Chromatography analysis of seminal plasma proteins in buffalo semen samples with high and low cryotolerance.

The aim of this study was to analyse and identify specific buffalo seminal plasma proteins (SPPs) responsible for sperm cryotolerance during low temperature storage. Computer Assisted Sperm Analysis (CASA) of the motility and viability of buffalo spermatozoa was performed before freezing and after thawing. Two sample groups were formed - ejaculates with high cryotol- erance (group A) and low cryotolerance (group B). CASA demonstrated that the initial progres- sive motility after thawing of the spermatozoa in group A is significantly higher than in group B (p⟨0.001). Group B showed a significant increase in the percentage of static and non-progressive spermatozoa at 240 min, when compared to group A (p⟨0.05). SPPs, proteins in the cryoprotec- tive medium (PM) and proteins in the mixture of PM and SP were separated by High Perfor- mance Liquid Chromatography (HPLC). Comparative analysis of the chromatographic profiles was performed to identify specific proteins related to sperm cryotolerance. SPPs profiles showed 5 distinct protein peaks in both groups, ranging from 500 kDa to 50 Da. Chromatograms of group A and group B showed quantitative and qualitative differences in protein content. Chromato- grams of proteins in PM showed 11 well-expressed peaks. HPLC analysis of the mixtures of SPPs from the two groups and PM visualized the formation of a new bio-complex structure expressed by a protein peak specific for group A (7.674 min, AU 1.50). This protein peak can be referred as a phenotypic trait for buffalo ejaculates with high sperm cryotolerance.

Dataset illustrating the construction of dislocations and running dislocation dynamics using the periodic array of dislocation quadrupoles in FCC Al.

The data illustrates the process of creation of dislocation quadrupoles in FCC Al. The density of dislocations in our data is 3.8 × 1013 m-2 which is the lowest published to date and is comparable to the density of dislocations in real crystals. The data set can be found at the following URL:https://data.mendeley.com/datasets/mvmychgk4j/1. The detailed research article is "A topologically correct method of dislocations construction for atomistic modeling" (K. Yu. Khromov et al., 2019).

Transposition of Saccharomyces cerevisiae Ty1 retrotransposon is activated by improper cryopreservation.

Ty1 is a retrotransposon of the yeast Saccharomyces cerevisiae whose transposition at new locations in the host genome is activated by stress conditions, such as exposure to UV light, X-rays, nitrogen starvation. In this communication, we supply evidence that cooling for 2 h at +4 degrees C followed by freezing for 1 h at -10 degrees C and 16 h at -20 degrees C also increased Ty1 transposition. The mobility of Ty1 was induced by cooling at slow rates (3 degrees C/min) and the accumulation of trehalose inside cells or the cooling at high rates (100 degrees C/min) inhibited significantly the induction of the transposition. The freeze-induced Ty1 transposition did not occur in mitochondrial mutants (rho-) and in cells with disrupted SCO1 gene (Deltasco1 cells) evidencing that the Ty1 transposition induced by cooling depends on the mitochondrial oxidative phosphorylation. We also found that the freeze induced Ty1 transposition is associated with increased synthesis and accumulation of superoxide anions (O2-) into the cells. Accumulation of O2- and activation of Ty1 transposition were not observed after cooling of cells with compromised mitochondrial functions (rho-, Deltasco1), or in cells pretreated with O2- scavengers. It is concluded that (i) elevated levels of reactive oxygen species (ROS) have a key role in activation the transposition of Ty1 retrotransposon in yeast cells undergoing freezing and (ii) given the deleterious effect of increased ROS levels on cells, special precautions should be taken to avoid ROS production and accumulation during cryopreservation procedures.

Freeze drying of biomaterials for the medical practice.

Experimentally determined thermophysical characteristics of the new antifibrinolytic bacteriostatic biomedical preparation CAPROCOL are reported. Experimental and theoretical temperature histories at selected locations in a sample of CAPROCOL undergoing freeze drying are compared. The advance of the moving phase interface is also predicted.

Some seasonal changes in the thecal gland in the ovaries of hibernators (Citellus citellus L.).

The ovaries of 36 ground squirrels were studied in March, April, July, and December. Morphological and functional seasonal characteristics of theca interna were studied by histological methods, electron microscopy, and quantitative methods for determinating the volume density and histoenzyme activity of NADH2-tetrazolium reductase, glucose-6-phosphate dehydrogenase, and delta 5-3 beta-hydroxysteroid dehydrogenase. Maximal activity of theca interna as a steroid-producing structure was observed during spring awakening. A significant increase in the histoenzyme activity of delta 5-3 beta-hydroxysteroid dehydrogenase in December, versus two other enzymes studied, indicated steroid synthesis in theca terna.

[The HLA system in normal and pathological pregnancies]. / HLA-sistemata pri normalna i patologichna bremennost.

The authors investigated 205 women, 50 of whom were nonpregnant, 32 with normally progressing pregnancy, 30 parturients, 30 with abortions, 30 with prolonged pregnancy and 35 with pre-eclampsia. There were no statistically significant changes between the percentage of cases with positive lymphocytotoxic test, presence of leucoagglutinins and thrombocytic antibodies among the single groups. There were HLA-antibodies, which were not titrated due to the low titre in 4 of the women with pre-eclampsia.

Studies on the estrus cycle of the ground squirrel (Citellus citellus L.) at experimentally induced hibernation.

Eighty-five ground squirrels (C. citellus L.) were examined immediately after arousal (March-June). It was established that the animals were in estrus at the end of March; in April they were pregnant; in May they were in lactation; and in June they were already at rest, a postlactation period. Fifteen ground squirrels were treated with Hibernal and then placed in a chamber for 30 days at external temperature, 8-12 degrees C. In this case the estrus cycle of hypothermic ground squirrels was not changed. A retarded involution of corpus luteum persisted for a month and cystic follicles appeared in the ovary.

Seasonal changes of the ovarian atretic follicles of the ground squirrel (Citellus citellus L.).

The ovaries of 84 ground squirrels (C. Citellus L.) were studied during the four seasons of the year. The ovarian atretic follicles were examined by histological methods and by electron microscopy. The histoenzyme activities of NAD.H2-tetrazolium reductase, glucose-6-phosphate dehydrogenase, and 3 beta-hydroxy-delta 5-steroid dehydrogenase were photometrically demonstrated. The steroid-producing atretic follicles were mainly described as they reached their highest enzyme activity during the lactation period in April. The atresia of the primordial and primary follicles was manifested by disappearance of the oocyte and preservation of the granulose cells surrounded by basal lamina. Atresia of follicles with two oocytes was a typical process for the ground squirrel. Later on the oocytes and the granulose cells around it disappeared. The remaining part of the follicle continued its development and reached maturity. Atresia was observed mainly in March, April, and May.

[Acquisition of the sequence of a new subclass of human alpha- satellite DNA, localized in the centromere regions of two pairs of acrocentric chromosomes]. / Poluchenie posledovatel'nosti novogo podklassa al'fa-satellitnoi DNK cheloveka, lokalizovannoi v tsentromernykh raionakh dvukh par akrotsentricheskikh khromosom.

The results are presented in the paper on obtaining of a clone of sub-group of alpha-satellite human DNA localized in heterochromatin regions of two pairs of acrocentric chromosomes. The obtained repeated DNA sequence is represented in a genome by some 20,000 copies. Using of the blot-hybridization method made it possible to show the polymorphic restriction variant distribution being preserved in different individuals genomes. Intergenomic structural heterogeneity in this alpha-satellite DNA sub-group has been found only by using restriction endonuclease BamHI.

Effect of trehalose on the phase properties of hydrated and lyophilized dipalmitoylphosphatidylcholine multilayers.

The structure and thermal behavior of hydrated and lyophilized dipalmitoylphosphatidylcholine (DPPC) multilayers in the presence of trehalose were investigated by differential scanning calorimetry and X-ray diffraction methods. Trehalose enters the aqueous space between hydrated bilayers and increases the interbilayer separation (from 0.36 to 1.37 nm in the different DPPC phases at 1 M trehalose). It does not affect the lipid chain packing and also the slow isothermal conversion at 4 degrees C of the metastable L beta' phase into the equilibrium crystalline Lc phase. Addition of trehalose leads to a slight upward shift (about 1 degrees C at 1 M trehalose) of the three phase transitions (sub-, pre-, and main transition) in fully hydrated DPPC while their other properties (enthalpy, excess specific heat, and transition width) remain unchanged. The effect of trehalose on the thermal behavior of DPPC multilayers freeze-dried from an initially completely hydrated state is qualitatively similar to that of water. These data support the "water replacement" hypothesis about trehalose action. It is suggested that trehalose prevents the formation of direct interbilayer hydrogen bonds in states of low hydration.

The ovarium of the hibernator (Citellus citellus L.) after spring arousal.

The ovaries of 20 ground squirrels (Citellus citellus L.) were studied after spring arousal, while the animals were in the stage of estrus. The membrana granulosa of the preovulatory folic was luteinized. Epithelial cords were observed in the ovarial stroma. Cells in different stages of differentiation were found in the cords. For the first time nucleoli-like bodies in the nucleus and in the interstitial cells of the ovary of the ground squirrels. Probably, these nucleoli-like bodies played a definite role in the process of membrane formation during the cell differentiation. The NADH2 reductase and 3 beta-hydroxy-delta 5-steroid dehydrogenase activity confirmed that the process of differentiation of the interstitial cells was still in an initial stage. The findings were compared with the changes that took place during the puberty period.

Cytology of oocytes in ground squirrels (Citellus citellus L.).

Oocytes of primordial, primary, growing, and Graaf follicles of the ovaries of 96 ground squirrels (Citellus citellus L.) were studied during the four seasons of the year. Multilamellar bodies in the oocytes were observed by electron microscopy. The bodies underwent changes during the follicle growth. The vesicles included in the multilamellar bodies were filled with an amorphous substance, while in Graaf follicles trapezoidal, rhomboidal, and hexagonal inclusions were found. The primordial follicle of the oocyte had no Balbiani body. In the hibernating ground squirrel the functions of the Balbiani body were probably performed by the multilamellar bodies, which were considered to be analogous to the Balbiani body.

Changes in interstitial gland in the ovary of the bat Rinolaphus eurale Blas: arousal from hibernation.

The ovaries of 36 mature bats (Rinolaphus eurale Blas) were studied in March, June, and December. Quantitative determinations of the volume densities of the interstitial glands were studied. The histoenzymatic activities of NADH2 reductase and hydroxysteroid dehydrogenase showed the highest activity in the gland immediately after spring arousal.

Dependence of trehalose protective action on the initial phase state of dipalmitoylphosphatidylcholine bilayers.

Dipalmitoylphosphatidylcholine (DPPC) bilayers hydrated in the presence of trehalose were equilibrated at various temperatures (4, 20, and 60 degrees C) corresponding to the crystalline Lc, gel L beta', and liquid-crystalline L alpha phases, respectively, and then desiccated at these temperatures or freeze-dried at -80 degrees C to ca. DPPC dihydrate. The thermotropic behavior of the resulting DPPC/trehalose mixtures was investigated by differential scanning calorimetry and found to be dependent not only on the trehalose concentration but also on the phase state of the hydrated bilayers prior to their drying. Trehalose was most effective when the desiccation was carried out from the L alpha phase at 60 degrees C. In this case, one trehalose molecule per two DPPC molecules was sufficient to depress the melting temperature from values typical of DPPC dihydrate to 45 degrees C. Trehalose's influence decreased when dried from the L beta' phase and was significantly less pronounced when dried from the Lc phase. These data show that trehalose's protective influence depends on the initial phase state of the lipid bilayer and reaches its maximum in the liquid-crystalline state. The possible role of this effect in anhydrobiosis is pointed out.

Histoenzymatic activity of the delta 5-3 beta-hydroxysteroid dehydrogenase of ovary in hibernators (Citellus citellus L).

The ovaries of 24 ground squirrels (Citellus citellus L) were studied in spring (March, April), summer (July), and winter (December). The animals hibernated in a chamber at a temperature of 6-8 degrees C. The activities of the interstitial gland, theca interna, atretic follicles with theca interna and yellow bodies were measured densitometrically and studied by a specially modified histoenzymatic technique. Measurements showed that the endocrine structures were most active in March and least active in July. The atretic follicles had the highest enzymatic activity in April. The quantitative histoenzymatic approach presents an objective base for morphofunctional studies of organs and tissues at different body temperature levels in hibernators.

Activity of ATP synthetase complex after low temperature treatment or freeze-drying of mitochondria isolated from skeletal muscles.

The influence of freezing, thawing, or freeze-drying on ATP synthetase complex of isolated skeletal muscle mitochondria was studied. Cooling to -60 or to -196 degrees C and rapid thawing did not change activity significantly. Slow warming stimulated the release of latent ATP-ase activity and decreased ATP synthesis. These changes were more pronounced after freeze-drying.

Isolation and cryopreservation of human peripheral blood monocytes.

A modification of the Freundlich and Avdalovic method (J. Immunol. Methods 62, 31 (1983] is reported. Buffy coats, separated and pooled together, are used for isolation of monocytes (70% yield, 100% purity). Cell density of working suspension is increased up to 0.65 X 10(9) cells/75 cm2 surface by multiplication of the active fibronectin sites. For the purpose, cryoprecipitate is used instead of plasma for coating the glass-gelatin surface. Monocytes, isolated by that procedure, could be successfully cryopreserved with dimethyl sulfoxide cryoprotective solution.

A quantitative evaluation of the extent of inner mitochondrial membrane destruction after freezing-thawing based on functional studies.

An assay based on comparative investigation of ATPase and ATP synthetase changes in the activity in rat liver mitochondria after low temperature and uncoupler 2,4-dinitrophenol treatment is considered. By varying the activity of the respiratory chain, three extents of membrane cryoinjury could be distinguished by monitoring the changes in ATPase activity under different types of cryoinfluence. The first extent corresponds to a minimum membrane destruction, where the action of the respiratory chain compensates the changes in proton permeability and the cryotreatment does not change the ATPase activity. The second extent corresponds to changes in ion permeability which is partially compensated by the respiratory chain action. The third extent corresponds to a maximum membrane destruction and, therefore, maximum increase in permeability which is related to the irreversible stimulation of the ATPase activity and complete inhibition of the phosphorylation. In this study the extent of cryoinjury in mitochondrial preparations frozen and thawed at different rates was evaluated using this method.