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Abstract Background Massive industrial production of engineered nanoparticles poses questions about health risks to living beings. In order to understand the underlying mechanisms, we studied the effects of TiO2 and ZnO agglomerated engineered nanoparticles (EPs) on erythrocytes, platelet-rich plasma and on suspensions of giant unilamelar phospholipid vesicles. Results Washed erythrocytes, platelet-rich plasma and suspensions of giant unilamelar phospholipid vesicles were incubated with samples of EPs. These samples were observed by different microscopic techniques. We found that TiO2 and ZnO EPs adhered to the membrane of washed human and canine erythrocytes. TiO2 and ZnO EPs induced coalescence of human erythrocytes. Addition of TiO2 and ZnO EPs to platelet-rich plasma caused activation of human platelets after 24 hours and 3 hours, respectively, while in canine erythrocytes, activation of platelets due to ZnO EPs occurred already after 1 hour. To assess the effect of EPs on a representative sample of giant unilamelar phospholipid vesicles, analysis of the recorded populations was improved by applying the principles of statistical physics. TiO2 EPs did not induce any notable effect on giant unilamelar phospholipid vesicles within 50 minutes of incubation, while ZnO EPs induced a decrease in the number of giant unilamelar phospholipid vesicles that was statistically significant (p Conclusions These results indicate that TiO2 and ZnO EPs cause erythrocyte aggregation and could be potentially prothrombogenic, while ZnO could also cause membrane rupture.

Abstract Background Microvesicles isolated from body fluids are membrane - enclosed fragments of cell interior which carry information on the status of the organism. It is yet unclear how metabolism affects the number and composition of microvesicles in isolates from the peripheral blood. Aim To study the post - prandial effect on microvesicles in isolates from the peripheral blood of 21 healthy donors, in relation to blood cholesterol and blood glucose concentrations. Results The average number of microvesicles in the isolates increased 5 hours post - prandially by 52%; the increase was statistically significant (p = 0.01) with the power P = 0.68, while the average total blood cholesterol concentration, average low density lipoprotein cholesterol concentration (LDL-C) and average high density lipoprotein cholesterol concentration (HDL-C) all remained within 2% of their fasting values. We found an 11% increase in triglycerides (p = 0.12) and a 6% decrease in blood glucose (p < 0.01, P = 0.74). The post - prandial number of microvesicles negatively correlated with the post - fasting total cholesterol concentration (r = - 0.46, p = 0.035) while the difference in the number of microvesicles in the isolates between post - prandial and post - fasting states negatively correlated with the respective difference in blood glucose concentration (r = - 0.39, p = 0.05). Conclusions In a population of healthy human subjects the number of microvesicles in isolates from peripheral blood increased in the post - prandial state. The increase in the number of microvesicles was affected by the fasting concentration of cholesterol and correlated with the decrease in blood glucose.

In order to mount high-affinity antibody responses, B cells internalise specific antigens and process them into peptides loaded onto MHCII for presentation to T helper cells (TH cells). While the biochemical principles of antigen processing and MHCII loading have been well dissected, how the endosomal vesicle system is wired to enable these specific functions remains much less studied. Here, we performed a systematic microscopy-based analysis of antigen trafficking in B cells to reveal its route to the MHCII peptide-loading compartment (MIIC). Surprisingly, we detected fast targeting of internalised antigen into peripheral acidic compartments that possessed the hallmarks of the MIIC and also showed degradative capacity. In these vesicles, internalised antigen converged rapidly with membrane-derived MHCII and partially overlapped with cathepsin-S and H2-M, both required for peptide loading. These early compartments appeared heterogenous and atypical as they contained a mixture of both early and late endosomal markers, indicating a specialized endosomal route. Together, our data suggest that, in addition to in the previously reported perinuclear late endosomal MIICs, antigen processing and peptide loading could have already started in these specialized early peripheral acidic vesicles (eMIIC) to support fast peptide-MHCII presentation. [ABSTRACT FROM AUTHOR]

The effect of carbon black agglomerated nanomaterial on biophysical properties of canine red blood cell and platelet membranes that are reflected in changes of cell shape, was studied. Samples of canine blood diluted with citrated and phosphate buffered saline were incubated with carbon black nanomaterial and observed by scanning electron microscope and optical microscope. Interaction of agglomerated nanomaterial with erythrocyte membrane was observed. The relative abundance of different erythrocyte shape types (discocytes, echinocytes, spherically shaped erythrocytes) was determined on populations of cells, in suspensions with added carbon black nanomaterial and in control suspensions. Ensembles composed of representative images of cell populations were assessed by statistical methods. A two dimensional mathematical model of the erythrocyte shape was constructed to illustrate and explain erythrocyte swelling of intially discocytic/echinocytic shape to the final spherical shape, which preceeds membrane rupture. Micrometre-sized agglomerates were formed in the blood and interacted with erythrocyte membrane without evidently disturbing local membrane curvature or global cell shape. Incubation of blood with citrated and phosphate buffered saline caused a time dependent decrease of the number of intact erythrocytes in samples that was ascribed to a disintegration of erythrocyte membranes. The presence of carbon black nanomaterial in the samples suppressed this effect. Relative proportions of cell shape types remained largely unchanged within 24 hours of observation of the test and the control sample. The observed effects of carbon black nanomaterial can be described as originating from osmosis. Incubation of canine platelets with carbon black nanomaterial within 24 hours preserved the disc-like shape that is characteristic for resting platelets. It was concluded that carbon black nanomaterial interacts with membranes of blood cells but does not have a direct effect on the local or global membrane shape. However, large size of carbon black agglomerates, that can be formed in blood plasma could present mechanical obstacles in the cardiovascular system. [ABSTRACT FROM AUTHOR]

The formation of the immunological synapse upon B cell activation critically depends on the rearrangement of the submembranous actin cytoskeleton. Polymerization of actin monomers into filaments provides the force required for B cell spreading on the antigen-presenting cell (APC). Interestingly, the actin network also participates in cellular signaling at multiple levels. Fluorescence microscopy plays a critical role in furthering our understanding of the various functions of the cytoskeleton, and has become an important tool in the studies on B cell activation. The actin cytoskeleton can be tracked in live cells with various fluorescent probes binding to actin, or in fixed cells typically with phalloidin staining. Here, we present the usage of TIRF microscopy and an image analysis workflow for studying the overall density and organization of the actin network upon B cell spreading on antigen-coated glass, a widely used model system for the formation of the immunological synapse.