulate matter dispersed in biofluids are missing due to a lack of suitable methods. Here we report that fluorescence single particle tracking (fSPT) with maximum entropy analysis is the first technique suited for accurate sizing of nanoparticles dispersed in biofluids, such as whole blood. After a thorough validation, the fSPT sizing method was applied to liposomes that have been under investigation for decades as nanocarriers for drugs. The tendency of these liposomes to form aggregates in whole blood was tested in vitro and in vivo. In addition, we have demonstrated that the fSPT sizing technique can be used for identifying and sizing natural cell-derived microparticles directly in plasma. fSPT sizing opens up the possibility to systematically study the size and aggregation of endogenous or exogenous nanoparticles in biofluids

Matrix metalloproteinase-10 effectively reduces infarct size in experimental stroke by enhancing fibrinolysis via a thrombin-activatable fibrinolysis inhibitor-mediated mechanism

BACKGROUND: The fibrinolytic and matrix metalloproteinase (MMP) systems cooperate in thrombus dissolution and extracellular matrix proteolysis. The plasminogen/plasmin system activates MMPs, and some MMPs have been involved in the dissolution of fibrin by targeting fibrin(ogen) directly or by collaborating with plasmin. MMP-10 has been implicated in inflammatory/thrombotic processes and vascular integrity, but whether MMP-10 could have a profibrinolytic effect and represent a promising thrombolytic agent is unknown. METHODS AND RESULTS: The effect of MMP-10 on fibrinolysis was studied in vitro and in vivo, in MMP-10-null mice (Mmp10(-/-)), with the use of 2 different murine models of arterial thrombosis: laser-induced carotid injury and ischemic stroke. In vitro, we showed that MMP-10 was capable of enhancing tissue plasminogen activator-induced fibrinolysis via a thrombin-activatable fibrinolysis inhibitor inactivation-mediated mechanism. In vivo, delayed fibrinolysis observed after photochemical carotid injury in Mmp10(-/-) mice was reversed by active recombinant human MMP-10. In a thrombin-induced stroke model, the reperfusion and the infarct size in sham or tissue plasminogen activator-treated animals were severely impaired in Mmp10(-/-) mice. In this model, administration of active MMP-10 to wild-type animals significantly reduced blood reperfusion time and infarct size to the same extent as tissue plasminogen activator and was associated with shorter bleeding time and no intracranial hemorrhage. This effect was not observed in thrombin-activatable fibrinolysis inhibitor-deficient mice, suggesting thrombin-activatable fibrinolysis inhibitor inactivation as one of the mechanisms involved in the MMP-10 profibrinolytic effect. CONCLUSIONS: A novel profibrinolytic role for MMP-10 in experimental ischemic stroke is described, opening new pathways for innovative fibrinolytic strategies in arterial thrombosis

New methodological strategies in haematology using cell-derived microvesicles: New methodological strategies in haematology

International audienceCellular microvesicles are membrane nanometric vesicles, 0.1-1 µm in size, released into body fluids by activated cells or during apoptosis in a variety of pathological conditions. Microvesicles expose phosphatidylserine and tissue factor and have been proposed as pathogenic markers, key players of the haemostatic response. In recent years, tumour microvesicles have evolved as potential biomarkers. Indeed, tumour cells are able to constitutively release large amounts of microvesicles bearing tumour specific antigens. In hematologic malignancies the study of microvesicles is gaining increased interest and the generation of leukaemia/lymphoma cell-derived microvesicles may constitute a new tool for diagnosis and clinical/therapeutic follow-up

Protéolyse de la matrice extracellulaire et survie cellulaire

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Fibrinolysis, new concepts and new mechanisms: fibrinolytic microvesicles and fibrinolytic crosstalk: Fibrinolysis, new concepts and new mechanisms

International audienceThrombus lysis is the consequence of a restricted number of reactions localised to the surface of fibrin. A functional defect or an insufficient fibrinolytic response may lead to thrombosis with severe or fatal clinical consequences, e.g. myocardial infarction and ischemic stroke. Despite this clinical exigency and a real progress in the knowledge of the different components of this system (plasminogen and its activators, inhibitors and receptors), its functional evaluation still remains a challenge in haemostasis. The absolute requirement of a template for molecular assembly of plasminogen and its activators (tissue- and urokinase-type plasminogen activators: tPA and uPA) restricts the formation of plasmin and protects its activity onto the surface of, respectively, fibrin and cells. In contrast, plasmin and tPA released from the clot during its lysis are immediately neutralised by their respective inhibitors α2-antiplasmin and plasminogen activator inhibitor 1, PAI-1). It seems therefore almost impossible to detect fibrinolytic activity in plasma with methods currently in use. Because of its unavailability, it is also impossible to measure the degree of fibrinolysis directly on the clot. Notwithstanding, it was recently discovered that circulating membrane microvesicles might be indicators of the fibrinolytic response to an inflammatory or prothrombotic process. These cell-derived fibrinolytic microvesicles bear at their membrane the plasminogen activators expressed by the parent cell: tPA from endothelial cells and uPA from leukocytes. These molecules are localised at the membrane surface and have the capacity to activate plasminogen into plasmin in situ. Moreover, it was recently discovered that these microvesicles might participate in a new mechanism of plasmin formation requiring a cross-talk between two different surfaces. In this fibrinolytic cross-talk one of the surfaces bear plasminogen (fibrin, extracellular matrix or platelets) whereas the other surface carry the plasminogen activator, typically leukocyte-derived microvesicles bearing uPA. These new actors and concepts in plasminogen activation represent hitherto unknown pathways in our comprehension of fibrinolysis and potential novel biomarkers in clinical practice

Des microparticules cellulaires dévoilent leur fonction fibrinolytique et protéolytique

Des microparticules cellulaires dont la taille varie entre 0,1 et 1 μm sont émises par des cellules activées ou en apoptose. Elles portent, associées à leur membrane, des protéines indiquant leur origine cellulaire et de la phosphatidylsérine qui transforme le feuillet membranaire externe en surface d’assemblage des facteurs de la coagulation. Cette propriété procoagulante est accentuée par la présence de facteur tissulaire (FT) sur certaines microparticules. Une nouvelle fonction pro-fibrinolytique et protéolytique est maintenant décrite pour des microparticules d’origine tumorale ou endothéliale portant des métalloprotéinases matricielles et/ou des composants du système d’activation du plasminogène. Ces microparticules concentrent le plasminogène à leur surface où il est transformé en plasmine par l’urokinase (uPA, urokinase plasminogen activator) liée à son récepteur uPAR. La fibrinolyse, la migration cellulaire, l’angiogenèse, la dissémination tumorale, ainsi que l’apoptose induite par le détachement cellulaire pourraient ainsi être modifiées par la présence de ces microparticules chargées en plasmine

Formation de plasmine à la surface cellulaire (de la vésiculation à l'apoptose)

CAEN-BU Médecine pharmacie (141182102) / SudocSudocFranceF