Injection of recombinant tissue plasminogen activator (rt-PA) is the standard drug treatment for thrombolysis. However, rt-PA shows risk of hemorrhages and limited efficiency even at high doses. Polysaccharide-poly(isobutylcyanoacrylate) nanoparticles functionalized with fucoidan and loaded with rt-PA were designed to accumulate on the thrombus. Fucoidan has a nanomolar affinity for the P-selectin expressed by activated platelets in the thrombus. Solid spherical fluorescent nanoparticles with a hydrodynamic diameter of 136 ± 4 nm were synthesized by redox radical emulsion polymerization. The clinical rt-PA formulation was successfully loaded by adsorption on aminated nanoparticles and able to be released in vitro. We validated the in vitro fibrinolytic activity and binding under flow to both recombinant P-selectin and activated platelet aggregates. The thrombolysis efficiency was demonstrated in a mouse model of venous thrombosis by monitoring the platelet density with intravital microscopy. This study supports the hypothesis that fucoidan-nanoparticles improve the rt-PA efficiency. This work establishes the proof-of-concept of fucoidan-based carriers for targeted thrombolysis.

The technique of differential dynamic microscopy is extended here, showing that it can provide a powerful and objective method of video analysis for optical microscopy videos of in vitro samples of live human bronchial epithelial ciliated cells. These cells are multiciliated, with motile cilia that play key physiological roles. It is shown that the ciliary beat frequency can be recovered to match conventional analysis, but in a fully automated fashion. Furthermore, it is shown that the properties of spatial and temporal coherence of cilia beat can be recovered and distinguished, and that if a collective traveling wave (the metachronal wave) is present, this has a distinct signature and its wavelength and direction can be measured.

Aim:We report the physicochemical analysis of nanosystems intended for cardiovascular applications and their toxicological characterization in static and dynamic cell culture conditions. Methods: Size, polydispersity and ζ-potential were determined in 10 nanoparticle systems including liposomes, lipid nanoparticles, polymeric and iron oxide nanoparticles. Nanoparticle effects on primary human endothelial cell viability were monitored using real-time cell analysis and live-cell microscopy in static conditions, and in a flow model of arterial bifurcations. Results & conclusions: The majority of tested nanosystems were well tolerated by endothelial cells up to the concentration of 100 μg/ml in static, and up to 400 μg/ml in dynamic conditions. Pilot experiments in a pig model showed that intravenous administration of liposomal nanoparticles did not evoke the hypersensitivity reaction. These findings are of importance for future clinical use of nanosystems intended for intravascular applications.

Thrombotic diseases rarely cause symptoms until advanced stage and sudden death. Thus, early detection of thrombus by a widely spread imaging modality can improve the prognosis and reduce mortality. Here, polymer microbubbles (MBs) made of degradable poly(IsoButylCyanoAcrylate) and functionalized with fucoidan (Fucoidan-MBs) were designed as a new targeted ultrasound contrast agent to image venous thrombus. The physicochemical characterizations demonstrate that the MBs with fucoidan surface exhibit a size of 2-6 μm and stability in suspension at 4°C up to 2 months. MBs exhibit high echogenicity and could be completely burst under high destructive pulse. Flow chamber experiments on activated human platelets show a higher affinity of Fucoidan-MBs than control anionic MBs (CM-Dextran-MBs) under shear stress conditions. In vivo analysis by ultrasound and histological results demonstrate that Fucoidan-MBs are localized in rat venous thrombotic wall, whereas few CM-Dextran-MBs are present. In addition, the binding of Fucoidan-MBs in healthy vein is not observed. Collectively, Fucoidan-MBs appear as a promising functionalized carrier for ultrasound molecular imaging in thrombotic diseases.

No abstract

Angiogenesis, the formation of blood vessels from pre-existing ones, is of vital importance during the early stages of bone healing. Extracellular stiffness plays an important role in regulating endothelial cell behavior and angiogenesis, but how this mechanical cue affects proliferation kinetics, gene regulation, and the expression of proteins implicated in angiogenesis and bone regeneration remains unclear. Using collagen-coated polyacrylamide (PAAm) hydrogels, human umbilical vein endothelial cells (HUVECs) are exposed to an environment that mimics the elastic properties of collagenous bone, and cellular proliferation and gene and protein expressions are assessed. The proliferation and gene expression of HUVECs are not differentially affected by culture on 3 or 30 kPa PAAm hydrogels, henceforth referred to as low and high stiffness gels, respectively. Although the proliferation and gene transcript levels remain unchanged, significant differences are found in the expressions of functional proteins and growth factors implicated both in the angiogenic and osteogenic processes. The down-regulation of the vascular endothelial growth factor receptor-2 protein with concomitant over-expression of caveolin-1, wingless-type 2, bone morphogenic protein 2, and basic fibroblast growth factor on the high stiffness PAAm hydrogel suggests that rigidity has a pro-angiogenic effect with inherent benefits for bone regeneration.

New tools for molecular imaging and targeted therapy for cardiovascular diseases are still required. Herein, biodegradable microcapsules (MCs) made of polycyanoacrylate and polysaccharide and functionalized with fucoidan (Fuco-MCs) are designed as new carriers to target arterial thrombi overexpressing P-selectin. Physicochemical characterizations demonstrated that microcapsules have a core-shell structure and that fucoidan is present onto the surface of Fuco-MCs. Furthermore, their sizes range from 2 to 6 µm and they are stable on storage over 30 d at 4 °C. Flow cytometry experiments evidenced the binding of Fuco-MCs for human activated platelets as compared to MCs (mean fluorescence intensity: 12 008 vs. 9, p < 0.001) and its absence for nonactivated platelets (432). An in vitro flow adhesion assay showed high specific binding efficiency of Fuco-MCs to P-selectin and to activated platelet aggregates under arterial shear stress conditions. Moreover, both types of microcapsules reveal excellent compatibility with 3T3 cells in cytotoxicity assay. One hour after intravenous injection of microcapsules, histological analysis revealed that Fuco-MCs are localized in the rat abdominal aortic aneurysm thrombotic wall and that the binding in the healthy aorta is low. In conclusion, these microcapsules appear as promising carriers for targeting of tissues characterized by P-selectin overexpression and for their molecular imaging or treatment.

This review highlights the preclinical and clinical research based on the use of nano- and micro-carriers in thrombolytic drug delivery. Ischemic heart and stroke caused by thrombosis are the main causes of death in the world. Because of their inactivation in the blood, high doses of thrombolytics are administered to patients, increasing the risk of intracranial hemorrhage. Preclinical research conducted with lipid, polymer or magnetic nanoparticles loaded with thrombolytic drugs showed an enhancement of thrombolysis and a reduction of undesirable side effects. Targeted nanocarriers exhibited an increased accumulation into clot. Clinical trials were already conducted with lipid-based microbubbles combined with ultrasound and thrombolytic drug and showed thrombolysis improvement. Future validation of nanosystems is awaited in clinic. This research opens new strategies for the management of thrombotic diseases.