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Neeraj Prabhakar,1,2 Jixi Zhang,3 Diti Desai,1 Eudald Casals,1 Tina Gulin-Sarfraz,1 Tuomas Näreoja,2,4 Jukka Westermarck,5,6 Jessica M Rosenholm1 1Pharmaceutical Sciences Laboratory, Faculty of Science and Engineering, Åbo Akademi University, 2Laboratory of Biophysics, Faculty of Medicine, University of Turku, Turku, Finland; 3College of Bioengineering, Chongqing University, Chongqing, People’s Republic of China; 4Department of Neuroscience, Karolinska Institute, Stockholm, Sweden; 5Centre for Biotechnology, University of Turku and Åbo Akademi, 6Department of Pathology, University of Turku, Turku, Finland Abstract: Small interfering RNA (siRNA) is a highly potent drug in gene-based therapy with the challenge being to deliver it in a sustained manner. The combination of mesoporous silica nanoparticles (MSNs) and polycations in the confined pore space allows for incorporation and controlled release of therapeutic siRNA payloads. We hereby constructed MSNs with expanded mesopores and pore-surface-hyperbranched poly(ethyleneimine) (PEI) tethered with redox-cleavable linkers that could carry a high payload of siRNA (120 mg·g-1). The developed nanocarriers were efficiently taken up by cancer cells and were subsequently able to escape to the cytoplasm from the endosomes, most likely owing to the integrated PEI. Triggered by the intracellular redox conditions, the siRNA was sustainably released inside the cells over a period of several days. Functionality of siRNAs was demonstrated by using cell-killing siRNA as cargo. Despite not being the aim of the developed system, in vitro experiments using cell-killing siRNAs showed that the efficacy of siRNA transfection was comparable to the commercial in vitro transfection agent Lipofectamine. Consequently, the developed MSN-based delivery system offers a potential approach to hybrid nanocarriers for more efficient and long-term siRNA delivery and, in a longer perspective, in vivo gene silencing for RNA interference (RNAi) therapy. Keywords: mesoporous silica nanoparticles, RNAi therapy, siRNA delivery, stimuli-responsive drug release, hybrid nanocarriers

M Rosa Hernández1, Patricia Urbán2, Elisenda Casals3, Joan Estelrich3, Ginés Escolar1, Ana M Galán11Department of Hemotherapy and Hemostasis, Hospital Clinic, CDB, IDIBAPS, UB, Barcelona, Spain; 2Nanobioengineering Group, Institute for Bioengineering of Catalonia, Barcelona, Spain; 3Department of Physical Chemistry, Faculty of Pharmacy, University of Barcelona, SpainBackground: The contribution of fibrinogen (FBN) to hemostasis acting on platelet aggregation and clot formation is well established. It has been suggested that FBN-coated liposomes could be useful in restoring hemostasis. In the present study, we evaluated the modifications induced by multilamellar raw liposomes (MLV) or fibrinogen-coated liposomes (MLV-FBN) on hemostatic parameters.Materials and methods: Different experimental settings using whole blood or thrombocytopenic blood were used. Thromboelastometry, aggregation studies, platelet function analyzer (PFA-100®) tests and studies under flow conditions were applied to detect the effect of MLV-FBN on hemostatic parameters.Results: The presence of MLV-FBN in whole blood modified its viscoelastic properties, prolonging clot formation time (CFT) (226.5 ± 26.1 mm versus 124.1 ± 9.4 mm; P < 0.01) but reducing clot firmness (45.4 ± 1.8 mm versus 35.5 ± 2.3 mm; P < 0.05). Under thrombocytopenic conditions, FIBTEM analysis revealed that MLV-FBN shortened clotting time (CT) compared to MLV (153.3 ± 2.8 s versus 128.0 ± 4.6 s; P < 0.05). Addition of either liposome decreased fibrin formation on the subendothelium (MLV 8.1% ± 4.7% and MLV-FBN 0.8% ± 0.5% versus control 36.4% ± 6.7%; P < 0.01), whereas only MLV-FBN significantly reduced fibrin deposition in thrombocytopenic blood (14.4% ± 6.3% versus control 34.5% ± 5.2%; P < 0.05). MLV-FBN inhibited aggregation induced by arachidonic acid (52.1% ± 8.1% versus 88.0% ± 2.1% in control; P < 0.01) and ristocetin (40.3% ± 8.8% versus 94.3% ± 1.1%; P < 0.005), but it did not modify closure times in PFA-100® studies. In perfusion experiments using whole blood, MLV and MLV-FBN decreased the covered surface (13.25% ± 2.4% and 9.85% ± 2.41%, respectively, versus control 22.0% ± 2.0%; P < 0.01) and the percentage of large aggregates (8.4% ± 2.3% and 3.3% ± 1.01%, respectively, versus control 14.6% ± 1.8%; P < 0.01).Conclusion: Our results reveal that, in addition to the main contribution of fibrinogen to hemostasis, MLV-FBN inhibits platelet-mediated hemostasis and coagulation mechanisms.Keywords: thrombocytopenia, hemostasis, fibrinogen, liposomes, procoagulant activity, fibrin

This work deals with the short-time effective diffusion coefficient of charged and uncharged liposomes, measured (as a function of the volume fraction) using fiber optic dynamic light scattering. Particularly, we are interested in the interplay between electrostatic and hydrodynamic interactions on the diffusion of these lipid vesicles. Regarding the charged liposome, it has been found that the effective diffusion coefficient can be theoretically justified for volume fractions not exceeding certain critical value. In applying the theoretical approach, a surface charge has been obtained which is consistent with the electrokinetic characterization of the liposome. Regarding the uncharged liposome, the hard-sphere model seems to account for reasonably well the self-diffusion data. In addition, comparing the measurements of the short-time self-diffusion coefficient for both liposomes, we conclude that strong electrostatic forces (direct interactions) slow down diffusion processes. [ABSTRACT FROM AUTHOR]

Electrostatically stabilized aqueous dispersions of both slightly and highly charged liposomes have been analyzed by quasielastic light scattering. The obtained static structure factor S(q) revealed a liquidlike ordering due to long range electrostatic interactions. The fit of S(q) assuming a Derjaguin-Landau-Verwey-Overbeek interaction potential within the Debye-Hückel approximation, the Ornstein-Zemike equation and the hypernetted chain approximation, as a closure relation, allowed obtaining an effective charge value. Simultaneously, alternative charge values were estimated from electrophoretic measurements, turning out to be very close to the effective ones. This study also suggests that the charge seems to be unequally distributed over the vesicles in the case of liposomes formed by mixtures of charged and uncharged lipids. Size polydispersity was also considered. [ABSTRACT FROM AUTHOR]