Nanoparticles of perfluorocarbon emulsion contribute to the reduction of methemoglobin to oxyhemoglobin.

Here we show that methemoglobin is converted to oxyhemoglobin in the presence of perfluorocarbon (PFС) emulsion. Methemoglobin in blood at the level of above 30% can cause severe complications and lethal outcome. Some pharm chemicals in blood in vivo and in vitro can lead to oxidation of iron, Fe(2+)→Fe(3+), and to increased level of methemoglobin. The oxidized heme is not able to carry oxygen, hypoxia arises and irreversible changes are developing in vital organs. We added NaNO2 solution in different concentrations to blood in vitro in order to yield methemoglobin. Then the suspension of PFC nanoparticles was added. As methemoglobin interacted with PFC nanoparticles the optical density of peaks typical for oxyhemoglobin increased and spectral peak of methemoglobin decreased. The greater the concentration of PFC and the more was the incubation time, the more efficient was the process of reduction of methemoglobin to oxyhemoglobin. We proved experimentally that with an initial concentration of methemoglobin ​in average 95% the addition of nanoparticles of PFC decreases its concentration to 9% ​in average. At the same time the concentration of oxyhemoglobin increased in average from 5% to 81%.

Nanodefects of membranes cause destruction of packed red blood cells during long-term storage.

Packed red blood cells (PRBC) are used for blood transfusion. PRBC were stored for 30 days under 4 °Ð¡ in hermetic blood bags with CPD anticoagulant-preservative solution. Hematocrit was 50-55%. The distortions of PRBC membranes nanostructure and cells morphology during storage were studied by atomic force microscopy. Basic measurements were performed at the day 2, 6, 9, 16, 23 and 30 of storage and additionally 2-3 days after it. Topological defects occurred on RBC membranes by day 9. They appeared as domains with grain-like structures ("grains") sized up to 200 nm. These domains were appeared in almost all cells. Later these domains merged and formed large defects on cells. It was the formation of domains with the "grains" which was onset process leading eventually to destruction of PRBC. Possible mechanisms of transformation of PRBC and their membrane are related to the alterations of spectrin cytoskeleton. During this storage period potassium ions and lactat concentrations increased, pH decreased, intracellular concentration of reduced glutathione diminished in the preservative solution. Changes of PRBC morphology were detected within the entire period of PRBC storage. Discocytes predominated at the days 1 and 2. By day 30 PRBC transformed into irreversible echinocytes and spheroechinocytes. Study of defects of membranes nanostructure may form the basis of assessing the quality of the stored PRBC. This method may allow to work out the best recommendations for blood transfusion.

Transformation of membrane nanosurface of red blood cells under hemin action.

Hemin is the product of hemoglobin oxidation. Some diseases may lead to a formation of hemin. The accumulation of hemin causes destruction of red blood cells (RBC) membranes. In this study the process of development of topological defects of RBC membranes within the size range from nanoscale to microscale levels is shown. The formation of the grain-like structures in the membrane ("grains") with typical sizes of 120-200 nm was experimentally shown. The process of formation of "grains" was dependent on the hemin concentration and incubation time. The possible mechanism of membrane nanostructure alterations is proposed. The kinetic equations of formation and transformation of small and medium topological defects were analyzed. This research can be used to study the cell intoxication and analyze the action of various agents on RBC membranes.

Opposite effects of electroporation of red blood cell membranes under the influence of zinc ions.

The goal of the study was to investigate the effects of zinc ions of various concentrations on the nanostructure of membrane of red blood cells in in vitro experiment. The suspension of red blood cells extracted from whole human blood was used. The calibrated electroporation and the atomic force microscopy (AFM) were used to analyse damage to membrane nanostructure. We studied the haemolysis after the electroporation at different zinc concentrations. A low concentration of zinc (0.15-0.5 mM) increased significantly the rate of haemolysis and reduced the residual level of non-haemolyzed cells. At high concentrations of zinc ions (0.5-10 mM), the rate constant was sharply reduced, at the same time the residual level increased. The relationship between haemoglobin coagulants and the zinc concentration was examined. High concentration of zinc caused haemoglobin aggregation. It was shown by AFM that the membrane nanostructure was essentially changed. It was experimentally established that there existed a special point of zinc concentration C = 0.5±0.1 mM at which the course of the conjugate processes on the membranes of red blood cells was changed.