Spontaneous level of sister chromosome exchanges and their distribution in human cells]

Blood of practically healthy donors of both sexes (27 females and 23 males) was cultured under the standard conditions during 96 hours. Bromodeoxyuridine (BUdR) was added at the final concentration of 10 mkg/ml 28 hours before harvesting. The slides were stained with acridine orange and Giemsa for differential staining of chromatids. In each culture sister chromatid exchanges (SCE) were analysed in 50 cells, and the part of cells undergoing the first, second and third mitoses at the time of harvesting, was calculated. According to the mean number of SCE per cell, the distribution of individuals was consistent with the normal law, the mean being 6.525 and standard deviation--0.956. A significant heterogeneity in the speed of cell cycle of cultures was observed. The coefficient of variation for the part of cells undergoing the first mitosis was 50%, for the cells in the second mitosis--15%, and for the cells in the third mitosis--154%. Correlation analysis showed a positive dependence of the mean level of SCF upon the age of a donor and upon the part of cells in the second mitosis in this individual. No reliable correlation of the SCE level with the donor's sex was observed. The distribution of cells, obtained from the culture of one individual, was best approximated by beta-distribution, and the distribution of cells obtained from the cultures of different individuals--by gamma-distribution. In both there was obtained a satisfactory approximation by Pearson's distribution of the 1 type, and significant deviations were found from the normal, Poison's and the negative binomial distribution. The conditions were found of similarity of empirical distribution of SCE in cells to the normal one. For that, it is not the value of SCE for a separate cell that should be used as a unit of measurement, but the mean from the values of frequencies for 5-10 cells. Hence, it was shown that for the evaluation of the mean frequency of SCE with the precision of 1 exchange in separate individuals it is necessary to analyse 40 cells, and to observe the 15% increase of spontaneous SCE level under the action of deleterious factors--8 individuals are enough to analyse.     

Selectivity characteristics of cation channels in Nitella flexilis plasmalemma

Ionic selectivity of Nitella flexilis plasmalemma cation channels is studied by voltage-clamp method with consecutive replacing of cations in the bathing medium. The selectivity sequence received by measuring the ionic current reversal potentials, psi alpha is: Ba++ approximately equal to Sr++ approximately equal to Ca++ greater than Mg++ greater than Cs+ approximately equal to K+ greater than Na+ greater than Li+. An analysis of results based on the three-barrier channel model suggests that when ions of the same valency are compared, the channel selectivity is determined by specific interactions between the ion and the nearest water molecules, which is possible both in a narrow and wide pore. On the other hand, when monovalent and divalent ions are compared the effects of ions binding in the channel or near the membrane surface prevail, thus causing the channel preference for divalent cations.     

Supramolecular structures of peptide assemblies in membranes by neutron off-plane scattering: method of analysis

In a previous paper (Yang et al., Biophys. J. 75:641-645, 1998), we showed a simple, efficient method of recording the diffraction patterns of supramolecular peptide assemblies in membranes where the samples were prepared in the form of oriented multilayers. Here we develop a method of analysis based on the diffraction theory of two-dimensional liquids. Gramicidin was used as a prototype model because its pore structure in membrane in known. At full hydration, the diffraction patterns of alamethicin and magainin are similar to gramicidin except in the scale of q (the momentum transfer of scattering), clearly indicating that both alamethicin and magainin form pores in membranes but of different sizes. When the hydration of the multilayer samples was decreased while the bilayers were still fluid, the in-plane positions of the membrane pores became correlated from one bilayer to the next. We believe that this is a new manifestation of the hydration force. The effect is most prominent in magainin patterns, which are used to demonstrate the method of analysis. When magainin samples were further dehydrated or cooled, the liquid-like diffraction turned into crystal-like patterns. This discovery points to the possibility of investigating the supramolecular structures with high-order diffraction.     

Changes in Sphingomyelinase Activity,Tumor Necrosis Factor α Level,and Lipid Peroxidation Intensity in the Course of Development of Cholestatic Liver Injury

Changes in sphingomyelinase activity, tumor necrosis factor α expression, and level of lipid peroxidation products in the course of development of cholestatic liver injury have been studied. The same type phase shifts in the analyzed parameters were observed, which included a marked decrease at the early stages of cholestasis (days 3–6) and a pronounced increase at the later stages (days 12–16), i.e., under the conditions of developed pathology. There was a significant positive linear correlation between tumor necrosis factor α expression, sphingomyelinase activity, and lipid peroxidation intensity in cholestatic injury. We propose that detected changes may reflect the balance between the effects of the two major bile components—bilirubin, which is accumulated in the liver at the early stages of cholestasis, and bile acids, whose influence dominates at the later stages of pathologic process. Our results indicate that tumor necrosis factor α overexpression, sphingomyelin cycle activation, and lipid peroxidation intensification may cause apoptosis of hepatocytes at the late stages of cholestasis.     

Coagulation factors and fibrinolytic system in subretinal liquid in patients with rheumatogenous retinal detachment

The work deals with estimation of some factors of blood coagulation and fibrinolytic systems, which include antithrombin III, factor X, prothrombin, plasminogen, protein C concentrations in the subretinal fluid of the patients with rhegmatogenous retinal detachment retina. The tendency to increase of the blood coagulation and fibrinolysis factor levels, except protein C, was revealed in the patients with complicated forms of the disease. The investigations mentioned above are capable of serving as a diagnostical and forecasting test characterizing the rhegmatogenous retinal detachment retina and surgical treatment proceeding.     

Cloning and characterization of SK2 channel from chicken short hair cells

In the inner ear of birds, as in mammals, reptiles and amphibians, acetylcholine released from efferent neurons inhibits hair cells via activation of an apamin-sensitive, calcium-dependent potassium current. The particular potassium channel involved in avian hair cell inhibition is unknown. In this study, we cloned a small-conductance, calcium-sensitive potassium channel (gSK2) from a chicken cochlear library. Using RT-PCR, we demonstrated the presence of gSK2 mRNA in cochlear hair cells. Electrophysiological studies on transfected HEK293 cells showed that gSK2 channels have a conductance of approximately 16 pS and a half-maximal calcium activation concentration of 0.74±0.17 M. The expressed channels were blocked by apamin (IC₅₀=73.3±5.0 pM) and d-tubocurarine (IC₅₀=7.6±1.0 M), but were insensitive to charybdotoxin. These characteristics are consistent with those reported for acetylcholine-induced potassium currents of isolated chicken hair cells, suggesting that gSK2 is involved in efferent inhibition of chicken inner ear. These findings imply that the molecular mechanisms of inhibition are conserved in hair cells of all vertebrates.     

In situ preparation of peptidylated polymers as ready-to-use adsorbents for rapid immunoaffinity chromatography

Summary The preparation method of peptide ligands employing polymer-supported solid-phase synthesis and leading to biospecific sorbents has been designed and optimized. This approach directly affords porous polymer sorbents for biospecific chromatography and avoids the cleavage of the synthesized peptide moieties from the carrier and their isolation. The specifics of both peptide synthesis and biospecific chromatography using hydrophilic macroporous polymer supports based on porous poly(glycidyl methacrylate-co-ethylene dimethacrylate) beads and discs were also investigated. The protecting groups can be removed from the target peptide (bradykinin) attached to the polymer support by trifluoromethylsulfonic acid without any significant loss of the attached peptide from the polymer carrier. Introduction of styrene as a comonomer into the copolymer structure improves the reactivity of the support. However, no nonspecific adsorption of proteins in the course of the biospecific isolation of antibradykinin antibodies was observed with these media. In contrast, the nonspecific sorption of proteins increases as a result of increasing peptide loading.     

Effect of the Pro-Gly-Pro peptide on resistance of rats to acute hypobaric hypoxia and posthypoxic behavioral disorders

Outbred white rats were subjected to acute hypobaric hypoxia in an altitude chamber at a reduced pressure of 145 mm Hg. Parameters of behavioral activity were evaluated in the tests of elevated cross maze and hole chamber and by the Porsolt method. Posthypoxic behavioral changes were characterized by an increase in the level of anxiety and depression and by a decrease in the level of orienting and exploratory activity. Preliminary intraperitoneal administration of the peptide 15 min before the hypoxia at a dose of 3.7 mol/kg increased the coefficient of individual resistance of the animals owing to a decrease in restitution time and significantly decreased posthypoxic behavioral disorders. We previously found that the Pro-Gly-Pro peptide had positive influence on homeostasis and maintained an adequate blood supply to organs and tissues. It was proposed that this ability of the Pro-Gly-Pro peptide could be one of the important mechanisms for its protective antihypoxic effect.