Nucleic acid metabolism in rat liver after single and prolonged exposure to tritium oxide

It was shown that after a single administration of tritium oxide in a dose of 22.2 MBq/g body mass the liver mass increased, the concentration of nucleic acids decreased and the biosynthesis rate increased during a one-month observation. By the end of the observation period (the first year) the parameters under study were normalized. The long-term administration of tritium oxide in daily doses of 0.37, 0.925 and 1.85 MBq/g body mass caused changes in the nucleic acid metabolism which were less manifest (at early times), than in the case of a single injection. At the same time, the long-term administration of tritium oxide in the dose of 0.925 MBq/g caused a substantial disturbance of the nucleic acid metabolism at later times (after 2-9 months).     

Nucleic acid metabolism in rat hematopoietic tissues during and after prolonged administration of different doses of tritium oxide

During the long-term administration to rats of tritium oxide in doses of 0.37, 0.925 and 1.85 MBq/g body mass the content of karyocytes and nucleic acids in the bone marrow and spleen was decreased, the rate of their biosynthesis changed, the DNA structure impaired, the content of salt-soluble polydeoxynucleotides increased, and DNAases activated. The observed changes were function of dose. After the end of the administration of the isotope the animals which had received a lesser tritium dose exhibited a more rapid and complete recovery.     

Dependence of the damage and recovery of nucleic acid metabolism on the dose rate in tritium oxide exposure

Disturbance and normalization of nucleic acid metabolism in rat thymus was studied after the effect of tritium oxide delivered in similar cumulative doses but at different dose rates. Both the disturbance and normalization were shown to be a function of dose rate, the slightest damage and the complete recovery being registered at the lowest dose rate (the amount of tritium oxide administered being 0.37 MBq/g/day). The rate of restoration was also a function of dose rate; with tritium oxide dose of 1.85 MBq/g/day (the dose rate at the stage of the equilibrium tritium content in the aqueous phase being 0.38 Gy/day) it was 9 times as high as that after a dose of 0.37 MBq/g/day (0.11 Gy/day dose rate).     

Relative biological effectiveness of tritium oxide based on nucleic acid metabolic indices

Relative biological effectiveness (RBE) of tritium oxide, as compared to gamma-rays (137Cs), with regard to LD50/30 is 2,32 +/- 0,69 for rats. The RBE coefficients for tritium oxide are obtained with regard to some indices of nucleic acid metabolism in the thymus and spleen during the dose formation (0-14 days). The RBE of tritium oxide increases with a decrease in radiation dose as determined according to the concentration and content of DNA per organ and activity of thymus DNAases.     

Comparison of the structure and catabolism of rat thymus DNA exposed to tritium oxide and gamma-radiation in equal doses

A decrease in DNA concentration and in the molecular mass of single-stranded DNA, an increase in the PDN content, and activation of acid DNAses in rat thymus were observed after a single administration of tritium oxide in a dose of 22 mBq/g (a cumulative dose of 7.8 Gy) and gamma-irradiation at a corresponding dose-rate and value of the cumulative dose. These changes were most pronounced during the period of dose accretion, i.e. during 14-30 days after the beginning of irradiation. The degree to which the indices under study varied from the controls was 2-3 times in rats given tritium oxide than in those exposed to gamma-irradiation.     

Incorporation of tritium into the DNA of hematopoietic tissues during prolonged administration of tritium oxide

With long-term (90 days) administration of tritium oxide (0.37 MBq/g body weight) to ras the carbon-bound tritium accumulated in DNA of haemopoietic tissues during two-month administration of the isotope (the accumulation half-time of 15-25 days); during the next month, the isotope level remained nearly constant (about 20 X 10(6) decay/min/g DNA). Elimination of tritium from DNA started 3 days after termination of its administration and proceeded with two half-times (4-8 days and 12-18 days). The ratio of the tritium content per 1 M hydrogen of DNA to tritium content per 1 M hydrogen of tissue water increased up to 0.5-0.7 during the uptake of tritium oxide, and up to 4-7 after the administration of the isotope had ceased.     

Study of the structure and metabolism of the rat thymus gland during long-term administration of tritium oxide in different doses

During-3-month administration of tritium oxide (0.37, 0.925 and 1.85 MBq/g body weight) to rats one could observe the loss of the thymus weight, decrease in the concentration, content and production of DNA and RNA per organ, reduction of the molecular weight of single-stranded DNA, increase in the content of the salt-soluble polydeoxyribonucleotides, and activation of acidic DNAases. These changes were function of tritium dose daily administered to rats.     

Mutation induction by tritiated water and effects of deuterium oxide in cultured mouse leukemia cells

Induction of mutation to 6-thioguanine resistance was studied in L5178Y mouse leukemia cells after exposure to low-dose-rate gamma rays or tritiated water at dose rates of approximately 0.025 to 0.4 Gy/hr for 20 hr in the presence or absence of 45% (v/v) deuterium oxide. The effect of acute gamma-ray exposure was also examined. A higher frequency of induced mutations was observed after tritium beta rays than after gamma rays, both at equivalent doses and cell survival. Deuterium oxide enhanced the mutation induced by gamma rays and tritium beta rays but did not affect the survival-mutation correlation of the two radiations.     

A STUDY OF THE PENETRATION OF MAMMALIAN CELLS BY DEOXYRIBONUCLEIC ACIDS

Tritium-labeled deoxyribonucleic acid (DNA) from pneumococci and from human leukocytes was added to growing cultures of HeLa cells at 37°C. Autoradiography revealed an extensive localization of tritium in the nuclear regions. The label could not be removed by treatment with ribonuclease or dilute perchloric acid, but quantitative removal from the cells could be effected with deoxyribonuclease. Chemical and radioactivity determinations on nucleic acids isolated from the exposed HeLa cells revealed the presence of tritium in all 4 DNA bases. About 12 µg. of tritiated DNA was recovered from 6 x 10⁶ HeLa cells which had been exposed for 24 hours to 240 µg. of the human DNA. From this, it is concluded that the amount of DNA, or its degradation products, taken up by the cells was equivalent to at least 10 per cent of the normal HeLa cell complement.     

Normal killer function in CBA mice as affected by long-term intake of tritium oxide

A study was made of the cytotoxic function of normal killers of CBA mice during long-term administration of tritium oxide in potable water (a cumulative absorbed dose of 8.7 Gy, dose-rate, 4.5 Gy/day) and at different times after termination of the radionuclide injection. A mean decrease of 20-30% in the lytic effect of the effectors from spleen of the experimental mice on cell-targets of K-562 human erythroleukosis was demonstrated by a 17-19-hour test with 51Cr. At later times after termination of action of tritium oxide, the cytotoxic effect of normal killers increased by 1.8 times as compared to intact controls of the same age.     

The immunity indices of rats after the long-term action of tritium oxide or gamma irradiation

In experiments with Wistar rats, a study was made of the content of antibody-forming cells and cytotoxic activity of normal killers after long-term administration of tritium oxide (3HOH) (370 kBq.g-1 of body mass daily, cumulative dose, 8.1 Gy, and dose rate, 8.5 cGy/day), and after gamma irradiation with corresponding doses. The long-term radiation effect caused a decrease in the immunity indices: the impairment of the immune reactions was more pronounced after the effect of 3HOH than after gamma irradiation. Damages to the immune system of mice and rats after irradiation with similar doses were compared.     

Voltammetric determination of gamma radiation-induced DNA damage

Homopolydeoxyribonucleotides, poly[dGuo], poly[dAdo], poly[dThd], and poly[dCyd], and calf thymus single-stranded DNA (ssDNA) and double-stranded DNA (dsDNA) aqueous solutions previously exposed to gamma radiation doses between 2 and 35 Gy, were studied by differential pulse voltammetry using a glassy carbon electrode. The interpretation of the voltammetric data was also supported by the electrophoretic migration profile obtained for the same ssDNA and dsDNA gamma-irradiated samples by nondenaturing agarose gel electrophoresis. The generation of 8-oxo-7,8-dihydroguanine, 2,8-dihydroxyadenine, 5-formyluracil, base-free sites, and single- and double-stranded breaks in the gamma-irradiated DNA samples was detected voltammetrically, with the amount depending on the irradiation time. It was found that the current peaks obtained for 8-oxoguanine increase linearly with the radiation dose applied to the nucleic acid sample, and values between 8 and 446 8-oxo-7,8-dihydroguanine (8-oxoGua) per 10(6) guanines per Gy were obtained according to the nucleic acid sample. The results showed that voltammetry can be used for monitoring and simultaneously characterizing different kinds of DNA damage caused by gamma radiation exposure.     

Improved method to prepare RNA-free DNA from mammalian cells

To isolate DNA for nucleoside analog incorporation studies, many investigators use RNase A to remove RNA from total cellular nucleic acid. We observed persistence of ribonucleotides from RNA in nucleic acid samples treated with RNase A alone. Although incubation of [5-³H]uridine-labeled nucleic acid with 50 μg/ml RNase A decreased tritium by 97%, HPLC analysis of the resulting DNA preparation digested to nucleosides revealed high levels of ribonucleosides. Increasing RNase A 10-fold (500 μg/ml) effected only a 1.7-fold reduction in ribonucleosides. Overall, the level of ribonucleosides was one-fourth that of the deoxynucleosides, primarily due to the high levels of guanosine. It was hypothesized that the ribonucleosides originated from guanosine-rich tracts of RNA since RNase A cuts preferentially 3′ to pyrimidine monophosphates and to some extent after AMP. The addition of 0.05 μg/ml RNase T1, which preferentially cleaves RNA 3′ to GMP, decreased total ribonucleosides by nearly 20-fold. In conclusion, we have developed a rapid method which removes greater then 99% of cellular RNA from nucleic acid extracts and a reversed-phase HPLC procedure that detects RNA contamination more sensitively than [5-³H]uridine labeling. These methods are useful for the determination of analog incorporation into DNA, especially for agents which incorporate into both DNA and RNA.     

Relation of the biosynthesis of intracellular and export proteins in rat liver cells during induction of proliferation by cycloheximide

After a single injection of a sublethal dose of cycloheximide (CHI) the biosynthesis of extracellular proteins in rat hepatocytes was rapidly suppressed, the reconstitution being very slow. On the contrast, the biosynthesis of intracellular proteins (e.g., histones, and other acid-soluble liver proteins) was more resistant to CHI. The activation of biosynthesis of acid-soluble and acid-insoluble proteins was found to occur stepwise. It was assumed that the activation of synthesis and accumulation of intracellular proteins after CHI release accompanied by a decreased synthesis of extracellular proteins is one of possible causes of stimulation of DNA synthesis in the hepatocytes following a single injection of CHI.     

壳寡糖对烟草悬浮细胞茉莉酸合成基因转录的影响

采用RT-PCR方法研究了不同浓度壳寡糖对烟草悬浮细胞茉莉酸合成酶基因的转录调控。结果表明, 50 μg.mL-1壳寡糖能够明显诱导烟草悬浮细胞茉莉酸合成途径的关键酶——磷脂酶A2、13-脂氧合酶、丙二烯氧化物合成酶、丙二烯氧化物环化酶和12-氧-植物二烯酸还原酶基因的表达, 而且该浓度的壳寡糖对这些基因的诱导作用相同(似)。在实验设定时间内均诱导表达编码磷脂酶A2的基因, 对其它基因的诱导时间均为8小时, 表明50 μg.mL-1壳寡糖在诱抗过程中启动了茉莉酸合成途径。而200 μg.mL-1壳寡糖的处理对这些基因的表达无显著影响。表明不同浓度的壳寡糖对烟草悬浮细胞的作用模式存在差异, 且高浓度的壳寡糖在烟草悬浮细胞中启动的信号通路可能没有茉莉酸信号的参与。     

壳寡糖对烟草悬浮细胞茉莉酸合成基因转录的影响

采用RT-PCR方法研究了不同浓度壳寡糖对烟草悬浮细胞茉莉酸合成酶基因的转录调控。结果表明,50μg·mL^-1壳寡糖能够明显诱导烟草悬浮细胞茉莉酸合成途径的关键酶——磷脂酶A2、13-脂氧合酶、丙二烯氧化物合成酶、丙二烯氧化物环化酶和12-氧-植物二烯酸还原酶基因的表达,而且该浓度的壳寡糖对这些基因的诱导作用相同（似）。在实验设定时间内均诱导表达编码磷脂酶A2的基因,对其它基因的诱导时间均为8小时,表明50μg·mL^-1壳寡糖在诱抗过程中启动了茉莉酸合成途径。而200μg·mL^-1壳寡糖的处理对这些基因的表达无显著影响。表明不同浓度的壳寡糖对烟草悬浮细胞的作用模式存在差异,且高浓度的壳寡糖在烟草悬浮细胞中启动的信号通路可能没有茉莉酸信号的参与。     

Relative biological effectiveness of tritium oxide by the criterion of death of the germ cells in mice

It was shown that low-level beta-radiation of tritium is much more effective than gamma-radiation of 137Cs with respect to reduction of the mouse testis weight. The RBE coefficient increases from 1.8 at a dose of 1 Gy to 2.2-2.3 at 0.1 Gy. On the basis of the data obtained by the authors and those reported in the literature a quality factor is proposed for tritium: QF = 2. Using THO and Na36Cl labels a mean water content of the testis cells, necessary for the estimation of a tritium-radiation dose absorbed, has been determined: gamma ct = 0.70 +/- 0.02 ml/g.     

The binding of metabolites formed from aminostilbene derivatives to nucleic acids in the liver of rats.

Carcinogenic trans-4-dimethylaminostilbene (trans-DAS) and trans-4-acetylaminostilbene (trans-AAS) as well as inactive cis-DAS and DABB were highly and specifically labeled with tritium and administered orally to female Wistar rats. Covalent binding to liver rRNA and DNA was measured and found to be higher for the carcinogenic compounds. Digests from these nucleic acids were chromatographed on Sephadex LH-20 and 16 different nucleoside adducts were characterised by their retention volumes. Labeled trans-DAS was administered in doses ranging from 0.025--250 mumol/kg. Binding to nucleic acids was directly proportional to the dose at low doses (0.025--2.5 mumol/kg) and less than proportional at higher doses (25--250 mumol/kg). The pattern of nucleoside adducts remained practically constant over the wide range of doses. A pharmacokinetically determined threshold of metabolic activation thus could not be demonstrated for this compound. A modified procedure is described to simultaneously isolate pure liver rRNA and DNA from nonfasted rats in high yields.     

γPNA一种新型高效的肽核酸

肽核酸(peptide nucleic acid,PNA)是一种人工合成的具有类多肽骨架的DNA类似物,具有与核酸结合特异性强、组织和细胞内生物稳定性好、半衰期长等优点。通过靶向结合DNA/RNA而抑制其复制、转录和翻译过程,进行基因调控。在PNA骨架结构中γ位点引入带手性的官能团,能形成右手螺旋结构,显著提高其与靶DNA/RNA的杂交特性,这种PNA衍生物称之为γPNA。γPNA的溶解性、热稳定性和特异性等化学与生物学特性明显改善,在基因编辑和作为探针检测等方面具有良好的应用前景。通过对γPNA结构、性质及其研究进展进行总结,以期为γPNA反义应用提供理论依据和参考。     

Study of the chemical makeup of the mycelium from an active strain of Act. rimosus and from an inactive mutant in relaiton to oxytetracycline biosynthesis]

Chemical composition of the mycelium of the active and inactive mutants of Act. rimosus grown under conditions favourable for oxytetracycline biosynthesis on the starch or maltose medium and under favourable conditions on the glucose medium was studied. It was shown that according to its chemical composition the above strains did not practically differ. When grown on the starch medium the mycelium of both strains contained great amounts of carbohydrates and comparatively small amounts of nucleic acids and nitrogen. Replacement of starch in the medium by glucose or maltose induced significant changes in the mycelium composition: the synthesis of intracellular polysaccharides was markedly suppressed and the synthesis of nucleic acids and nitrogen containing compounds increased. RNA was the main nucleic acid in both strains on starch and glucose media. The content of DNA was low and did not practically change. The mycelium of both strains contained small amounts of lipids which did not significantly change during the process of cultivation and did not correlate with the antibiotic activity.