[The polymorphism of DNA repair genes XRCC1, XRCC3 and the level of chromosomal aberrations in the Uranium workers].

Association between the polymorphism of DNA repair genes XRCC1 Arg399ln and XRCC3 Thr241Met and the frequency of chromosomal aberrations in the uranium workers was studied. The Gln/Gln genotype of gene XRCC1 was associated with a significant increase in the number of chromosomal aberrations as compared to the corresponding homozygous wild type Arg/Arg (p < 0.05). The frequency of chromosomal aberrations in heterozygous carriers of the XRCC3gene Thr/Met was lower than in the homozygous carriers of the wild type Thr/Thr (p < 0.001).

Integrity of mTORC2 is dependent on the rictor Gly-934 site.

Growth factor signaling coupled to activation of the phosphatidylinositol-3-OH kinase (PI3K)/Akt pathway plays a crucial role in the regulation of cell proliferation and survival. The key regulatory kinase of Akt has been identified as mammalian target of rapamycin complex 2 (mTORC2), which functions as the PI3K-dependent Ser-473 kinase of Akt. This kinase complex is assembled by mTOR and its essential components rictor, Sin1 and mLST8. The recent genetic screening study in Caenorhabditis elegans has linked a specific point mutation of rictor to an elevated storage of fatty acids that resembles the rictor deficiency phenotype. In our study, we show that in mammalian cells the analogous single rictor point mutation (G934E) prevents the binding of rictor to Sin1 and the assembly of mTORC2, but this mutation does not interfere with the binding of the rictor-interacting protein Protor. A substitution of the rictor Gly-934 residue to a charged amino acid prevents formation of the rictor/Sin1 heterodimer. The cells expressing the rictor G934E mutant remain deficient in the mTORC2 signaling, as detected by the reduced phosphorylation of Akt on Ser-473 and a low cell proliferation rate. Thus, although a full length of rictor is required to interact with its binding partner Sin1, a single amino acid of rictor Gly-934 controls its interaction with Sin1 and assembly of mTORC2.

Genetic polymorphisms and expression of minisatellite mutations in a 3-generation population around the Semipalatinsk nuclear explosion test-site, Kazakhstan.

We have reported previously that a population near the Semipalatinsk nuclear explosion test site had significantly increased minisatellite mutations (MM), suggesting increased germ-line mutation rates from the exposure in 3 generations. We hypothesize that the MM can be used as a surrogate biomarker for functional genetic alterations, e.g. gene mutations and chromosome aberrations. Therefore, we have investigated the influence of polymorphisms in genes on the expression of MM in the same two populations (247 and 172 individuals, for exposed and control, respectively, in 3 generations), and their relationships with radiation exposure. We have chosen the analyses of three polymorphic DNA - repair genes (XRCC1, XRCC1 and XRCC3) and two xenobiotic detoxification genes (GSTT1 and GSTM1). Among the exposed and in comparison with the wild-type gene, the functionally active XRCC1 Arg194Trp was significantly associated with low MM and over-represented in the exposed compared with the control populations. In a similar analysis, the functionally deficient XRCC1 Arg399Glu and XRCC3 Trp241Met were associated with increased and significantly reduced MM, respectively, but these variant genes were under-represented in the exposed population. Both GSTT1 and GSTM1 nulls were significantly associated with increased MM. The former was under-represented but the latter was significantly over-represented in the exposed compared with the control populations. In summary, the data indicate that the expected enzymatic functions of the polymorphic genes are consistent with the MM expression, except the XRCC1 Arg399Glu variant gene. In addition, the variant genes were retained in the three generations in association with their useful function, except for the GSTM1 null. However, the MM frequencies in the exposed were not consistently and significantly higher than those in the control populations, radiation exposure may therefore not have been the only cause for the high MM frequency among the exposed individuals. Since we studied three generations of citizens, the over- and under-representations of variant genes in the exposed population indicate their persistence and elimination, respectively, from the exposed individuals, suggesting their functional influence on survivability. The latter observation also indicates the complexity of gene and environmental interactions, e.g. the GSTM1 null was significantly over-represented in the exposed population.

[Mutagenic effect of the rocket fuel component asymmetric dimethylhydrazine on rats of various ages].

Mutagenic effect of asymmetric dimethylhydrazine (ADMH) on rats of different age groups upon acute and subacute treatment and protective effect of a Limonium gmelinii preparation. Genotoxic effect of ADMH depending on the dose and duration of treatment was established. The phytopreparation lacked mutagenicity and toxicity and had a protective effect in combination with the xenobiotic.

[Role of different isoforms of nitric oxide synthase in development of tumor mutants in Drosophila melanogaster]. / Rol' razlichnykh izoform sintazy okisi azota v razvitii opukholevykh mutantov Drosophila melanogaster.

We studied the role of nitric oxide synthase during tumor growth in oncovirus-induced tumor mutants of Drosophila melanogaster. The lines with different capacity for malignancy differed reliably in the level of enzymatic activity. It was shown using specific inhibitors of neuronal and inducible isoforms that the neuronal isoform was not involved in tumor formation, while the inducible one appears to play an important role in tumor growth inhibition. This isoform was identified with the help of immunoblotting and monoclonal antibodies against inducible nitric oxide synthase.

Effect of nitric oxide on apoptotic activity in the rat gastrointestinal tract.

The effect of nitric oxide (NO) on apoptosis in the gastrointestinal mucosa was investigated. Experiments involved long-term exposure of rat gastric mucosal cells in vitro to exogenous NO delivered from the NO, donor S-nitroso-N-acetyl-penicillamine, and the effect of intravenous administration of lipopolysaccharide in vivo, in the presence and absence of the selective inhibitor of inducible NO synthase N-(3-(aminomethyl)benzyl) acetamidine (1400 W). S-nitroso-N-acetyl-penicillamine produced a dose-related inhibition of caspase 3-like activity and DNA fragmentation in isolated gastric mucosal cells. Caspase 3-like activity and DNA fragmentation in gastric, ileal and colonic mucosa were increased both 5 and 24 h after injection of lipopolysaccharide (3 mg/kg, i.v.) to rats in vivo. Administration of 1400 W (5 mg/kg, i.v.) immediately after lipopolysaccharide enhanced caspase 3-like activity and DNA fragmentation above that found with lipopolysaccharide alone. In conclusion, data obtained both in vitro and in vivo suggest that NO exerts an anti-apoptotic effect on rat gastrointestinal mucosal cells.

[Chromosomal localization of a gamete-killing gene in a wheat line]. / Khromosomnaia lokalizatsiia gametotsidnogo gena v linii pshenitsy.

Monosomic analysis revealed that a gametocidal gene in a common wheat line derivative from Aegilops speltoides is located on chromosome 2B. This gene induces semisterility, low level of chromosome breaks, shriveled seeds, and endosperm degeneration. The obtained data indicate that the localized gene is allelic to well known gametocidal genes Gc1a and Gc1b.

[Oncovirus-induced permanent genetic instability in Drosophila melanogaster]. / Onkovirus-indutsirovannaia prodlennaia geneticheskaia nestabil'nost' u Drosophila melanogaster.

Mutant alleles of a system of genetic instability induced by oncoviral DNAs were shown to demonstrate an unstable manifestation 500 generations after their emergence. A cytogenetic analysis of oncovirus-induced unstable lines has revealed numerous chromosome rearrangements. For the Lobe alleles of this system, a specific chromosome rearrangement, Df(2L) = 35C-36B, was found on the left arm of chromosome 2. We used recessive lethal mutations involving DNA rearrangements in a successful construction of cross systems for "explosive" instability.

[Cellular mechanisms of postnatal growth of rat liver in during chronic exposure of cadmium sulfate and strontium chloride]. / Kletochnye mekhanizmy postnatal'nogo rosta pecheni krys pri khronicheskom vozdeistvii sul'fata kadmiia i khlorida strontsiia.

A cytophotometric investigation was performed to study the ploidy level and total protein content in hepatocytes of rats of different ages (1, 7, 14, 21, 30, 90, 180, 365 days), both intact and chronically treated with cadmium sulfate or strontium chloride. It was established that during the first month of postnatal ontogenesis, compositions of liver parenchyma cell population of intact and treated rats did not differ. Compared to control animals, the process of cell polyploidization in the liver of rats treated with heavy metal salts of 30-90 days proceeded slower, especially in Cd(2+)-treated rats. Within 180-365 days the cell polyploidization in the treated animals increased. The proportion of (4c x 2)-hepatocytes in 1 year old Cd(2+)- or Sr(2+)-treated rats increased, resp., by 2.7 and 1.5 times, and that of 8c hepatocytes was higher by 3.9 and 1.5 times than in the control, the average ploidy level rising by 20 and 5%. respectively. It was established that until 90 days the rate of protein accumulation in liver cells of intoxicated rats was slower than in intact animals. Thus, the average protein content per diploid hepatocyte in Cd(2+)- or Sr(2+)-treated 30 day old rats was lower by 20 and 16%, respectively, compared to control animals. The protein content increased in liver cells of Cd(2+)- or Sr(2+)-intoxicated rats following 90 and 180 days, respectively, and this process was exclusively associated with cell polyploidization. During the first 3 weeks after birth, no significant difference was observed in the extent of involvement of cell proliferation, polyploidization and hypertrophy in the growth of liver in intact and intoxicated animals. At this period the liver was growing due completely to cell proliferation and hypertrophy. During 21-30 days the contribution of cell proliferation to the liver growth of intact rats was not significant (29%), whereas it remained at higher level (50%) in the treated animals. In 30-90 days after birth, the involvement of proliferation process to the liver growth of intoxicated rats decreased to 25-28%, while in intact animals it increased up to 37%. At this period the cell polyploidization plays an essential role in the growth of liver in both intact and intoxicated animals to reach in average 37-46%. The contribution of polyploidization and hypertrophy to the liver growth of Cd(2+)-treated rats within 30-90 days was obviously higher than in Sr(2+)-treated animals. Both at the late (3-12 months) and at the early (1-21 days) stages of experiments, the pattern of correlation of different cell components in the growing liver of intact and intoxicated rats differed only a little.

Effects of prostaglandins on [Ca2+]i and adenylate cyclase activity in isolated porcine gastric mucous cells.

In porcine gastric mucous cells, isolated enzymatically from the fundic mucosa and enriched by counterflow centrifugation, PGE2 (1 microM) increased adenylate cyclase activity to 225% and, distinct from that documented for other species, also [Ca2+]i, measured fluorimetrically with Fura2/AM, in Ca(2+)-containing and Ca(2+)-free incubation medium to 182% and 165% of control values, respectively. PGF2 alpha, PGD2, the stable prostacyclin analogue iloprost and the thromboxane-mimetic U46619 had no significant effects on adenylate cyclase activity and [Ca2+]i. Histamine (10 microM) stimulated adenylate cyclase activity to 236% of control value, an effect which could be blocked by the H2-receptor antagonist ranitidine. However, histamine and the activators of the cAMP system forskolin and dibutyryl cAMP had no significant effect on [Ca2+]i, indicating that an activation of the adenylate cyclase/cAMP system per se does not result in an increase in [Ca2+]i. These data suggest that prostanoids stimulate adenylate cyclase activity and [Ca2+]i in gastric mucous cells via activation of EP-receptors linked to both second messenger systems.

[Participation of protein kinase C in hormonal regulation of pepsin secretion in the rat stomach]. / Uchastie proteinkinaszy C v gormonal'noi reguliatsii sekretsii pepsinav zheludke krys.

It was shown that pentagastrin (0.5 micrograms/100 g of body mass) increases the activity of Ca2+ and phospholipid-dependent protein kinase C in the membrane fraction of rat gastric mucosa cells. This effect of pentagastrin is accompanied by a decrease of the protein kinase C activity in the cytosolic fraction. Chromatography of the membrane fraction revealed an additional peak of the enzyme activity. Analysis of isolated gastric mucosa cells demonstrated that pentagastrin (10(-8)-10(-6) M) (but not 10(-4) M histamine) added to the incubation mixture increased the protein kinase C concentration in the membranes. The pentagastrin effect was directly correlated with the amount of pepsin-producing chief cells in the cellular pools. Carbacholine, another well-known pepsin secretion stimulator, was able to activate, similar to pentagastrin, the protein kinase C activity. It is concluded that protein kinase C plays a prominent role in hormonal regulation of the chief gastric cell function.

[Secondary messengers in the hormonal regulation of the functional activity of the main and mucoid cells in the stomach]. / Vtorichnye messendzhery v gormonal'noi reguliatsii funktsional'noi aktivnosti glavnykh i mukoidnykh kletok zheludka.

The messenger role of Ca+2, cyclic nucleotides and inositol triphosphates in the stimulation of pepsinogen and mucous secretion were studied using isolated pig [correction of nug] gastric chief cells and guinea pig mucous cells, resp. Pepsinogen secretion was stimulated by agents either working at the postreceptor adenylate cyclase (AC) level (db-cAMP, forskolin) or after 12-o-tetradecanoyl-phorbol-13-acetate (TPA) stimulation of protein kinase C (PK C). Similar secretory effects were observed with histamine (H), carbachol (C) and cholecystokinin (CCK). [Ca-2] in was elevated by C and by CCK, but not by H in both types of cells. Like TPA, both C and CCK, but not H, stimulated the Ca+2-sensitive particulate PK C. H increased the activity of cAMP-dependent PK A. PGE2, C and CCK were found to increase inositol-1,4,5-triphosphate content in mucous cells. The findings indicate that two pathways of the regulation of pepsinogen and mucous secretion (AC-cAMP-PK C and phosphoinositol breakdown cascade) can act synergistically.

Biochemical mechanisms of regulation of mucus secretion by prostaglandin E2 in rat gastric mucosa.

The effects of prostaglandin E2 (PGE2) and inhibitors of RNA and protein synthesis on rat gastric mucosa were investigated in order to study the cellular and biochemical mechanisms involved in the PGE2-stimulated formation and secretion of gastric mucus. It was shown that PGE2 caused significant stimulation of gastric mucus secretion and this effect of PGE2 was inhibited by cycloheximide but not actinomycin D. The influence of PGE2 on the in vitro incorporation of N-acetyl-[3H]glucosamine and 14C-labelled amino acids in to the glycoproteins representing a major mucus component of the isolated gastric mucosa cells was also studied. The stimulatory effect of PGE2 on incorporation of labelled precursors into glycoproteins of gastric cells was also inhibited by cycloheximide. These results suggest that the effect of PGE2 on mucus production requires ongoing protein synthesis. cAMP can fully reproduce the effect of PGE2 on the formation and the secretion of gastric mucus. The binding of [3H]PGE2 to rat gastric non-parietal cell fractions consisting predominantly of mucoid cells correlated with the ability of PGE2 to increase adenylate cyclase activity in these cells. PGE2 had no effect on adenylate cyclase activity in cell suspensions enriched in parietal cells. These data suggest further that the stimulatory effect of PGE2 on mucus secretion may be mediated by cAMP as a messenger.

Inhibition by estradiol of binding and mitogenic effect of epidermal growth factor in primary cultures of Xenopus hepatocytes.

We demonstrate here the presence of two classes of EGF (epidermal growth factor) binding sites in primary cultures of male Xenopus liver parenchymal cells. One of these corresponds to the high-affinity receptor described in other tissues and species, and which exhibits the property of autophosphorylation. The number of EGF receptors decreased sharply in freshly prepared cultures but recovered to maximum levels within 24 h thereafter. Addition of EGF and insulin to the hepatocyte cultures enhanced the rate of DNA synthesis as measured by the incorporation of [3H]thymidine. Estrogen abolished this increase, reducing the incorporation to that seen with hydroxyurea. At the same time, the addition of estradiol reduced the number or activity of EGF receptors in a dose-dependent manner. The latter paralleled the activation of transcription of vitellogenin genes in Xenopus hepatocytes so that a high rate of DNA synthesis is unnecessary for or incompatible with the activation of the steroid hormone-induced vitellogenin genes.

[Prostaglandin E2 binding by gastric mucosa cells in the rat]. / Sviazyvanie prostaglandina E2 kletkami slizistoi obolochki zheludka u krys.

After incubation of gastric pieces with [3H] prostaglandin E2 (PGE2) the label is accumulating predominantly over the cells situating in the middle third of the gastric glands proper, where mucus-secreting cells are mainly situated. As demonstrate biochemical experiments, [3H] PGE2 combines 5.5 times as intensively with the isolated cells fraction, which consists mainly of mucocytes but not of parietal cells. A suggestion is made that prostaglandin E2 contributes to biosynthesis and mucus secretion in the stomach when it immediately combines with mucocytes.

[Biochemical mechanisms of the regulation of gastric mucus secretion in the rat: the role of prostaglandin E2 and histamine]. / Biokhimicheskie mekhanizmy reguliatsii vydeleniia zheludochnoi slizi u krys: rol' prostaglandina E2 i gistamina.

Both prostaglandin E2 (PGE2) and histamine activated the adenylate cyclase system of rat gastric cells. Parietal acid-producing cells of the stomach were the target-cells for histamine, while PGE2 affected the mucous cells of gastric glands. The stimulating effect of PGE2 on mucus production occurred due to activation of protein synthesis. Electron microscopy of mucous cells detected alterations in structure of cells involved in biosynthesis of mucoids. Histamine appears to cause its effect on secretion of gastric mucus via the activating influence on production of hydrochloric acid.

[Effect of prostaglandin E2, cAMP and histamine on glycoprotein synthesis in isolated cells of the rat gastric mucosa]. / Vliianie prostaglandina E2, cAMP i gistamina na sintez glikoproteinov v izolirovannykh kletkakh slizistoi obolochki zheludka krys.

It was discovered that prostaglandin E2 (PGE2), but not histamine, increased the incorporation of 3H-N-acetyl-D-glucosamine and 14C-amino acids into the acid-insoluble protein fraction of isolated, mainly mucoid cells of rat gastric mucosa. The cAMP at the dose of 1 mM enhanced, like the PGE2, the synthesis of gastric mucoids. Cycloheximide inhibited the basal incorporation of labelled N-acetyl-D-glucosamine and the amino acid mixture by 28 and 72%, respectively, and blocked completely the PGE2 effect on glycoproteins formation. It is suggested that the PGE2, unlike histamine, enhances the biosynthesis of glycoproteins in the mucoid cells of rat gastric mucosa. The cAMP is believed to be a messenger of the PGE2 effect.

[Effect of pentagastrin, histamine, cyclic-3',5'-AMP and prostaglandin E2 on the secretion of mucus in the rat stomach]. / Vliianie pentagastrina, gistamina, tsiklicheskogo 31,51-AMF i prostaglandina E2 na sekretsiiu slizi v zheludke krys.

Pentagastrin as well as transmitters of its effect histamine and cAMP, affecting the HCl secretion, increased the gastric mucus secretion apparently due to stimulation of HCl production. This assumption is supported by experiments in which the histamine effect was not inhibited by cycloheximide but was completely eliminated in presence of burimamide--a drug blocking histamine H2-receptors. Prostaglandin E2 which inhibited distinctly basal secretion of HCl but did not affect the pepsin secretion, elevated markedly the mucus production. The effect of this stimulator of mucus production, which differed from histamine in its ability to inhibit HCl secretion, was mediated via the processes of translation but not of transcription. Regulation of mucus secretion in stomach, similarly to HCl and pepsin secretion, appears to involve a multicomponent cascade system, including prostaglandin E2 as one of transmitters. cascade system, including prostaglandin E2 as one of the transmitters.

[Localization of (3H)histamine and (3H)prostaglandin E2 receptors in isolated cells of rat gastric mucosa]. / Lokalizatsiia retseptorov [3H]gistamina i [3H]prostaglandina E2 v izolirovannykh kletkakh slizistoi zheludka krys.

Isolated cells of rat gastric mucosa were obtained by treatment of rat stomach with pronase. Two fractions were isolated, one of which was rich (up to 90%) and the second one poor (to 25%) of parietal cells. Using specific antagonists and agonists of H1- and H2-receptors of histamine (diphenhydramine, metiamide, cimetidine, impromidine, dimaprit) the H2-receptors of histamine were shown to be localized in parietal cells. A preferential binding of (3H)prostaglandin E2 by the receptor proteins of plasma membranes of non-parietal (presumably mucoid) cells was found. The data obtained indicate that rat gastric mucosa contains receptors of histamine and PGE2 which differ in their intracellular localization and strictly selectively bind (3H)histamine and (3H)PGE2. It is assumed that the starting point in the mechanism of action of these intercellular regulators on gastric secretion is probably the process of their specific recognition by the protein receptors localized in functionally different cells.