Effect of vitamins A and E on the structural-functional state of retinal lysosomes

Vitamins A and E (alpha-tocopheryl-acetate and retinol-palmitate) are studied for their effect on structural and functional state of retina lysosomes. These vitamins are shown to exert a pronounced membrane-tropic effect on lysosomes. Vitamin E in chosen concentrations stabilizes membranes of retina lysosomes both in the in vitro and in vivo experiments. Vitamin A acts on them as a labilizing agent. The mentioned effect of vitamins may be corrected by low-energy radiation of the monochromatic coherent light (lambda = 632.8 nm). It is substantiated experimentally that the stabilizing effect of vitamin E may be intensified in case of the action combined with laser radiation on lysosomes. The labilizing effect of vitamin A on membranes of organelles under study may be weakened by application of laser radiation of low intensities.     

Optical radiation and the eyes with special emphasis on children

The Sun is the most abundant source of optical radiation for the child eye. New hand-held visible lasers are a threat to the child eye. Some scientific data suggest that near infrared radiation may cause cumulative damage in the ocular lens. The child eye usually is exposed to ambient solar radiation, gazing at the horizon. Ambient Sun ultraviolet radiation (UVR) exposure to the child is complex due to atmospheric scattering and strong dependence on background reflection. Solar exposure causes biological damage, only by photochemical mechanisms. UVR exposure to a child eye is mainly a threat to the anterior segment of the eye, but also age dependently to the retina. Above threshold exposure to UVR, for short delay onset of damage, causes a toxic reaction on the surface of the eye, snow blindness, and cataract. Sub-threshold daily exposure to UVR over decades is associated with several ocular surface pathologies and eye lid cancer. Visible radiation is a threat to the retina. A single above threshold exposure, for short delay onset of damage to the retina causes immediate photochemical Type II retinal damage, Sun blindness. A single exposure of the retina to a very high intensity laser beam may cause thermal or thermo-mechanical damage in the retina. In environments with high irradiance of optical radiation, the child eye should be protected. Legislation and public information is required for avoidance of damage from high intensity laser systems. More research is urgently needed to exclude the potential hazard of near infrared radiation.     

Overexpression of a novel gene , Cms1, can rescue the growth arrest of a Saccharomyces cerevisiae mcm10 suppressor

INTRODUCTIONDNA replication is a fundamenial process thatmust occur only once at each ce1l cycle. This restrictcontrol appears to be achieved through the coordi-nated actiVities of numerous proteins. The buddingyeast Saccharompes cerevhaae provides an excellenteukaryotic model fOr study of proteins invo1ved inthe control of DNA replication.In the budding yeast, minichromosome mainte-nance (MCM) proteins, MCM2-7, are a family of strsequence-related proteins that play crucia1 roles inr…     

Reflexotherapeutic effects on biologically active points of the concha auriculae

By means of histochemical method, for revealing cholinergic nervous structures, and electron microscopy, innervation of biologically active points (BAP) and that of neutral areas of the rabbit ear skin has been studied, normal and after acu-, electro- and electroacupuncture. The BAP have more intensive vascularization and innervation, their specific feature is presence of well pronounced nervous fasciculi. The latter are formed by 6-10 fibers 1-6 mcm in diameter. The diameter of the fasciculi is within the limits 25-30 mcm up to 40-45 mcm. Under the electron microscopic investigation myelin and amyelin fibers are revealed in the nervous fasciculi. In the area of the epidermal basal layer and in the epidermis itself, single nerve terminals are found; they are considered as the point pain receptors. After acu-, electro- and electroacupuncture, intensity of the nervous fibers staining increases, thus demonstrating an increment of acetylcholine esterase activity. After insertion of acupuncture needles and after electrical irritation, the changes in the nervous and cellular elements in the BAP areas are studied electron microscopically. After the effects mentioned, mast cells situating in the BAP area become, as a rule, degranulated. After repeated electroacupuncture irritation of the BAP, an inflammatory focus appears with peculiarities specific for the given process. The reflexotherapeutic effect is supposed to be transferred via the nervous system. The mechanical irritation of the nerve fasciculi and the accompanying traumatization of the surrounding cellular elements initiate the mechanism of reflexotherapy.     

Wave optical simulation of retinal images in laser safety evaluations

Lasers with wavelengths in the visible and near infrared region, pose a potential hazard to vision as the radiation can be focused on the retina. The laser safety standard IEC 60825–1:2014 provides limits and evaluation methods to perform a classification for such systems. An important parameter is the retinal spot size which is described by the angular subtense of the apparent source. In laser safety evaluations, the radiation is often described as a Gaussian beam and the image on the retina is calculated using the wave optical propagation through a thin lens. For coherent radiation, this method can be insufficient as the diffraction effects of the pupil aperture influence the retinal image. In this publication, we analyze these effects and propose a general analytical calculation method for the angular subtense. The proposed formula is validated for collimated and divergent Gaussian beams.     

Overexpression of a novel gene, Cms1, can rescue the growth arrest of a Saccharomyces cerevisiae mcm10 suppressor.

MCM10 protein is an essential replication factor involved in the initiation of DNA replication. A mcm10 mutant (mcm10-1) of budding yeast shows a growth arrest at 37 degrees C. In the present work, we have isolated a mcm10-1 suppressor strain, which grows at 37 degrees C. Interestingly, this mcm10-1 suppressor undergoes cell cycle arrest at 14 degrees C. A novel gene, YLR003c, is identified by high-copy complementation of this suppressor. We called it as Cms1 (Complementation of Mcm 10 Suppressor). Furthermore, the experiments of transformation show that cells of mcm10-1 suppressor with high-copy plasmid but not low-copy plasmid grow at 14 degrees C, indicating that overexpression of Cms1 can rescue the growth arrest of this mcm10 suppressor at non-permissive temperature. These results suggest that CMS1 protein may functionally interact with MCM10 protein and play a role in the regulation of DNA replication and cell cycle control.     

The phenotype of the minichromosome maintenance mutant mcm3 is characteristic of mutants defective in DNA replication.

MCM3 is an essential gene involved in the maintenance of minichromosomes in yeast cells. It encodes a protein of 971 amino acids that shows striking homology to the Mcm2 protein. We have mapped the mcm3-1 mutation of the left arm of chromosome V approximately 3 kb centromere proximal of anp1. The mcm3-1 mutant was found to be thermosensitive for growth. Under permissive growth conditions, it was defective in minichromosome maintenance in an autonomously replicating sequence-specific manner and showed an increase in chromosome loss and recombination. Under nonpermissive conditions, mcm3-1 exhibited a cell cycle arrest phenotype, arresting at the large-bud stage with an undivided nucleus that had a DNA content of nearly 2n. These phenotypes are consistent with incomplete replication of the genome of the mcm3-1 mutant, possibly as a result of limited replication initiation at selective autonomously replicating sequences leading to cell cycle arrest before mitosis. The phenotype exhibited by the mcm3 mutant is very similar to that of mcm2, suggesting that the Mcm2 and Mcm3 protein may play interacting roles in DNA replication.     

Phosphorylation of Mcm2 modulates Mcm2-7 activity and affects the cell's response to DNA damage

The S-phase kinase, DDK controls DNA replication through phosphorylation of the replicative helicase, Mcm2-7. We show that phosphorylation of Mcm2 at S164 and S170 is not essential for viability. However, the relevance of Mcm2 phosphorylation is demonstrated by the sensitivity of a strain containing alanine at these positions (mcm2(AA)) to methyl methanesulfonate (MMS) and caffeine. Consistent with a role for Mcm2 phosphorylation in response to DNA damage, the mcm2(AA) strain accumulates more RPA foci than wild type. An allele with the phosphomimetic mutations S164E and S170E (mcm2(EE)) suppresses the MMS and caffeine sensitivity caused by deficiencies in DDK function. In vitro, phosphorylation of Mcm2 or Mcm2(EE) reduces the helicase activity of Mcm2-7 while increasing DNA binding. The reduced helicase activity likely results from the increased DNA binding since relaxing DNA binding with salt restores helicase activity. The finding that the ATP site mutant mcm2(K549R) has higher DNA binding and less ATPase than mcm2(EE), but like mcm2(AA) results in drug sensitivity, supports a model whereby a specific range of Mcm2-7 activity is required in response to MMS and caffeine. We propose that phosphorylation of Mcm2 fine-tunes the activity of Mcm2-7, which in turn modulates DNA replication in response to DNA damage.     

Distribution of nerve fiber nodes with different cytochemical properties in bushy interoceptors

Distribution of generation zones of the action potential (AP) in the bushy interoceptors of the frog urinary bladder has been studied by means of D. C. Quick and S. G. Waxman cytochemical technique (1977). A positive reaction is obtained from the nodes of the neural fibers and a part of semi-nodi. The dependence of active nodi distribution on the length of the myelin branches is revealed. Three variants in distribution of precipitate are found in the divided fibers. If the distance between the semi-nodi is 60-350 mcm, the precipitate deposits more proximal to the node division. When the distance between the semi-nodi is 1,300-1,650 mcm, the active nodi are in both branches of the divided fiber. When the distance between the semi-nodi is 460-940 mcm, the active nodi are either in one branch only, or no branches give any positive reaction, and the precipitate deposits in the node of branching. Since the precipitate deposition characterizes, according to D. C. Quick and S. G. Waxman (1977) data, the AP generation places, it is possible to suppose that several impulse generators are present in one receptor, when the distance between its semi-nodi is not less than 1,300 mcm. A specific precipitate is supposed to deposit in the plates. It speaks in favour of a great density of transmembranous currents that determine transformation of a mechanical stimulus into a receptory potential.     

Specificity of the lethal and mutagenic actions of pico-second laser pulses of 532-nm wavelength

A study was made of the lethal effect of pulse laser (second harmonic Nd+3:YAG laser of 532 nm, pulse length 3.3.10(-11) s, peak intensity from 4.10(12) to 1.10(14) W/m2) on HeLa cells at the phases of active and stationary growth, and lethal and mutagenic effects of this radiation on E. coli cells. As was shown, HeLa cells at both growth phases and E. coli cells exhibited low sensitivity to laser radiation at lambda = 532 nm.     

Ultrastructure of the renal corpuscle of fresh water fishes

By means of light (semithin slices) and electron microscopes structural organization of the corpuscle and glomerulus of the capsule has been studied in freshwater fishes--carp (Cyprinus carpio) and crucian carp (Carassius carassius). The diameter of their glomeruli are in the carp from 80.8 +/- 1.4 mcm up to 59.8 +/- 2.4 mcm and in the crucian carp--from 86.4 +/- 1.8 mcm up to 69.25 +/- 0.2 mcm. Between these dimensions there are significant differences. Components of the external wall of the capsule in the fishes studied have similar structure. In epitheliocytes, situating on the basement membrane, electron opaque inclusions have been revealed. They resemble, by their ultrastructure, granules of peripolar cells, described in mammals and in man. In places, where epitheliocytes have contacts, thin folds are formed, they turn into the capsule lumen. Touching the cell surface, they can form border vacuoles. The main peculiarity of endotheliocytes in the glomerular capillaries is presence of diaphragmated and open fenestrae (their size is from 30 to 50 nm) in the peripheral zone of their cytoplasm. Unlike the higher vertebrates, the fishes have a well developed pericapillary mesangium. It contains mesangial matrix and rare processes of mesangiocytes.     

Mcm2 phosphorylation and the response to replicative stress

ABSTRACT: BACKGROUND: The replicative helicase in eukaryotic cells is comprised of minichromosome maintenance (Mcm) proteins 2 through 7 (Mcm2-7) and is a key target for regulation of cell proliferation. In addition, it is regulated in response to replicative stress. One of the protein kinases that targets Mcm2-7 is the Dbf4-dependent kinase Cdc7 (DDK). In a previous study, we showed that alanine mutations of the DDK phosphorylation sites at S164 and S170 in Saccharomyces cerevisiae Mcm2 result in sensitivity to caffeine and methyl methanesulfonate (MMS) leading us to suggest that DDK phosphorylation of Mcm2 is required in response to replicative stress. RESULTS: We show here that a strain with the mcm2 allele lacking DDK phosphorylation sites (mcm2AA) is also sensitive to the ribonucleotide reductase inhibitor, hydroxyurea (HU) and to the base analogue 5-fluorouracil (5-FU) but not the radiomimetic drug, phleomycin. We screened the budding yeast non-essential deletion collection for synthetic lethal interactions with mcm2AA and isolated deletions that include genes involved in the control of genome integrity and oxidative stress. In addition, the spontaneous mutation rate, as measured by mutations in CAN1, was increased in the mcm2AA strain compared to wild type, whereas with a phosphomimetic allele (mcm2EE) the mutation rate was decreased. These results led to the idea that the mcm2AA strain is unable to respond properly to DNA damage. We examined this by screening the deletion collection for suppressors of the caffeine sensitivity of mcm2AA. Deletions that decrease spontaneous DNA damage, increase homologous recombination or slow replication forks were isolated. Many of the suppressors of caffeine sensitivity suppressed other phenotypes of mcm2AA including sensitivity to genotoxic drugs, the increased frequency of cells with RPA foci and the increased mutation rate. CONCLUSIONS: Together these observations point to a role for DDK-mediated phosphorylation of Mcm2 in the response to replicative stress, including some forms of DNA damage. We suggest that phosphorylation of Mcm2 modulates Mcm2-7 activity resulting in the stabilization of replication forks in response to replicative stress.     

Replication of chromosomal DNA in Chinese hamster cells, cultivated at different temperatures

DNA fiber autoradiography was used to measure the rate of replication and the size of replication units in Chinese hamster cells cultured at 34, 37 and 39 degrees C. The average rate of DNA replication per single fork is 0.6 mcm/min at 34 degrees and 0.8 mcm/min at 37 and 39 degrees. In contrast to the change in the rate of DNA replication, no change was found in the size of replication units, which are more than 200 mcm at 31, 37 and 39 degrees. The change of the length of the S phase at various temperatures is determined only by the rate of DNA replication.     

Effect of infrared impulse laser irradiation on the development of albino rat embryos]

Development of the white rat embryos has been studied under effect of infrared laser beams (wave length--0.89 mcm, impulse frequency--300 Hz) during various periods of pregnancy. Exposure of the female rats for 256 and 128 sec produces an increased preimplantational death of embryos and certain disturbances in formation of the osseous skeleton in some embryos.     

Characterization of Schizosaccharomyces pombe mcm7(+) and cdc23(+) (MCM10) and interactions with replication checkpoints

MCM proteins are required for the proper regulation of DNA replication. We cloned fission yeast mcm7(+) and showed it is essential for viability; spores lacking mcm7(+) begin S phase later than wild-type cells and arrest with an apparent 2C DNA content. We isolated a novel temperature-sensitive allele, mcm7-98, and also characterized two temperature-sensitive alleles of the fission yeast homolog of MCM10, cdc23(+). mcm7-98 and both cdc23ts alleles arrest with damaged chromosomes and an S phase delay. We find that mcm7-98 is synthetically lethal with the other mcmts mutants but does not interact genetically with either cdc23ts allele. However, cdc23-M36 interacts with mcm4ts. Unlike other mcm mutants or cdc23, mcm7-98 is synthetically lethal with checkpoint mutants Deltacds1, Deltachk1, or Deltarad3, suggesting chromosomal defects even at permissive temperature. Mcm7p is a nuclear protein throughout the cell cycle, and its localization is dependent on the other MCM proteins. Our data suggest that the Mcm3p-Mcm5p dimer interacts with the Mcm4p-Mcm6p-Mcm7p core complex through Mcm7p.     

A genome-wide screen identifies genes required for formation of the wobble nucleoside 5-methoxycarbonylmethyl-2-thiouridine in Saccharomyces cerevisiae

We recently showed that the gamma-subunit of Kluyveromyces lactis killer toxin (gamma-toxin) is a tRNA endonuclease that cleaves tRNA(mcm5s2UUC Glu), tRNA(mcm5s2UUU Lys), and tRNA(mcm5s2UUG Gln) 3' of the wobble nucleoside 5-methoxycarbonylmethyl-2-thiouridine (mcm(5)s(2)U). The 5-methoxycarbonylmethyl (mcm(5)) side chain was important for efficient cleavage by gamma-toxin, and defects in mcm(5) side-chain synthesis correlated with resistance to gamma-toxin. Based on this correlation, a genome-wide screen was performed to identify gene products involved in the formation of the mcm(5) side chain. From a collection of 4826 homozygous diploid Saccharomyces cerevisiae strains, each with one nonessential gene deleted, 63 mutants resistant to Kluyveromyces lactis killer toxin were identified. Among these, eight were earlier identified to have a defect in formation of the mcm(5) side chain. Analysis of the remaining mutants and other known gamma-toxin resistant mutants revealed that sit4, kti14, and KTI5 mutants also have a defect in the formation of mcm(5). A mutant lacking two of the Sit4-associated proteins, Sap185 and Sap190, displays the same modification defect as a sit4-null mutant. Interestingly, several mutants were found to be defective in the synthesis of the 2-thio (s(2)) group of the mcm(5)s(2)U nucleoside. In addition to earlier described mutants, formation of the s(2) group was also abolished in urm1, uba4, and ncs2 mutants and decreased in the yor251c mutant. Like the absence of the mcm(5) side chain, the lack of the s(2) group renders tRNA(mcm5s2UUC Glu) less sensitive to gamma-toxin, reinforcing the importance of the wobble nucleoside mcm(5)s(2)U for tRNA cleavage by gamma-toxin.     

Formation of structuro-functional units of the microcirculatory bed of muscles in the postnatal period of the ontogenesis of the white rat

By means of biomicroscopy, histological methods and scanning electron microscopy topological relationships between the white rat m. cremaster microvessels and changes of their spatial organization have been studied in the postnatal development beginning from the 3d to the 14th week. In the terminal link of the microcirculatory bed zonal functional complexes of microvessels--myoangions are revealed. They represent rather autonomic and regularly repeating constructions of microvessels, having spatial regularity in respect to muscle fibers. With age the myoangion increases its stretchness from 303 +/- 14 in 3-week-old up to (70 +/- 3) X 10 mcm in 14-week-old animals. Capillary density in the m. cremaster during the development decreases from 354 +/- 12 up to 210 +/- 4 mcm, respectively. Distance between the longitudinal capillaries increases from 16.1 +/- 1.3 up to 31.2 +/- 1.8 mcm. Increase of transversal connections takes place in the capillary network, increasing number of the branching knots; this demonstrates certain complication of the nutritive link of the muscle microcirculatory bed. By the time of sexual maturation (6 weeks) there is a sharp decrease of the inflow and outflow coefficients. Higher meanings of the coefficient (0.70-0.74) are noted before sexual maturation period in comparison to that in mature animals (0.53-0.55).     

Differential requirement of DNA replication factors for subtelomeric ARS consensus sequence protosilencers in Saccharomyces cerevisiae

The establishment of silent chromatin requires passage through S-phase, but not DNA replication per se. Nevertheless, many proteins that affect silencing are bona fide DNA replication factors. It is not clear if mutations in these replication factors affect silencing directly or indirectly via deregulation of S-phase or DNA replication. Consequently, the relationship between DNA replication and silencing remains an issue of debate. Here we analyze the effect of mutations in DNA replication factors (mcm5-461, mcm5-1, orc2-1, orc5-1, cdc45-1, cdc6-1, and cdc7-1) on the silencing of a group of reporter constructs, which contain different combinations of "natural" subtelomeric elements. We show that the mcm5-461, mcm5-1, and orc2-1 mutations affect silencing through subtelomeric ARS consensus sequences (ACS), while cdc6-1 affects silencing independently of ACS. orc5-1, cdc45-1, and cdc7-1 affect silencing through ACS, but also show ACS-independent effects. We also demonstrate that isolated nontelomeric ACS do not recapitulate the same effects when inserted in the telomere. We propose a model that defines the modes of action of MCM5 and CDC6 in silencing.