[ORGANIZED PNEUMONIA IN A PATIENT WITH ULCERATIVE COLITIS].

The article presents a case of organized pneumonia development in 44-year-old patient with ulcerative colitis, successfully permitted on the background of treatment with high doses of systemic steroids. The authors consider the case as extra-intestinal IBD-associated pulmonary disease.

[Indexes of immunity and local protection in humans with intestine dysbacteriosis].

AIM: Study indexes of immunity and local protection in humans with intestine dysbacteriosis. MATERIALS AND METHODS: Qualitative and quantitative intestine microbiocenosis, content of gamma-interferon (EIA method) in coprofiltrates in 204 individuals were studied, data from immunograms of 123 individuals with bacteriologically confirmed dysbacteriosis were analyzed. RESULTS: The presence of immune deficiency mainly by T-cell type was established in 92.7+/-2.4% of individuals with intestine dysbacteriosis. Significant variations in -interferon content in coprofiltrates of examined individuals was detected (from no less than 5 pcg/ml to 240 pcg/ml), a statistically significant dependence of gamma-interferon content in coprofiltrates on the number of opportunistic microbes and atypical escherichia (including hemolytic) in intestine microbiocenosis was determined. CONCLUSION: The presence of T-cell type immune deficiency in individuals with intestine dysbacteriosis combined with a reduced local protection, and the content of gamma-interferon in coprofiltrates gives evidence not only on the reduction of local protection but also to some extent mirrors the degree of this reduction.

Chromatin structure of two genomic sites for targeted transgene integration in induced pluripotent stem cells and hematopoietic stem cells.

Achieving transgene integration into preselected genomic sites is currently one of the central tasks in stem cell gene therapy. A strategy to mediate such targeted integration involves site-specific endonucleases. Two genomic sites within the MBS85 and chemokine (C-C motif) receptor 5 (CCR5) genes (AAVS1 and CCR5 zinc-finger nuclease (CCR5-ZFN) sites, respectively) have recently been suggested as potential target regions for integration as their disruption has no functional consequence. We hypothesized that efficient transgene integration maybe affected by DNA accessibility of endonucleases and therefore studied the transcriptional and chromatin status of the AAVS1 and CCR5 sites in eight human induced pluripotent stem (iPS) cell lines and pooled CD34+ hematopoietic stem cells (HSCs). Matrix chromatin immunoprecipitation (ChIP) assays demonstrated that the CCR5 site and surrounding regions possessed a predominantly closed chromatin configuration consistent with its transcriptional inactivity in these cell types. In contrast, the AAVS1 site was located within a transcriptionally active region and exhibited an open chromatin configuration in both iPS cells and HSCs. To show that the AAVS1 site is readily amendable to genome modification, we expressed Rep78, an AAV2-derived protein with AAVS1-specific endonuclease activity, in iPS cells after adenoviral gene transfer. We showed that Rep78 efficiently associated with the AAVS1 site and triggered genome modifications within this site. On the other hand, binding to and modification of the CCR5-ZFN site by a ZFN was relatively inefficient. Our data suggest a critical influence of chromatin structure on efficacy of site-specific endonucleases used for genome editing.

[Features of microbiocenosis composition of large intestine of humans with various degree of local metabolic disorders].

AIM: Detection of features of qualitative and quantitative composition of large intestine microbiocenosis of humans with various degree of local metabolic disorders during dysbacterioses due to various causes. MATERIALS AND METHODS: Microflora of large intestine and content of malonic dialdehyde (MDA) in coprofiltrates of 330 adult humans with large intestine dysbacterioses due to various causes were studied. RESULTS: It was established, that high MDA content in coprofiltrates matches higher quantities of opportunistic microorganisms and atypical escherichia in microflora composition of large intestine. Relation of MDA composition in coprofiltrates and factors that cause dysbacteriosis were not detected. CONCLUSION: The studies performed give evidence that changes in local metabolic activity may be a single mechanism of development of large intestine dysbacterioses that are caused by various factors.

The product of the murine homolog of the Drosophila extra sex combs gene displays transcriptional repressor activity.

The heterogeneous nuclear ribonucleoprotein K protein represents a novel class of proteins that may act as docking platforms that orchestrate cross-talk among molecules involved in signal transduction and gene expression. Using a fragment of K protein as bait in the yeast two-hybrid screen, we isolated a cDNA that encodes a protein whose primary structure has extensive similarity to the Drosophila melanogaster extra sex combs (esc) gene product, Esc, a putative silencer of homeotic genes. The cDNA that we isolated is identical to the cDNA of the recently positionally cloned mouse embryonic ectoderm development gene, eed. Like Esc, Eed contains six WD-40 repeats in the C-terminal half of the protein and is thought to repress homeotic gene expression during mouse embryogenesis. Eed binds to K protein through a domain in its N terminus, but interestingly, this domain is not found in the Drosophila Esc. Gal4-Eed fusion protein represses transcription of a reporter gene driven by a promoter that contains Gal4-binding DNA elements. Eed also represses transcription when recruited to a target promoter by Gal4-K protein. Point mutations within the eed gene that are responsible for severe embryonic development abnormalities abolished the transcriptional repressor activity of Eed. Results of this study suggest that Eed-restricted homeotic gene expression during embryogenesis reflects the action of Eed as a transcriptional repressor. The Eed-mediated transcriptional effects are likely to reflect the interaction of Eed with multiple molecular partners, including K protein.

Point mutations in the WD40 domain of Eed block its interaction with Ezh2.

The Polycomb group proteins are involved in maintenance of the silenced state of several developmentally regulated genes. These proteins form large aggregates with different subunit compositions. To explore the nature of these complexes and their function, we used the full-length Eed (embryonic ectoderm development) protein, a mammalian homolog of the Drosophila Polycomb group protein Esc, as a bait in the yeast two-hybrid screen. Several strongly interacting cDNA clones were isolated. The cloned cDNAs all encoded the 150- to 200-amino-acid N-terminal fragment of the mammalian homolog of the Drosophila Enhancer of zeste [E(z)] protein, Ezh2. The full-length Ezh2 bound strongly to Eed in vitro, and Eed coimmunoprecipitated with Ezh2 from murine 70Z/3 cell extracts, confirming the interaction between these proteins observed in yeast. Mutations T1031A and T1040C in one of the WD40 repeats of Eed, which account for the hypomorphic and lethal phenotype of eed in mouse development, blocked binding of Ezh2 to Eed in a two-hybrid interaction in yeast and in mammalian cells. These mutations also blocked the interaction between these proteins in vitro. In mammalian cells, the Gal4-Eed fusion protein represses the activity of a promoter bearing Gal4 DNA elements. The N-terminal fragment of the Ezh2 protein abolished the transcriptional repressor activity of Gal4-Eed protein when they were coexpressed in mammalian cells. Eed and Ezh2 were also found to bind RNA in vitro, and RNA altered the interaction between these proteins. These findings suggest that Polycomb group proteins Eed and Ezh2 functionally interact in mammalian cells, an interaction that is mediated by the WD40-containing domain of Eed protein.

[Dynamic health state control in personnel of the Radiology Department of Bryansk Regional Oncological Dispensary].

The health state of personnel of the Radiology Department of Bryansk Regional Oncological Dispensary before and after Chernobyl Nuclear Disaster is analyzed using an automated classifying system. The system operation is based on analysis of peripheral blood count.

Regulation of LacZ mRNA translatability in a cell-free system at heat shock by the last four sense codons.

Under heat shock conditions translation of Xenopus laevis normal mRNAs in a rabbit reticulocyte cell-free system is blocked whereas hsp70 mRNA is translated. mRNA for E. coli beta-galactosidase containing the last four sense codons of Drosophila hsp70 at its 3'-end was constructed. This mRNA is efficiently translated in a rabbit reticulocyte cell-free system at 43 degrees C.

Alterations in protein synthesis induced by C2 toxin in 3T3 cells.

We have studied the effect of actin skeleton depolymerisation induced by C2 toxin on protein synthesis in 3T3 cells. The toxin that was purified from culture medium of Clostridium botulinum type C was shown to specifically ADP-ribosylate actin in vitro and in vivo. Cells exposed to C2 toxin were rounded off, which was accompanied by disappearance of stress fibers. The rate of total protein synthesis decreased two-three times in the treated cells. This correlated with the reduction in amount of polyribosomes. The rates of specific protein synthesis were compared using 2D electrophoresis of pulse-labeled proteins. Dramatic changes were observed in the synthesis of a small group of cellular proteins. Our results indicate that actin filament depolymerization affects gene expression at the level of translation and/or through the control of mRNA concentrations.

Diverse molecular interactions of the hnRNP K protein.

The hnRNP K protein is a versatile molecule that interacts with RNA, DNA, the proto-oncoprotein VaV, Src-like tyrosine and inducible serine/threonine kinases, the transcription factor TBP and a number of zinc-finger transcriptional repressors. The interaction of K protein with some of its protein partners is modulated by nucleic acids and K protein can alter the in vivo and in vitro rate of transcription. K protein can simultaneously engage several proteins and may facilitate molecular cross-talk. Taken together these diverse interactions suggest that K protein may act as a nucleic acid-regulated docking platform.

An inhibitor of protein synthesis initiation from Alhagi kirgisorum S.

Polyproanthocyanidin--a plant phenolic compound from Alhagi kirgisorum S. effectively inhibited protein synthesis in rabbit reticulocyte and wheat germ cell-free systems. Poly-proanthocyanidin inhibited translation only at the level of initiation and not at the elongation level and aminoacylation of tRNA. The inhibitory effect of the phenolic compound is due to the blockage of the ternary complex formation of eIF-2 with GTP and initiator Met-tRNA.

Stimulation of actin synthesis in phalloidin-treated cells. Evidence for autoregulatory control.

We used the technique of scrape loading to introduce phalloidin into mouse embryo fibroblasts in mass culture. Phalloidin almost completely destroyed actin microfilament bundles, but the amount of polymerized cytoskeleton-associated actin was increased approximately two-fold and the amount of monomeric (Triton X-100 extractable) actin was significantly reduced. The major result of the present study is that the rate of actin synthesis in the phalloidin-treated cells was 2-3 times higher than in the control cells. Northern blot and translation in a cell-free system from rabbit reticulocytes showed that the actin mRNA level significantly increased as a result of phalloidin treatment.

A novel open reading frame in tobacco mosaic virus genome coding for a putative small, positively charged protein.

From sequence comparisons between the tobramovirus genomes an open reading frame (ORF-X) potentially encoding a small, positively charged protein (33- to 45-amino-acids long) was found to overlap the immediate 3' and 5' sides of the transport protein gene and coat protein gene, respectively. In vitro translation of the monocistronic artificial transcripts generated with T7 RNA polymerase yielded a protein of M(r) 4000 (p4) and an unexpected trypsin-sensitive complex of M(r) 54,000 that was resistant to reduction with 2-mercaptoethanol but could be dissociated by 8 M urea. Assembly of this complex was inhibited completely by site-directed mutagenesis within a conserved, positively charged 5-amino-acid long segment of the ORF-X protein. After centrifugation in low salt buffer the 54-kDa complex remained mostly associated with ribosomes. Apparently this complex represents a specific aggregate of the p4 product of ORF-X with a protein of approximate M(r) 50,000 that is a component of the translation apparatus.

The interactions of anti-MLI monoclonal antibodies with isoforms of the lectin from Viscum album.

Monoclonal antibodies (mAb) reacting with native (TA5, TB12) and denatured (T33, T35) plant toxin mistletoe lectin I (MLI) from Viscum album have been obtained. The interaction between mAbs and native toxin with ML isoforms (MLII, MLIII) has been investigated. An immunological cross-reaction has been shown to take place for mAb TA5 (anti-A-chain of MLI) between MLII and MLIII isoforms of toxin. TA5 has not inhibited enzyme activity of the A-chain in a rabbit reticulocyte cell-free system. TB12 has been shown to react with the galactose-binding site of the B-chain. TA5 and TB12 have shown no cross-reaction with plant toxin ricin. The association constants for mAbs have been determined. The nature of heterogeneity of the lectins from Viscum album is discussed.

Physical, chemical and serological properties of C-type virus associated with transplantable rat Thurzo-Svee leukemia.

Cells of rat Thurzo-Svee leukemia produce C-type virus. Viral particles are shown to contain high-molecular weight RNA and RNA-dependent DNA polymerase the virus possesses mammalian gs-3 antigen and rat species-specific gs-1 antigen and thus appears to be of rat origin.

ESR and TL dosimetry systems: comparative measurements for human phantom.

Mixtures of small fragments of tooth enamel as well as thermoluminescence (TL) dosimeters were placed into the tissue-equivalent phantom of the human head with skeleton (approximately at the level of the jaws) and irradiated using 137Cs low dose-rate gamma therapeutic sources ('SELEKTRON' LDR 137Cs). Phantom, samples of teeth and TL detectors were irradiated behind water tank to produce scattered irradiation. The same irradiation with the same geometry was performed in air too. For gamma-spectrometry 137Cs sources with very low activity were used but with the same geometry as therapeutic sources. The absorbed dose in enamel was estimated with the help of ESR spectrometer 'ESP-300 E' (Brucker). The samples of tooth enamel were partially used for preliminary dose evaluation by ESR signal before starting of experiment. TL dosimetry was performed by TL reader model 8800 (HARSHAW) using TL dosimeters calibrated with 137Cs. The paper presents data obtained in comparative aspects.

[Study of plant lectins from Viscum album using monoclonal antibodies]. / Issledovanie rastitel'nykh lektinov iz omely beloi s pomoshch'iu monoklonal'nykh antitel.

Monoclonal antibodies (monAT) against both native (TA5, TB12) and denatured (TB33, TB35) plant toxin ML1 from Viscum album have been obtained. The interaction of monAT against native toxin with its isoforms ML2 and ML3 was investigated. It was shown that monAT TA5 to A-chain of ML1 toxin cross-reacted with ML2 and ML3 isoforms. TA5 did not inhibit enzyme activity of A-chain in cell-free rabbit reticulocyte system. It was shown that monAT TB12 reacted with galactose-binding site of B-subunit. Both monAT had no cross-reactions with plant toxin ricin. The binding constants for TA5 with ML1, ML2, ML3 respectively were 4.3.10(7) M-1, 1.2.10(7) M-1, and 0.3.10(7) M-1. The binding constants for TB12 were 2.10(7) M-1 with ML1 toxin, and more than 10(6) M-1 with ML2 and ML3. The nature of heterogeneity in ML toxin family is discussed. Test-systems for ML1 determination in different V. album extracts are suggested.

[The effect of ts-mutation on expression of genes induced by heat shock in Drosophila melanogaster. III. Synthesis of proteins cognate to HSP70]. / Vliianie ts-mutatsii na ékspressiiu genov, indutsiruemykh teplovym shokom u Drosophila melanogaster. Soobshchenie III. Sintez belkov, rodstvennykh BShT70.

2D-electrophoresis performed as described by O'-Farrel has revealed clear-cut differences in the pattern of proteins synthesized in the cells of wild-type flies and in the cells of ts-lethal. The cells of the mutant studied after heat-shock exhibit not only the lack of HSP83 and HSP35 but also the absence of a few of the heat-shock proteins belonging to the HSP70 group. Moreover, in the cells of the mutant an intensive synthesis of a protein with mol. weight 72 KDa was observed after heat-shock. This protein belongs to highly abundant heat-shock cognate proteins (HSCP) which are usually not induced by temperature elevation.