[A biotechnical system for diagnosis and treatment of binocular vision impairments].

Automation of the binocular vision biorhythm diagnosis and improvement of the efficacy of treatment of vision impairments are important medical problems. In authors' opinion, to solve these problems, it is necessary to take into account the correlation between the binocular vision and the electrical activity of the brain. A biotechnical system for diagnosis and treatment of binocular vision impairments was developed to implement diagnostic and treatment procedures based on the detection of this correlation.

["Stationary phase aging" of cell culture: an attempt of evaluation of growth medium "age" effect].

Cell proliferation rate and 3H-thymidine labeling index of "young" (i. e. harvested in 3 days after subcultivation) cultured Chinese hamster cells (B11 dii-FAF28 line) have been determined in growth medium conditioned by the same cells for various periods of time during their growth and subsequent "stationary phase aging" (medium of different "age"). Cells were serially cultured in Eagle's medium with 10 % bovine serum. The experiment was conducted as follows. The "young" cells were seeded in Carrel's flasks (4500 cells/cm2) with fresh growth medium and placed at 37 degreesC. At definite time intervals, media from 3 randomly selected flasks were filtrated and stored in small glass flasks at 4 degreesC. The cells from all 3 flasks were collected by trypsin treatment and counted with hemocytometer. During the period of 26 day cultivation we collected a set of media of different "age" corresponding to certain points of the growth and "stationary phase aging" curve of the culture. Then, the "young" cells in fresh medium were seeded into tissue culture plates with cover slips placed into wells of the plates (26,600 cells/cm2) and grown at 37degreesC, 5 % CO2 for 2 h. At this point, the medium was replaced with media of different "age". 22 h later (i. e. on the first day after seeding) cell density was evaluated microscopically in all the wells. On the next day (i. e. in 2 days after seeding) 3H-thymidine was added to every well to final concentration 1.85 x 10(4) Bq/ml. After next 24 h (i. e. in 3 days after seeding) cell density was counted again, and the medium was removed. The cover slips were rinsed with Hank's solution and air-dried. Autoradiography was performed in standard manner by photoemulsion exposing for 5 days and subsequent developing in amidol developer. The relative number of nuclei with 10 and more "grains" was revealed microscopically. Based on the obtained results, two basic parameters were evaluated for every "age" medium: 1) cell proliferation activity index calculated as log2 (N3/N1), where N1 - cell density on the first day after seeding, and N3 - the same parameter on the third day after seeding; 2) cell labeling index calculated as percentage of cells with nuclei labeled by 3H-thymidine during incubation from 2nd to 3rd day of cultivation. These two indexes for cell growth in different "age" media appeared to be highly correlating (R = 0.85). Besides, it was found that the observed "age-related" diminishing of ability of the growth media of different "age" to stimulate proliferation of "young" cells cannot completely explain the "stationary phase aging" phenomenon (in particular, even for the "oldest" medium cell labeling index was 65 %). We conclude that the phenomenon is based on exactly intrinsic changes of cells, most likely on molecular level, though environmental effects cannot be entirely excluded. The authors are grateful to the Russian Basic Research Foundation for support (grants 03-04-49030 and 00-04-48049).

[Study of protein products of gene expression in cells with altered chromosome sets for genetic mapping]. / Izuchenie belkovykh produktov gennoi ékspressii v kletkakh s izmenennym naborom khromosom dlia geneticheskogo kartirovaniia.

Two-dimensional electrophoresis was used for analyzing proteins in hybrid cells that contained single human chromosomes (chromosome 5, chromosome 21, or chromosomes 5 and 21) against the background of the mouse genome. By comparing the protein patterns of hybrid and parent cells (about 1000 protein fractions for each kind of cell), five fractions among proteins of hybrid cells were supposedly identified as human proteins. The genes of two of them are probably located on chromosome 5, and those of other three, on chromosome 21. Moreover, analysis of proteins in fibroblasts of patients with the cri-du-chat syndrome (5p-) revealed a decrease in the content of two proteins, as compared with those in preparations of diploid fibroblasts. This fact was regarded as evidence that two corresponding genes are located on the short arm of chromosome 5. Methodological problems associated with the use of protein pattern analysis in cells with altered chromosome sets for the purposes of genetic mapping are discussed.

[Erythrocyte membrane proteins in patients with hereditary spherocytosis]. / Belki membran éritrotsitov u bol'nykh nasledstvennym sferotsitozom.

Red blood cell membrane proteins were studied in a group of patients with hereditary spherocytosis, in comparison with normal donors, to reveal anomalous proteins associated with this disease. For this purpose red blood cells of the patients and normal donors were fractionated, by the age, in Ficoll-400 gradient, as a result red blood cell membranes were obtained with proteins that were investigated by the method of two-dimensional electrophoresis. In comparison of two-dimensional electrophoregrams of red blood cell membrane proteins of normal donors and those of microspherocytosis patients it was found that the latters had additional peptides in the area of glyceraldehyde-3-phosphate hydrogenase and pyruvate kinase. The changes detected in the red blood cell membrane protein composition might be caused by age shifts in the red blood cell population or by the disease type.

[Comparative study of human erythrocyte membranes in normal people and in Huntington's chorea patients]. / Sravnitel'noe izuchenie membran éritrotsitov krovi chekoveka v norme ibol'nykh khoreei gentingtona.

Erythrocytes of healthy volunteers and of patients with hereditary chorea were studied. Evaluation of the state of cellular membrane was carried out by measuring osmotic resistance, activity of Na+, K(+)-ATPase and protein composition. In the patients osmotic resistance of erythrocytes was distinctly decreased down to 66.3 +/- 3.3, while the Na+, K(+)-ATPase activity was increased 4-fold as compared with controls. Protein composition of erythrocyte membranes, studied by means of two-dimensional electrophoresis, was similar both in healthy persons and in patients with hereditary chorea when the electrophoretograms were analyzed visually. An additional protein with Mr = 30,000 and r-1-0.25 was detected in one of the patients. Slowly sedimenting fraction of erythrocytes was found in almost all the patients with hereditary chorea when erythrocytes aging was studied by means of fractionation in Ficoll density gradient. The fraction was not observed in healthy persons. These data suggest that the cell membranes in Huntington's chorea are altered as compared with normal state.

[Effect of digitonin on the activity of Na,K-ATPase from bovine brain]. / O vliianii digitonina na aktivnost' Na,K-ATPazy iz mozga byka.

The kinetic properties of intact and digitonin-treated Na,K-ATPase from bovine brain were studied. The temperature dependence curve for the rate of ATP hydrolysis under optimal conditions (upsilon 0) in the Arrhenius plots shows a break at 19-20 degrees. The temperature dependence curves for Km' and Km" have breaks at the same temperatures, while the Arrhenius plot for V is linear. The value of the Hill coefficient (nH) for ATP at 37 degrees is variable depending on ATP concentration, i. e. it is less than 1 at ATP concentrations below 50 mkM and is increased up to 3.2 at higher concentrations of the substrate. At high ATP concentrations the value of nH depends on temperature, falling down to 2.1 at 23 degrees and then down to 1 within the temperature range of 21-19 degrees. A further decrease in temperature does not significantly affect the nH value. Digitonin irreversibly inhibits Na, K-ATPase. ATP hydrolysis is more sensitive to the effect of the detergent than is nNPP hydrolysis, i. e. after complete inhibition of the ATPase about 40% of the phosphatase activity are retained. Treatment of Na,K-ATPase by digitonin results in elimination of the breaks in the Arrhenius plots for upsilon 0, Km' and Km", whereas the temperature dependence plot of V remains linear. Simultaneously digitonin eliminates the positive cooperativity of the enzyme for ATP. It is assumed that Na, K-ATPase from bovine brain is an oligomer of the (alpha beta) 4 type. Digitonin changes the type of interaction between the protomers within the oligomeric complex by changing the lipid environment of the enzyme or the type of protein -- lipid interactions.