Biochemistry (Moscow), Supplement Series A: Membrane and Cell Biology最新文献

In this work we have studied the effect of the phytoecdysteroid 20-hydroxyecdysone (20E) on the functioning of mouse skeletal muscle mitochondria. It is shown that 20E at a concentration of 100 µM or more suppresses mitochondrial respiration fueled by glutamate and malate (substrates of complex I of the respiratory chain) or succinate (substrate of complex II of the respiratory chain). This effect of 20E is accompanied by a decrease in the mitochondrial membrane potential and is associated with inhibition of the activity of complex III, the total activity of complexes I + III and II + III of the mitochondrial respiratory chain. We have noted a prooxidant effect of 20E, which manifests itself in an increase in the production of hydrogen peroxide by skeletal muscle mitochondria. In addition, 20E reduces the ability of mitochondria to accumulate calcium ions in the matrix. We discuss the mechanisms of the possible toxic effect of 20E on the functioning of skeletal muscle mitochondria.

Thrombin is a key enzyme of the blood coagulation system, which has been actively studied since the beginning of the last century. The formation of thrombin from prothrombin in the vicinity of vessel injury leads not only to the formation of fibrin—an important structural component of the hemostatic clot—but also to the activation of platelets, endothelium and immune system cells. The binding of thrombin to the platelet surface is thought to play a critical role in the process of platelet activation and may also ensure the maintenance of a high concentration of thrombin within the thrombus due to the concentration of protease on the platelet surface. Nowadays, all major thrombin receptors on platelets have been thoroughly characterized: through various experimental methods, the physicochemical parameters of the corresponding intermolecular interactions have been established. Since the interaction of thrombin with platelets leads to their activation, which includes changes in the number of receptors as a result of granule secretion, the interpretation of the observed kinetic binding curves faces a number of difficulties. It is known that as a result of platelet activation some receptors are able to redistribute on the membrane and form dimers and clusters, which makes the kinetics of thrombin binding to platelets an extremely complex process dependent on many factors, such as activator concentrations, platelet state, and other local parameters of the system. This review aims to describe the current understanding of the interaction of thrombin with the platelet membrane and to outline important unresolved issues in this area of research. The review provides not only information on structural and kinetic features of thrombin binding to individual platelet membrane proteins, but also analyzes the relationship between the relevant interaction parameters and previously obtained data on the integral kinetics of protease binding to the platelet surface.

The effect of He-Ne laser radiation on the composition and content of cell membrane components, namely phospholipids (PL) and sterols, in wheat callus tissues was studied by chromato-mass spectrometry and thin-layer chromatography. It was shown that irradiation of calluses with laser light at a dose of 3.6 J/cm² resulted in significant changes in the content of these components. Thus, the content of phosphatidylinositol increased in irradiated callus by 8 times, phosphatidylethonolamine by 2 times, and the content of phosphatidic acid decreased by 20% of the sum of PL. For sterols it was found that irradiation caused the most significant changes in the content of β-sitosterol dominant in plants (increase from 1453 ± 170 μg/g dry weight in unirradiated control to 2001 ± 111 μg/g dry weight 1 h after exposure); due to this, the total content of sterols also increased. The analysis of the obtained results suggests that PLs and sterols, primarily those for which regulatory and signaling functions are known, participate in the reaction of plant tissue to exposure to low-intensity He-Ne laser irradiation. This participation is realized as a stress (nonspecific) response to intense radiation.

Tropical teleost fish Danio rerio is increasingly used as a model object for electrophysiological studies of human cardiac physiology and pathology. D. rerio is characterized by the similarity with humans in such functional parameters of the electrical activity of the heart as heart rate, action potential morphology, as well as in a set of ion currents depolarizing and repolarizing the cell membrane. D. rerio is easy to breed, easy to handle experimentally, and easy to genetically modify. This overview presents current data on the structural and functional organization of ion channels in D. rerio heart myocytes.

Hydrogen sulfide (H₂S), which under physiological conditions exists in cells mainly in the form of the HS^– anion, is considered as a gaseous transmitter of inter- and intracellular signals along with nitrogen monoxide and carbon monoxide. Analysis of the dynamics of H₂S content in living cells is impossible without the creation of sensitive and specific probes. The group of K.H. Ahn synthesized several acedan-based compounds, which in the presence of H₂S attached a sulfhydryl group, forming fluorescent carbocyclic compounds. According to the spectral characteristics and reaction rate with H₂S, the optimal substance was P3, which forms the carbocyclic compound csP3 with the same large Stokes shift as P3 (approx. 130 nm) and has a brighter fluorescence. In this work, we tested the suitability of csP3 for recording changes in H₂S in solutions simulating the minimum salt composition of the intracellular medium, as well as in cells of primary neuronal culture from the rat cerebral cortex. It was found that the fluorescence intensity of csP3, which was formed when Na₂S (H₂S donor, 100 and 300 µM) was added to the P3 solution, differed for solutions corresponding in salt composition to the extracellular medium and cytosol. In both cases, fluorescence increased in the presence of bicarbonate (NaHCO₃, 10 mM). A decrease in the polarity of solutions due to the addition of dimethyl sulfoxide (30% by volume) shifted the emission by approx. 10 nm to the shorter wavelength region and doubled the intensity. Glutamate (10 µM, in the presence of 10 µM of glycine, 0 Mg²⁺) increased the fluorescence of the probe, but only in those neurons in which delayed deregulation of calcium homeostasis did not occur. The addition of P3 or csP3 to the cell culture caused a rapid increase in the fluorescent signal, which was replaced by a slow signal growth after 3–5 min. It was concluded that the product of the reaction of P3 with H₂S was sensitive to changes in the salt composition of the intracellular medium and could be redistributed in cells between an aqueous and more hydrophobic environment. These circumstances made it difficult to interpret the growth of P3 fluorescence in cells as a quantitative indicator of the presence of H₂S and required additional studies of the properties of this and structurally related H₂S probes.

Disulfiram (DSF) and its oxidized derivatives (DSFoxy) are currently being investigated as potential anticancer agents. We previously found that DSFoxy initiate the paraptosis-like death of tumor cells, which is of potential interest for the treatment of tumors resistant to the initiation of apoptosis. Based on bioinformatics analysis of mass spectrometric data on protein ubiquitination, we formulated a conception about the important role of disruption of the retrograde transport of damaged proteins from the endoplasmic reticulum to the cytosol in the mechanism of initiation of paraptosis-like cell death. In the present work, it has been found that DSFoxy, in the process of initiating paraptosis-like death of human adenocarcinoma HEp-2 cells, also enhances the ubiquitination of histones and histone code enzymes. In particular, this applies to the ubiquitination of histone H2BC12, histone methyltransferases responsible for transcription and repair of damaged DNA, as well as acetylating and ubiquitin-conjugating proteins. Bioinformatics analysis of changes in ubiquitination of cell nuclear proteins using the STRING database revealed during this process an increase in the occurrence of ubiquitinated proteins (functional enrichment) of cell cycle regulation, cell response to DNA damage and DNA repair, the regulation of which also depends on the histone code. This directly indicates damage to the cell nucleus and is consistent with confocal microscopy data. These results indicate that paraptosis-like death initiated by DSFoxy is accompanied, along with impairment of retrograde transport and ER stress, also by a change in regulation of the histone code, which points to a pleiotropic mechanism of cell death induction.

Calcium signaling is a principal method of signal transduction in cells of non-excitable tissues. In both mouse and human sperm, it can be induced in response to progesterone, manifesting as oscillations or single peaks and followed by the acrosomal reaction. However, the molecular mechanisms of progesterone activation may vary between species. In this study, we aim to compare the calcium signaling mechanisms in human and mouse spermatozoa. We investigated the calcium response in mouse sperm activated by progesterone. We employed spectrofluorometry to quantify the rise in calcium concentration in response to progesterone in Fura-2 loaded mouse sperm cells in suspension. Our experiments demonstrated that mouse sperm cells respond to 50 μM progesterone with a peak 120 ± 35 s wide and 0.8 ± 0.3 μM high. Based on literature data, a scheme for the induction of calcium signaling was constructed, suggesting an intermediate stage with the synthesis of a certain prostanoid (possibly PGE2) and activation of mouse sperm by this prostanoid through a G-protein-coupled receptor. Based on the obtained reaction scheme, two computational models were developed: a point model and a three-dimensional model. As with human sperm, the point model provided only a qualitative description of calcium responses, whereas the three-dimensional model produced the shape of the calcium peak and the frequency of calcium oscillations in response to progesterone that were similar to the experimentally obtained values. Using in silico analysis, it was shown that in mouse sperm, the spatial distribution of signaling enzymes regulates the type and form of the calcium response. We conclude that the presence of time delays due to the diffusion and spatial distribution of calcium signaling enzymes regulates the calcium response in both human and mouse sperm.

The effect of ionizing radiation (IR) on plants is mainly realized by altering the status of signaling systems and modifying stress signals. Variation potential (VP) is one of the types of electrical signals in plants. IR contributes to an increase in the amplitude of the VP, but the mechanisms of such influence are practically unknown. A possible way to implement changes arising from the action of IR is the regulation of gene expression. In the present work, the changes in the gene expression of participants in the generation and propagation of VP in irradiated plants are investigated. The experiments were performed on 14–15-day-old wheat plants (Triticum aestivum L.) grown under chronic irradiation (source ⁹⁰Sr–⁹⁰Y) with a dose rate of 31.3 μGy/h. The maximum accumulated dose was about 11.3 mGy. The irradiated plants showed no changes in the expression of calcium (TPC1), anionic (ALMT1 and CLC1), potassium (AKT1) channels, H⁺-ATPase (HA1), and NADPH oxidase (RBOHs) genes. A decrease in the expression of the SKOR potassium channel gene was revealed. The potassium channel blocker, tetraethylammonium chloride, caused an increase in response amplitude in control plants comparable to the increase in amplitude in the irradiated group. The obtained results indicate that one of the ways IR influences the electrical signals of plants is to inhibit the expression of the potassium channel.

The study of the permeability to water and urea of plasma membranes of endothelial cells of the normal cornea and cornea with a pronounced form of keratoconus was carried out. Human corneal endothelial cells were obtained from surgical material. Determination of osmotic water permeability (P_f) of normal and keratoconus endothelial cells did not reveal significant differences in the value of this parameter in both groups, where cells had a similar osmotic water permeability (control P_f = 0.53 ± 0.045 cm/s; keratoconus P_f = 0.63 ± 0.041 cm/s) (n = 25; p ≥ 0.05). The urea permeability coefficients (P_u) in both groups also had no statistically significant differences (control P_u = 0.049 ± 0.003 cm/s; keratoconus P_u = 0.056 ± 0.003 cm/s) (n = 25; p ≥ 0.05). Analysis of cell volume dynamics based on exponential approximation showed that corneal endothelial cells with keratoconus reduced their volume in a hypertonic medium to a greater extent compared to cells of a healthy cornea. An increase in cell volume as a result of isotonic urea inflow in a hypertonic medium also occurred to significantly higher values compared to normal cells. The conclusion was made about significant changes in the mechanism of regulation of the volume of corneal endothelial cells in keratoconus.