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A nanocarrier was obtained by coating the natural clinoptilolite particle surface with a phosphatidylcholine layer. The shell effective size does not exceed the thickness of the phospholipid molecular layer, which was confirmed by UV spectrometry and molecular mass spectrometry. The hydrodynamic diameter of the formulated nanocarrier, which was determined by dynamic light scattering, is smaller than the clinoptilolite core size. This effect is assumed to be caused by the phospholipid shell, which reduces the aqueous medium friction. The nanosize of the formulated nanocarrier, the natural clinoptilolite core, and the phospholipid shell together allow a combination of fruitful features that can be used for the fabrication of multifunctional platforms for the delivery of biologically active substances, bioimaging, or as a basis for biosensors., (© 2022. Pleiades Publishing, Ltd.)

Using a non-invasive approach, quantitative laser scanning microtomography (QLSM), the morphology of human oocyte was studied layer-by-layer. Then, the cell volume was computed based on 3D reconstruction of a stack of optical sections obtained by QLSM. The integrity of oocyte membrane after cryopreservation was assessed by measuring the changes in oocyte volume in response to hypotonic shock.

For in vitro fertilization technology, the quality of oocytes has a direct impact on the egg fertilization and developmental competence of early embryo. The morphological criteria are used for the estimation of oocyte quality before its fertilization in vitro. To date, only one method is known to determine the maturity of oocyte. This is the routine observation with a light microscope. The aim of this article was to adapt the noninvasive quantitative laser scanning microtomography (QLSM) for the investigation of morphological features of a human oocyte in vitro. This approach was used to accumulate the Z-stack gallery of optical sections of IVF oocyte. The layer-by-layer acquisition allows the fine cytoplasmic structure imaging. Applying the QLSM Z-stack of optical sections, the cellular volume was calculated with quantitative 3D reconstruction of a human oocyte. The volume value and intracellular structure were used as novel criteria to assess the oocyte state after the stress evoked by cryopreservation procedure.

The aim of this work was to study the fine structure of bacterial films grown on the inner tube surface of a flow reactor. Using the scanning electron microscopy (SEM) approaches, the detailed biofilm relief was visualized. The action of electrochemically reduced water (ERW) on the biofilm ultrastructure generated by the plankton form of E. coli and/or lacto bacteria was investigated. The treatment with an ERW solution destroyed the biofilm organic polymer matrix and bacterial cells embedded in the matrix.

The structure of bacterial film formed at the inner surface of the recirculation reactor tube, is studied. The surface relief of the biofilm was visualized by scanning electron microscopy. The effect of electrochemically activated water solution on the film formed from planktonic lactobacteria or E. coli was studied. Treatment with electrochemically activated water solution destroys cells and polymeric matrix of the biofilm.

Analysis of the element composition of oviduct and uterine fluid in mammals showed high potassium concentrations in the early embryo microenvironment in vivo. The results of early embryogenesis of mammals in vitro in the presence of high potassium concentrations are discussed. The data are summarized in accordance with the conditions of experimentally modeled pre-implantation development. Comparative assessment of the quality of embryo development until the blastocyst stage in vitro proved the embryos more successfully developed at potassium concentrations close to those registered in the oviductal fluid.

Regulatory volume decrease in response to hypotonic stress is typical of the oocytes and early mouse embryos. Changes in the kinetics of osmotic reaction can be used as a marker of the modulating effect of the incubation medium on transmembrane transport in embryonic cells. Quantitative laser scanning microtomography (QLSM) was used to measure oocyte volume. In this paper, it is shown that addition of 5 μM glycine, taurine, or GABA, as well as ATP to Dulbecco's medium abolished the regulatory volume decrease in mature mouse oocytes.

The inner surface of the drainage catheter used in surgical interventions for biliary system pathologies was examined by scanning electron microscopy. Microflora in the catheter lumen reflects etiological characteristics of the pathological process and helps to predict possible complications. The developed scanning electron microscopy imaging technique of visualization of the fi ne spatial structure of microbial biofilm formed on the catheter surface allows describing the cell pool and structure of the biofilm.

Electron probe microanalysis was applied to study the kinetics of changes in potassium and sodium concentration in muscle cells of isolated heart from Wistar rat during experimental ischemia. Hypoxic perfusion without glucose was shown to evoke the potassium deficiency and sodium accumulation in cardiac myocells. Short-term action (10 min) of strophanthin (0.1 mM/l) recovered Na/K balance in ischemic myocells. Hypothermic perfusion exhibited the opportunity to conserve the cytoplasmic elemental contents in the state corresponding to the beginning of low temperature (4 degrees C) operation.

The surface of wound dressing Biocol was studied by scanning electron microscopy. This composite system consists of latex matrix with incorporated water-soluble polysaccharide. The peculiarities of the surface are important for manufacturing of the dressing and for modification of its surface upon contact with fluids, e.g. during de novo tissue reconstruction. The method for studying the fine structure of the polymeric film surface was developed. The relief of the wound dressing changes during interaction with the fluid and nanopores appear on the surface. Thus, scanning electron microscopy is an informative method for studying the surface of biosynthetic films.

In the given investigation contents of potassium and its physiological analog, rubidium, are determined in cardiomyocyte. Applying Electron Probe Microanalysis (EPMA), cytoplasmic concentrations of elements (K, Rb) are measured. The data obtained exhibit that for initial acute ischemia phase the active transport is involved in the uptake of rubidium which competes with potassium entry in cardiac myocell. Then, deep deenergization leads to the intracellular potassium depletion and rubidium retention. This suggests that Rb+ is physiologically not complete analog for K+. Results of combined perfusion with and without rubidium allow us to hypothesize the appearance of cascade of ionic transports to compensate acute ischemic disturbances following the oxygen and substrate deficiency.

Mouse single-cell embryos exhibit robust Regulatory Volume Decrease (RVD). In what manner the very early mammalian embryo following zygote stage is appreciably altered by the anisotonic extracellular solution is, as yet, totally unclear. Little attention was paid to this direction since there was no way to determine the blastomere volume. This work has served to quantitatively investigate the osmotic response of bicellular mouse embryos employing Laser Scanning Microtomography (LSM) followed with three-dimensional reconstruction (3 DR). We have shown that bicellular mouse embryos in hypotonic Dulbecco's experience RVD. Embryonic cells subjected to hyposmolar exhibit rapid osmotic swelling followed by gradual shrinking back toward their original volume. The van't Hoff law defines swelling phase with the effective hydraulic conductivity of 0.3 micron x min(-1) x atm(-1). Water release during RVD in bicellular mouse embryos is abolished by Cytochalasin B (Cyto B) and the volume recovery is insensitive to ouabain treatment.

Osmolarity of Dulbecco's medium at which the volume of two-cell mouse embryo remained similar to that of intact embryo was determined. The method is based on comparison of kinetic curves describing the volume of embryonic cell in solutions of different osmolarity. The blastomere volume was measured by quantitative laser microtomography after fixed osmotic stress intervals. It was found that Dulbecco's saline with 125 mM NaCl solution is an isotonic solution for two-cell mouse embryo. This concentration corresponds to 290 mOsm, which is lower than osmolarity (~310 mOsm) of media routinely used for culturing of differentiated cells or biological fluids, e.g. blood plasma.

We performed qualitative comparison of freeze drying and chemical drying as methods of preparing 3D wet specimens for scanning electron microscopy. Human fibroblasts immobilized in collagen gel were used as a model system. Specimens fixed with glutaraldehyde were frozen in liquid nitrogen and freeze-dried at low temperature in high vacuum. In parallel experiments, glutaraldehyde-fixed samples were dehydrated in ascending ethanol solutions, absolute ethanol, and 100% hexamethyldisilazane and then dried at room temperature. Scanning electron microscopy microphotographs of collagen fibers and cells were characterized by high resolution and the absence of collapsed or deformed structures even at high magnification (×50,000) for both chemical drying and high-vacuum freeze drying. However, high-vacuum freeze drying is superior to chemical drying for the investigation of the internal space of 3D scaffolds, because sample fracture can be prepared directly in liquid nitrogen. These techniques are a part of the sample preparation process for scanning electron microscopy and can also be used for studies of cell adhesion, morphology, and arrangement in wet specimens (3D gels and flexible tissue engineering scaffolds).

Electron probe microanalysis was applied to determine cytoplasmic elemental (K, Na, Cl) concentrations in cardiac cells of the rat (Wistar). Potassium, sodium and chlorine contents were measured in papillary muscle myocytes of the rat heart perfused by the Langendorff's procedure. Ischemic depletion was created by perfusion with deeply deoxygenated Tirode's solution in the absence of glucose. It was found that the initial phase of acute ischemia is characterized by the potassium deficiency and the accumulation of sodium and chlorine in cardiac myocytes. It should be noted that changes in the total charge of the main intracellular cations (K+, Na+) do not compensate for the increased chlorine concentration. This result can be accounted for by the appearance of ionic (K+ and Cl-) transport coupled with the removal of lactate anions produced in cardiomyocytes during anaerobic glycolysis.

The blastomer volume was measured by three-dimensional (3D) reconstruction of a series of successive optical sections of the early mouse embryo. Changes in cell volume during osmotic shock were studied. Incubation of a two-cell embryo in Dulbecco's medium induced slow shrinkage of the embryonic cells followed by recovery of its initial volume. A regulatory phase of osmotic shock compensation is characteristic of a blastomer under hypotonic conditions.

The impact of the osmotic component of the incubation medium for the volume of mouse early embryonic cell was studied by laser scanning microscopy. Common Dulbecco's medium caused a prolonged hyperosmotic effect. Adaptive phase of regulatory compensation for the osmotic shock was observed under hypotonic conditions. From these data, water permeability of the blastomer membrane is evaluated as 0.4 micro/(minxatm).

Osmotic adaptation in a blastomere of mouse embryo has been studied by the direct measurement of the cell volume using laser scanning microscopy microtomography followed by quantitative 3D reconstruction. Embryonic cells subjected to hypotonic shock first swelled and then returned to the initial size. At the beginning of osmotic stress, the swelling occurred by the van't Hoff equation with the water permeability coefficient (L(p)) of 0.4 micro x min(-1) atm(-1). The phase of the regulatory volume decrease was not abolished by Na+/K(+)-ATPase inhibition.

The concentration of the elements (K, P) in the cytoplasm of rat (Wistar) cardiomyocytes has been determined by the electron probe analysis. The intracellular content of potassium and phosphorus was determined in early organogenesis and in unfertilized animals. In pregnant animals, a decrease in the cytoplasmic concentration of phosphorus and an increase in the concentration of potassium were registered. It was shown that the acute hypobaric hypoxia changes the content of both elements in the cytoplasm. The data obtained are discussed in terms of the activation of the ion transport system, which is aimed at compensating the cellular acidosis and lactosis induced by the development of hypoxic deenergization in pregnant animals.

The cell volume alteration participates in a wide variety of cellular functions that may interfere with intra-cellular homeostasis. The most adequate approach of estimation of the volume changes induced by osmotic misbalance, alteration in shape and size due to the action shape forming substances, etc., is the direct measurement of volumetric parameters of embryos. In the given research, the volume magnitude and kinetics of changes in volume and surface area of blastomere and polar bodies of early mouse embryos were determined using three-dimensional reconstruction of the optical section stack obtained with laser scanning microscope (LSM). The size and surface area were determined for isotonic and anisosmotic conditions. The physiological significance of the findings is discussed.

The concentrations of potassium and phosphorus in the cardiomyocyte cytoplasm of the Wistar rat were measured by means of electron probe microanalysis. Pregnancy was accompanied by a decrease in cytoplasmic phosphorus concentration and an increase in cytoplasmic potassium concentration. Acute hypoxia modified the concentrations of these elements in the cytoplasm.

Cytoplasmic Na/K imbalance induced by cytotoxic substances (ethylene glycol and cytochalasin B) was studied. Incubation in Dulbecco's medium with these cytotoxins caused changes in K and Na concentrations in the mouse two-cell embryo blastomer. The effects of both substances resulted in a drop of Na content in the embryonic cell. Washing from cytochalasin B restored the initial Na/K balance in the cytoplasm. Possible adaptive mechanisms involved in the regulation of intracellular ionic homeostasis are discussed.

Administration of substrates of energy metabolism in combination with hypoxen (sulfur-containing oligoquinone) promoted the increase in blood lactate concentration during maximum exercise, accelerated lactate utilization, and reduced the lactate/pyruvate ratio during recovery. The in vivo effects are in line with hypoxen capacity to accelerate in vitro oxidation of exogenous NADH in mitochondria by the non-rotenone-dependent pathway realized with participation of cytochrome C.

Menopausal transition is often accompanied by a variety of adverse pathological symptoms, currently treated with hormone replacement therapy, which is associated with a number of health risks. This report investigated the role of a food supplement--a composition of energy-exchange metabolites, with succinate as the main component--for treating menopausal syndrome. We studied the impact of a 4-week succinate-based food composition (SBC) treatment on the estral cycle, and bone mass and calcium content of aging mice. The impact of SBC on hormone levels and on the progression of several neurovegetative and psycho-emotional symptoms was further investigated in a randomized, double-blind, placebo-controlled clinical study of early menopausal women. Data were collected from questionnaires, Kupperman index scores, Spielberger-Hanin tests, and blood analysis of hormone levels taken at baseline and throughout the 5-week study. A "rejuvenating" effect of SBC on menopausal animals was observed, expressed as restoration of the estral cycle and an increase in the weight and calcium content of bone tissue. Furthermore, in the randomized, placebo-controlled clinical study in menopausal women, SBC-based monotherapy significantly lowered most subjectively evaluated characteristics of menopausal syndrome and increased blood serum levels of estradiol fourfold. This monotherapy also alleviated symptoms of some neurovegetative and psycho-emotional disorders, such as hot flushes, headache, and anxiety. Succinate-based therapy alleviated many biochemical symptoms of menopause in aging mice and early menopausal women, as well as neurovegetative and psycho-emotional disorders in women. Succinate-based therapy appeared to be free of adverse side effects.

In the companion paper (CryoLetters, 2007, 28:403-408), we reported effects of exposure of two-cell mouse embryos to vitrification solutions containing different vitrificants (EG, PG and DMSO) on the intracellular potassium and sodium content. We also compared exposure of 30% (v:v) ethylene glycol for 1.5 min to the similar experiments with 3-min exposure reported previously (CryoLetters, 2006, 27:87-98). In all experiments, four step protocols (2 steps of addition and 2 steps of washing) were used. Here we present mathematical modeling of the cell osmotic response using the relativistic permeability (RP) approach, which allows calculation of the osmotic curves without using simulation software but by direct calculations of the cell volume, intracellular concentration, and amount of the permeable vitrificants (Cryobiology, 2006, 53:402-3). Magnitude of the maximum cell volume excursion and other important osmotic characteristics were calculated for each step of the protocol, and the results of the mathematical modeling were superimposed onto the experimental data reported and discussed in the companion paper. The osmotic damage vs. specific chemical toxicity of the vitrificants as the major cause of the elemental disturbance of intracellular potassium and sodium content are discussed.

Intracellular concentrations of potassium and sodium in two-cell mouse embryos in G1/S phase after exposure to vitrification solutions containing vitrificant agents (VFAs) ethane-1,2-diol (ethylene glycol, EG), propane-1,2-diol (propylene glycol, PG), or dimethyl sulfoxide (DMSO), and sucrose (S) after incubation in Dulbecco solution were measured by electron probe microanalysis (EPMA) as described earlier (CryoLetters, 2006, 27: 87-98). The 4-step protocol was as followed: 10% VFA for 10 min => 30% VFA + 0.7 M S for 1.5 min ==> 0.5 M S for 10 min ==> 10 min pure Dulbecco's. The cytoplasmic concentration of potassium and sodium in immediately flashed out from the oviduct embryos was in range of 120 +/- 2 mM, with good concordance with the previous data (CryoLetters, 2006, 27:87-98). Exposure in Dulbecco's for 30 min did not alter elemental composition, neither did exposure in PG or DMSO for 1.5 min. In contrast, exposure for 1.5 min in EG dropped the level of potassium to 96 +/- 2 mM while elevating level of cytoplasmic sodium to 136 +/- 3 mM. Further exposure to 30%-EG for 3 min led to a two-fold decrease of both elements (60 +/- 3 mM and 66 +/- 2 mM for K and Na, respectively).

The 3D structure of the heart tissue of Wistar rats from different age groups has been reconstructed by light microscopy of consecutive series of semithin sections. At the histological resolution level, connective tissue segments in the myocardium of aged (18 months) animals were found, while in the myocardium of young (2 months) animals no connective tissue segments were observed. A mathematical model and the results of the 3D reconstruction were used to simulate the formation of excitation wave circulation in the myocardium. The values of conductivity and the excitability threshold, and the pulse frequency rates were found at which the disturbances of the heart rhythm resulting from the excitation wave circulation are predicted by the mathematical modeling.

This work was performed to study changes in intracellular elemental (Na/K) concentrations caused by Cytochalasin B in two-cell mouse embryo using Electron Probe Microanalysis. The presence of Cytochalasin B is required to transfer a somatic cell nuclear into an early embryo cell. The direct effect of this chemical is cytoskeleton transformation, which would be able to cause the increase of potassium channel activity resulting in cytoplasmic Na/K imbalance. In our study Cytochalasin B was shown to decrease the intracellular sodium concentration. The Na/K balance in the cytoplasm of mouse embryos reverted to its intact level after treatment them with Cytochalasin B free Dulbecco's solution. Possible mechanisms responsible for the changes in the intracellular sodium concentration observed in the embryo cells are discussed.

Age-related changes in the content of the major cellular cations of potassium and sodium in heart muscle cells of Wistar rats have been studied. The cytoplasmic concentration of potassium and sodium was determined by the electron probe microanalysis. The results revealed differences in both the concentration of the elements in young and old control animals and the responses of a cardiomyocyte to the state of acute hypoxic deenergization modelled on a perfused heart. The data are consistent with the hypothesis about the presence of genetically-related age changes in the conductance of potassium channels, which in old animals are realized against the background of deficient supply of tissues with oxygen and substrate.

Electron probe microanalysis is a method for evaluation of concentrations of elements at the subcellular level, effective in studies of an individual cell in culture or suspension. Intracellular concentrations of cytoplasmic ions (K+, Na+, Cl-) most rapidly and markedly reacting to changes is an adequate criterion for evaluating the traumatism of manipulations used in cell technologies. Using electron probe microanalysis it will be possible to develop special procedures (for example, therapeutic cloning) for reprogramming or cryopreservation, when intactness of intracellular balance of element serves as the criterion of cell status.

Intracellular concentration of potassium and sodium in two-cell mouse embryos in G1/S phase after exposition to vitrification solutions containing ethylene glycol (EG) and sucrose or after incubation in Dulbecco solution were measured by electron probe microanalysis (EPMA). The embryos at room temperature were treated in 10 percent EG for 10 min, transferred into mixture of EG and 1.0 M sucrose in ratio of 3:7 (v/v) for 3 min, then to 0.5 M sucrose for 10 min followed by washing the cells with Dulbecco;s solution for 10 min prior to analysis. The cytoplasmic concentration of potassium and sodium in controlled untreated with EG embryos were in a range of 116-130 mM of potassium and 120 mM of sodium, with good concordance in two identical experiments. After exposition that mimicked vitrification protocols, the intracellular potassium dropped almost two-three-fold (47 + 3 mM in one experiment and to 70 mM in the second experiment. The intracellular sodium concentration also decreased two-fold in range 60-70 mM after treatment with EG. Possible mechanisms of changes in the intracellular elemental concentrations including the high intracellular sodium observed in intact embryos are discussed.

The mean potassium and sodium concentrations and distribution of potassium in clonogenic culture of human neural stem cells (neurosphere) were estimated by means of electron probe microanalysis. High sodium concentration was typical of undifferentiated cells. Potassium was irregularly distributed in the test structure. Our results confirm published data on heterogeneous morphological structure of neurospheres.

Intracellular concentrations of potassium and phosphorus were determined by Electron Probe Microanalysis in mouse mature oocytes and zygotes. The oocytes were characterized by insignificant variations in the concentrations of these elements in the cytoplasm: 60 +/- 4 and 103 +/- 6 mM, respectively. In zygotes, on the contrary, significant variations were observed: 64 +/- 16 and 84 +/- 14 mM, respectively. Changes in the potassium homeostasis during the first cell cycle have been discussed.

Method of 3-D reconstruction approaches for early mouse embryo in preimplantation stages was modified. The developed technique is based on application of light microscopy of serial thin sections and well known soft operating. The designed method enabled us 1) to get serial sections of a single mouse embryo; 2) to create an orthogonal system independent on the sample for orientation of virtual sections. The adequacy of 3-DR protocol was checked on reconstruction of air bubbles embedded in epoxy resin as a model of sphere.

Comparative analysis of potassium concentrations in the cytoplasm of intact and enucleated one-cell mouse embryos of showed that microsurgical manipulations during collection of pronuclei disordered potassium homeostasis in the embryonic cell.

We propose a method of evaluation of anticlimacteric efficiency of a drug by its effect on the estrous cycle. The study was carried out on 9-month-old mice with retained, but notably reduced reproductive function. Analysis of the cell components of the estrous cycle was carried out on histological preparations of vaginal smears.

The need for effective methods of cryopreservation of early mammalian embryos necessitates the study of the mechanisms of blastomere adaptation to the equilibration procedure and subsequent washing from the cryoprotector. The osmotic effects during these procedures can cause electrolyte imbalance in embryonic cells. Intracellular potassium concentrations at the stage of two-blastomere mouse embryo were studied by electron probe microanalysis.

Direct effects of cardiotropic preparations on the hearts isolated from Wistar rats were examined. Deenergization of cardiomyocytes was modeled under conditions of hypoxic perfusion. Recovery of cardiac function during reperfusion was assessed by changes in the heart rate and contraction amplitude.

3-D structures were obtained at rotatory cultivation of CH immortalyzed human fibroblasts attached to glass microcarrier beads. The morphology of cells from these cultures was studied by scanning electron microscopy. A number of structural alterations in fibrillar filopodia of CH cells were revealed as compared with cells grown in stationary monolayer cultures, namely, smaller length, uneven caliber, the presence of curvatures, and disturbed branching pattern. Filopodia displayed unusual formations: protuberance-like and "mammoth's tusk"-like off-shoots, foamy spreadings in distal segments, and spiral windings of filopodia. The susceptibility of CH cells morphology to mechanical environment makes them a promising model for gravitational biology studies.

Electron probe microanalysis was employed to determine the elemental concentration (K,Na,Cl) in a myocyte on cryosections of the papillary muscle of the isolated rat (Wistar) heart. Protocols of global ischemia and ischemic conditions under glucose-free anoxic perfusion were applied. It was shown that global ischemia induces potassium deficiency (94 +/- 2 mM) in the myocyte and an increase in the level of sodium (72 +/- 4 mM) and chlorine (42 +/- 1 mM) in the cytoplasm compared with intact cell (122 +/- 2; 36 +/- 1; 24 +/- 1 mM). Glucose-free anoxic perfusion leads to a smooth fall of potassium concentration in the cell up to 54 +/- 2 mM with the retention of intracellular sodium (40 +/- 1 mM) and chlorine (26 +/- 1 mM) level. The present finding suggest that, in early ischemia, specific membrane mechanisms of ion transport are activated. Among these are KNa channel, Hi(+)-Nao+ exchange, KATP channel, lactate transport from the cell, associated either with potassium efflux to the extracellular space or chlorine influx into the myocyte. It is assumed that Na/K-ATPase is also activated under ischemic conditions.

X-ray electron probe microanalysis of a Neurospora crassa hyphal preparation revealed a significant heterogeneity of the potassium content in the hyphae. Low-potassium areas were found in a close proximity of the growing tip and in the distal part of the hyphae in the submembrane area of about 1 microm width. The intracellular potassium concentration was shown to change more than 3-5 times from apical to distal regions of the leading hyphae, the potassium contents in the apical part of the hyphae usually being minimal. We suggest that the content of intracellular potassium is determined by the volume occupied by free intracellular water, i.e., by the size of the space available for diffusion of substances, including inorganic ions.