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- ItemNanoparticles Can Wrap Epithelial Cell Membranes and Relocate Them Across the Epithelial Cell Layer(Washington, DC : ACS Publ., 2018-7-24) Urbančič, Iztok; Garvas, Maja; Kokot, Boštjan; Majaron, Hana; Umek, Polona; Cassidy, Hilary; Škarabot, Miha; Schneider, Falk; Galiani, Silvia; Arsov, Zoran; Koklic, Tilen; Matallanas, David; Čeh, Miran; Muševič, Igor; Eggeling, Christian; Štrancar, JanezAlthough the link between the inhalation of nanoparticles and cardiovascular disease is well established, the causal pathway between nanoparticle exposure and increased activity of blood coagulation factors remains unexplained. To initiate coagulation tissue factor bearing epithelial cell membranes should be exposed to blood, on the other side of the less than a micrometre thin air-blood barrier. For the inhaled nanoparticles to promote coagulation, they need to bind lung epithelial-cell membrane parts and relocate them into the blood. To assess this hypothesis, we use advanced microscopy and spectroscopy techniques to show that the nanoparticles wrap themselves with epithelial-cell membranes, leading to the membrane’s disruption. The membrane-wrapped nanoparticles are then observed to freely diffuse across the damaged epithelial cell layer relocating epithelial cell membrane parts over the epithelial layer. Proteomic analysis of the protein content in the nanoparticles wraps/corona finally reveals the presence of the coagulation-initiating factors, supporting the proposed causal link between the inhalation of nanoparticles and cardiovascular disease.
- ItemComplementary studies of lipid membrane dynamics using iSCAT and super-resolved fluorescence correlation spectroscopy(Bristol : IOP Publ., 2018) Reina, Francesco; Galiani, Silvia; Shrestha, Dilip; Sezgin, Erdinc; de Wit, Gabrielle; Cole, Daniel; Christoffer Lagerholm, B.; Kukura, Philipp; Eggeling, ChristianObservation techniques with high spatial and temporal resolution, such as single-particle tracking based on interferometric scattering (iSCAT) microscopy, and fluorescence correlation spectroscopy applied on a super-resolution STED microscope (STED-FCS), have revealed new insights of the molecular organization of membranes. While delivering complementary information, there are still distinct differences between these techniques, most prominently the use of fluorescent dye tagged probes for STED-FCS and a need for larger scattering gold nanoparticle tags for iSCAT. In this work, we have used lipid analogues tagged with a hybrid fluorescent tag–gold nanoparticle construct, to directly compare the results from STED-FCS and iSCAT measurements of phospholipid diffusion on a homogeneous supported lipid bilayer (SLB). These comparative measurements showed that while the mode of diffusion remained free, at least at the spatial (>40 nm) and temporal (50 ⩽ t ⩽ 100 ms) scales probed, the diffussion coefficient was reduced by 20- to 60-fold when tagging with 20 and 40 nm large gold particles as compared to when using dye tagged lipid analogues. These FCS measurements of hybrid fluorescent tag–gold nanoparticle labeled lipids also revealed that commercially supplied streptavidin-coated gold nanoparticles contain large quantities of free streptavidin. Finally, the values of apparent diffusion coefficients obtained by STED-FCS and iSCAT differed by a factor of 2–3 across the techniques, while relative differences in mobility between different species of lipid analogues considered were identical in both approaches. In conclusion, our experiments reveal that large and potentially cross-linking scattering tags introduce a significant slow-down in diffusion on SLBs but no additional bias, and our labeling approach creates a new way of exploiting complementary information from STED-FCS and iSCAT measurements.
- ItemZooming in on virus surface protein mobility(London : Future Medicine Ltd, 2018) Chojnacki, Jakub; Eggeling, Christian[no abstract available]