Vitamin C was shown to partially protect red blood cells from oxidative changes during storage by noticeable reduction of mechanical fragility and hemolysis. In order to maintain the content of ascorbate in the reconstituted form in plasma, the latter is involved in a number of oxidative-reducing processes within red blood cells. This work is a continuation of studies of the effects of ascorbate on the metabolic processes that maintain the viability of red blood cells. Human red blood cells were incubated for five hours at 25 ºC in the oxidizing media system 1 1.0 · 10–4 M ascorbic acid (AscH), 5 · 10–6 M Cu2+, Na-phosphate buffer (0.015 M, pH 7.4), 0.15 M NaCl, and system 2, that contained o-phenanthroline at a concentration of 1.0 · 10–4 M in addition to the components of system 1 medium. For these cells, the changes in the content of reduced glutathione, glutathione enzyme activity, and the state of the membrane electron transport NADH: ferricyanide reductase were determined in time. The obtained data indicate that red blood cells undergo significant oxidative stress under the influence of the oxidative medium. During the first incubation period of erythrocytes in the AscH-Cu2+ environment, the activity of glutathione peroxidase and glutathione-S-transferase reached the maximum values, indicating the presence of H2O2 in the cell and the activation of lipid peroxidation processes. Glutathione-S-transferase activity remained above the control level throughout the entire study period. The activity of glutathione reductase and glucose-6-phosphate dehydrogenase was reduced. The oxidative loading of erythrocytes in the presence of o-phenanthroline was lower, the development of oxidative stress occurred in 90 minutes, but the binding of the o-phenanthroline complexes of Cu2+ to the membrane modified the SH-group of membrane proteins and this reduced the transport capabilities of the dehydroascorbate transporters and the electron transmembrane system, the consequence of which may be the accumulation of oxidized forms of ascorbate outside. We detected the participation of CO-signaling mechanism in hemoglobin deglutathionylation and increase in the content of glutathione. In this work we discuss the role of metabolic reprogramming in red blood cells through thiol-disulfide exchange as a mechanism that can be involved into adaptive responses aimed at counteracting stress in mammalian tissues.

One of commonly used approaches of biophysical modeling of muscle contractile apparatus is spatially explicit discrete lattice models in Monte Carlo simulation. Such models allow to reproduce structural features and actin-myosin interaction in the muscle contractile system more accurately. Limitation of such models is their low computational efficiency and stochasticity under certain circumstances. This work introduces deterministic approximation of stochastic model that considers a pair of rigid contractile filaments interaction. Approximation background is discreetness of spacing between cross-bridges and binding sites. Due to this property cross-bridges can be divided into discrete groups with the same strain, and considered statistically using the set of ordinary differential equations. Deterministic model is more computationally efficient, operates with average values. Within the given approach isotonic contraction was simulated. A comparison with Monte Carlo simulation demonstrates that approximation reproduces results for stochastic model with large number of cross-bridges. Also within the deterministic model a mechanism and essential conditions for oscillations appearance in isotonic transient response, relations of their parameters with geometrical ones of filaments lattice were examined, theoretical and experimental results were compared. The proposed approach can also be applied to approximation of continuous Huxley-based models solutions. Advantage over existing numerical methods is their greater numerical stability.

Background. Hemoglobin is a hemoprotein which in the presence of oxidative equivalents, such as H2O2, can act as peroxidase with a very high oxidative potential. Hemoglobin oxidation is accompanied by generation of highly oxidized forms of iron and globin radicals that have high oxidative activity and are toxic to cells. In addition, peroxidase activity may indicate structural changes that occur in the hemoglobin molecule as a result of chemical modification. Materials and Methods. Erythrocyte suspension was subjected to vibration for 3 h within the frequency range from 8 to 32 Hz with amplitudes of 0.5 ± 0.04 and 0.9 ± 0.08 mm. At certain intervals, hemoglobin peroxidase activity was determined together with the content of its ligand forms in the hemolysates of cells. Additionally, experiments were performed to investigate the mechanism and calculate the kinetic parameters of peroxidase reaction. Results and Discussion. Experimental data on low-frequency vibrations effect on erythrocyte hemoglobin peroxidase activity were analyzed. The kinetics of the oxidation reaction of p-phenylenediamine by hemoglobin in erythrocytes was studied. It was found that peroxidase oxidation has a ping-pong mechanism. The kinetic parameters of the peroxidase reaction involving hemoglobin were determined. The change of kinetic parameters after two-hour exposure to the incubation medium and low-frequency vibration was studied. A possible mechanism of action of hemoglobin in oxidation reactions involving H2O2 was proposed. Conclusion. Any effect that initiates the formation of methemoglobin leads to an increase in the peroxidase activity of hemoglobin due to the involvement of the latter in the pseudoperoxidase cycle and the formation of toxic reactive globin radicals. The high content of oxyhemoglobin in the cell, observed under vibrations within the frequency range of 16–32 Hz with an amplitude of 0.9 ± 0.08 mm, can prevent its oxidation and involvement in the pseudoperoxidase cycle.

The ability of protein to reversibly bind with membrane components is considered to be one of the oldest mechanisms of cell response to external stimuli. Erythrocytes have a well-developed mechanism of an adaptive response involving sorption-desorption processes, e.g. interactions of key glycolytic enzymes and hemoglobin with band 3 protein. A few publications have shown that under oxidative stress, cytoplasmic enzymes such as catalase, glutathione peroxidase and рeroxiredoxin bind to the erythrocyte membrane. The present work is a continuation of research in this direction to determine the causes and consequences of the interaction of cytoplasmic proteins with the membrane under conditions of oxidative stress and different glucose content. Human erythrocytes were incubated for five hours at 20 °C in an oxidizing medium of AscH – 1 · 10–4 M, Cu2+– 5 · 10–6 M with different glucose content (0–8 mM). Dynamic changes in the accumulation of membrane-bound hemoglobin, the distribution of ligand forms of hemoglobin in the cytoplasmic and membrane-bound fractions, the activity of membrane-associated and cytoplasmic forms of Cu/Zn superoxide dismutase (SOD1) and catalase, H2O2 content in extracellular and intracellular media were recorded. It was shown that binding of catalase and SOD1 to the erythrocyte membrane is initiated by oxidative stress and is a physiological function aimed at complete inactivation of extracellular and H2O2 and protection against their entry into the cell. It was shown that under conditions of glucose depletion and oxidative loading, catalase and SOD1 bind to the erythrocyte membrane, leading to inactivation of these enzymes. Membrane-bound hemoglobin was higher in cells incubated under these conditions than in glucose experiments. Glucose introduced into the incubation medium in an amount 4–8 mM causes complete binding of SOD1 to the membrane of erythrocytes, by involving it in the processes of casein kinase stabilization and glycolytic fluxes regulation. With mild oxidation, the amount of hemoglobin bound to the membrane does not change, indicating the presence of certain binding sites for hemoglobin with membrane proteins. We show that the activity of membrane-bound SOD1 along with the content of ligand forms in the composition of membrane-bound hemoglobin are informative indicators of the metabolic and redox state of erythrocytes.

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