A cryoultratomy method and its use in immunocytochemistry]

A method of frozen ultrathin sectioning for tissues and suspension material is presented. Practical aspects and results in the field of cryoultratomy are observed in detail. An example of a good preservation of tissue ultrastructure and application of cryosections to immunocytochemical investigations is given.     

Cationic Peptides Facilitate Iron-induced Mutagenesis in Bacteria

Pseudomonas aeruginosa is the causative agent of chronic respiratory infections and is an important pathogen of cystic fibrosis patients. Adaptive mutations play an essential role for antimicrobial resistance and persistence. The factors that contribute to bacterial mutagenesis in this environment are not clear. Recently it has been proposed that cationic antimicrobial peptides such as LL-37 could act as mutagens in P. aeruginosa. Here we provide experimental evidence that mutagenesis is the product of a joint action of LL-37 and free iron. By estimating mutation rate, mutant frequencies and assessing mutational spectra in P. aeruginosa treated either with LL-37, iron or a combination of both we demonstrate that mutation rate and mutant frequency were increased only when free iron and LL-37 were present simultaneously. Colistin had the same effect. The addition of an iron chelator completely abolished this mutagenic effect, suggesting that LL-37 enables iron to enter the cells resulting in DNA damage by Fenton reactions. This was also supported by the observation that the mutational spectrum of the bacteria under LL-37-iron regime showed one of the characteristic Fenton reaction fingerprints: C to T transitions. Free iron concentration in nature and within hosts is kept at a very low level, but the situation in infected lungs of cystic fibrosis patients is different. Intermittent bleeding and damage to the epithelial cells in lungs may contribute to the release of free iron that in turn leads to generation of reactive oxygen species and deterioration of the respiratory tract, making it more susceptible to the infection.     

Inhibitors of the heat shock protein 90 activity: a novel class of tumor radiosensitizers

The 90 kDa heat shock protein (HSP90) is one of major chaperones of eukaryotes which catalyzes maturation and activation of its client proteins. Among the identified client proteins there are oncogene products, hormone or growth factor receptors and key components of signaling pathways responsible for the malignant growth of tumors or their resistance to chemotherapy and radiotherapy. In the case of inhibition of the HSP90 chaperone function, such proteins are inactivated and degraded soon that leads to simultaneous blocking several pathways essential for proliferation and survival of malignant cells; therefore, pharmacological inhibitors of the HSP90 chaperone activity could be used in anticancer therapy. At present, several HSP90 inhibitors are in preclinical testing or I-III Phase clinical trials as mono-agents or in combinations with other anticancer drugs or radiation. In the present review, all the data are summarized which characterize HSP90 inhibitors as effective radiosensitizers of tumor cells. Molecular mechanisms and selectivity of the radiosensitizing action of HSP90 inhibitors are here discussed as well as a possibility of their application to improve the outcome of radiotherapy.     

Effect of enteropeptidase on survival of cultured hippocampal neurons under conditions of glutamate toxicity

The effects of full-size bovine enteropeptidase (BEK) and of human recombinant light chain enteropeptidase (L-HEP) on survival of cultured hippocampal neurons were studied under conditions of glutamate excitotoxicity. Low concentrations of L-HEP or BEK (0.1–1 and 0.1–0.5 nM, respectively) protected hippocampal neurons against the death caused by 100 μM glutamate. Using the PAR1 (proteinase-activated receptor) antagonist SCH 79797, we revealed a PAR1-dependent mechanism of neuroprotective action of low concentrations of enteropeptidase. The protective effect of full-size enteropeptidase was not observed at the concentrations of 1 and 10 nM; moreover, 10 nM of BEK caused death of 88.9% of the neurons, which significantly exceeded the cell death caused by glutamate (31.9%). Under conditions of glutamate cytotoxicity the survival of neurons was 26.8% higher even in the presence of 10 nM of L-HEP than in the presence of 10 nM BEK. Pretreatment of cells with 10 nM of either form of enteropeptidase abolished the protective effect of 10 nM thrombin under glutamate cytotoxicity. High concentrations of BEK and L-HEP caused the death of neurons mainly through necrosis.     

Molecular cloning of the human insulin gene

Isolation of genomic clones containing the human insulin gene by screening the human genomic library for this gene using the cDNA rat insulin probe is reported. The analysis of promoter-enhancer region and exons sequences has revealed their identity to analogous sequences determined earlier.     

Oxidative processes at initial stages of interaction of nodule bacteria (<Emphasis Type="Italic">Rhizobium leguminosarum</Emphasis>) and pea (<Emphasis Type="Italic">Pisum sativum</Emphasis> L.): A review

A possible physiological mechanism of legume-Rhizobium symbiosis, consisting in regulation of the intensity of oxidative processes by the macrosymbiont in response to infection with Rhizobium, was analyzed using our own and published data. The results used in the analysis included data on the content of reactive oxygen species (O ₂ ^·? and H₂O₂), activity of antioxidant enzymes (superoxide dismutase, catalase, and peroxidase), and intensity of lipid peroxidation proceeding with the involvement of lipophilic phenolic compounds of the microsymbiont.     

The effect of the phenolic compounds exuded by pea roots in darkness on the reproduction of <Emphasis Type="Italic">Rhizobium</Emphasis>

The exudation, composition, and biological activity of the phenolic compounds (PC) of pea (Pisum sativum L.) roots in the light and darkness were studied. The roots of leguminous plants grown for 5 days in darkness exuded a smaller amount of PC that displayed a weaker stimulation of Rhizobium reproduction. Moreover, the root exudates contained antimicrobial compounds, stilbenes. It is assumed that a lower PC exudation by roots and the specific features of PC composition influencing the biological activity are among the reasons causing a delayed nodulation of legumes grown in darkness.     

Postnatal development of corticosteroid function of the adrenals in C57BL/6J-Ay mice

We studied postnatal development of corticosteroid function of the adrenals in mice during the period of elevated activity of the hypothalamic-pituitary-adrenal system and the influence of mutant gene Ay on this process. Normally, a corticosterone peak in blood and increased basal and stimulated steroidogenesis in vitro are observed in 3-week old mice. In 3-week old Ay/a mice (hyperexpression of protein agouti) a corticosterone peak in blood is lowered and genotypic differences in steroidogenesis in vitro are absent, as compared to a/A mice (absence of agouti), while at the ages of 10 and 15 weeks, there were no genotypic differences in the blood level of corticosterone and steroidogenesis in vitro was elevated. Thus, a high level of corticosterone during the period of elevated activity of the hypothalamic-pituitary-adrenal system in 3-week old mice is determined by enhanced steroidogenic function of the adrenals. Mutant gene Ay in male mice affected the postnatal development of the adrenal function: the peak of corticosterone in blood was lowered during the period of elevated activity of the system.     

The effect of the phenolic compounds exuded by pea roots in darkness on the reproduction of Rhizobium

The exudation, composition, and biological activity of the phenolic compounds (PC) of pea (Pisum sativum L.) roots in the light and darkness were studied. The roots of leguminous plants grown for 5 days in darkness exuded a smaller amount of PC that displayed a weaker stimulation of Rhizobium reproduction. Moreover, the root exudates contained antimicrobial compounds, stilbenes. It is assumed that a lower PC exudation by roots and the specific features of PC composition influencing the biological activity are among the reasons causing a delayed nodulation of legumes grown in darkness.