[Effect of cryopreserved cord blood nucleated cells on course of pathological processes in progressive aging of brain (experimental study).]

The data on morphometric indices of the vascular bed and brain substance of spontaneously hypertensive SHR rats, which can be considered as an evidence of the developing pathological brain aging processes in these animals, are presented in the work. Chronic alcohol intoxication aggravates the neurodegeneration, significantly reducing the indices of the neuroglial index, the number of functioning vessels and activating the LPO processes. Single intraperitoneal administration of cryopreserved human cord blood nucleated cells in a dose of (1-5)‧107/kg promoted the regeneration of neurocells, stimulation of angiogenesis and an increased level and quality of neurotissue metabolism, reducing the signs of dystrophic, destructive and pathologically altered compensatory adaptive processes in the rat's brain as a result of the restoration of its microhemocirculation and cytoarchitecture.

Immunohistochemical expression of transforming growth factor alpha and epidermal growth factor receptor in pancreatic endocrine tumors.

Coexpression of transforming growth factor alpha (TGF-alpha) and its receptor epidermal growth factor receptor (EGFR) is known to be associated with aggressive biologic behavior and adverse clinical outcome in a variety of tumors, including pancreatic adenocarcinomas. However, very little information is currently available as to whether this is true of pancreatic endocrine tumors (PETs) as well. Thirty-five PETs were retrospectively studied for immunohistochemical expression of TGF-alpha, the intracellular and extracellular domains of EGFR, and various hormonal secretory products. Proliferative activity was additionally studied (in 20 cases only) using the MIB-1 antibody. Thirty-one (89%) of 35 tumors were reactive for 1 or more of the peptide hormones tested; 22 (63%) tumors were positive for TGF-alpha; and 23 (65%) were positive for the intracellular and/or extracellular domain of EGFR. Based on their TGF-alpha and EGFR expression, these tumors could be classified into 4 groups. Of the 10 tumors in group I (positive for TGF-alpha and the complete EGFR molecule), 3 were malignant, 6 were >2 cm in diameter, 5 were functional, and 1 had a proliferative index of >40%. The 12 tumors in group II (positive for TGF-alpha but negative for the intracellular and/or extracellular domain of EGFR) included 4 malignant tumors, 4 PETs >2 cm in diameter, 8 functional, and 1 with a proliferative index of >40%. The 7 PETs in group III (positive for the intracellular/extracellular domain of EGFR alone) included 3 malignant tumors, 3 PETs >2 cm in diameter, and 3 functional tumors. The 6 tumors in group IV (completely negative for both TGF-alpha and EGFR) included 4 malignant tumors, 3 PETs >2 cm in diameter, and 4 functional lesions. Therefore, immunohistochemical expression of TGF-alpha and EGFR, either alone or in concert, shows no correlation with size, functional status, secretory profile, or biologic behavior and hence cannot be used as a marker of malignancy in this group of tumors.

Molecular mechanisms of translation initiation in eukaryotes.

Translation initiation is a complex process in which initiator tRNA, 40S, and 60S ribosomal subunits are assembled by eukaryotic initiation factors (eIFs) into an 80S ribosome at the initiation codon of mRNA. The cap-binding complex eIF4F and the factors eIF4A and eIF4B are required for binding of 43S complexes (comprising a 40S subunit, eIF2/GTP/Met-tRNAi and eIF3) to the 5' end of capped mRNA but are not sufficient to promote ribosomal scanning to the initiation codon. eIF1A enhances the ability of eIF1 to dissociate aberrantly assembled complexes from mRNA, and these factors synergistically mediate 48S complex assembly at the initiation codon. Joining of 48S complexes to 60S subunits to form 80S ribosomes requires eIF5B, which has an essential ribosome-dependent GTPase activity and hydrolysis of eIF2-bound GTP induced by eIF5. Initiation on a few mRNAs is cap-independent and occurs instead by internal ribosomal entry. Encephalomyocarditis virus (EMCV) and hepatitis C virus epitomize distinct mechanisms of internal ribosomal entry site (IRES)-mediated initiation. The eIF4A and eIF4G subunits of eIF4F bind immediately upstream of the EMCV initiation codon and promote binding of 43S complexes. EMCV initiation does not involve scanning and does not require eIF1, eIF1A, and the eIF4E subunit of eIF4F. Initiation on some EMCV-like IRESs requires additional noncanonical initiation factors, which alter IRES conformation and promote binding of eIF4A/4G. Initiation on the hepatitis C virus IRES is even simpler: 43S complexes containing only eIF2 and eIF3 bind directly to the initiation codon as a result of specific interaction of the IRES and the 40S subunit.

A conserved HEAT domain within eIF4G directs assembly of the translation initiation machinery.

The X-ray structure of the phylogenetically conserved middle portion of human eukaryotic initiation factor (eIF) 4GII has been determined at 2.4 A resolution, revealing a crescent-shaped domain consisting of ten alpha helices arranged as five HEAT repeats. Together with the ATP-dependent RNA helicase eIF4A, this HEAT domain suffices for 48S ribosomal complex formation with a picornaviral RNA internal ribosome entry site (IRES). Structure-based site-directed mutagenesis was used to identify two adjacent features on the surface of this essential component of the translation initiation machinery that, respectively, bind eIF4A and a picornaviral IRES. The structural and biochemical results provide mechanistic insights into both cap-dependent and cap-independent translation initiation.

Physical association of eukaryotic initiation factor 4G (eIF4G) with eIF4A strongly enhances binding of eIF4G to the internal ribosomal entry site of encephalomyocarditis virus and is required for internal initiation of translation.

Mammalian eukaryotic initiation factor 4GI (eIF4GI) may be divided into three similarly sized regions. The central region (amino acids [aa] 613 to 1090) binds eIF3, eIF4A, and the encephalomyocarditis virus (EMCV) internal ribosomal entry site (IRES) and mediates initiation on this RNA. We identified the regions of eIF4GI that are responsible for its specific interaction with the IRES and that are required to mediate 48S complex formation on the IRES in vitro. Mutational analysis demarcated the IRES binding fragment of eIF4GI (aa 746 to 949) and indicated that it does not resemble an RNA recognition motif (RRM)-like domain. An additional amino-terminal sequence (aa 722 to 746) was required for binding eIF4A and for 48S complex formation. eIF4GI bound the EMCV IRES and beta-globin mRNA with similar affinities, but association with eIF4A increased its affinity for the EMCV IRES (but not beta-globin RNA) by 2 orders of magnitude. On the other hand, eIF4GI mutants with defects in binding eIF4A were defective in mediating 48S complex formation even if they bound the IRES normally. These data indicate that the eIF4G-eIF4A complex, rather than eIF4G alone, is required for specific high-affinity binding to the EMCV IRES and for internal ribosomal entry on this RNA.

The functional role of basic patch, a structural element of Escherichia coli transcript cleavage factors GreA and GreB.

The transcript cleavage factors GreA and GreB of Escherichia coli are involved in the regulation of transcription elongation. The surface charge distribution analysis of their three-dimensional structures revealed that the N-terminal domains of GreA and GreB contain a small and large basic "patch," respectively. To elucidate the functional role of basic patch, mutant Gre proteins were engineered in which the size and charge distribution of basic patch were modified and characterized biochemically. We found that Gre mutants lacking basic patch or carrying basic patch of decreased size bind to RNA polymerase and induce transcript cleavage reaction in minimally backtracked ternary elongation complex (TEC) with the same efficiency as the wild type factors. However, they exhibit substantially lower readthrough and cleavage activities toward extensively backtracked and arrested TECs and display decreased efficiency of photocross-linking to the RNA 3'-terminus. Unlike wild type factors, basic patch-less Gre mutants are unable to complement the thermosensitive phenotype of GreA(-):GreB(-) E. coli strain. The large basic patch is required but not sufficient for the induction of GreB-type cleavage reaction and for the cleavage of arrested TECs. Our results demonstrate that the basic patch residues are not directly involved in the induction of transcript cleavage reaction and suggest that the primary role of basic patch is to anchor the nascent RNA in TEC. These interactions are essential for the readthrough and antiarrest activities of Gre factors and, apparently, for their in vivo functions.

The joining of ribosomal subunits in eukaryotes requires eIF5B.

Initiation of eukaryotic protein synthesis begins with the ribosome separated into its 40S and 60S subunits. The 40S subunit first binds eukaryotic initiation factor (eIF) 3 and an eIF2-GTP-initiator transfer RNA ternary complex. The resulting complex requires eIF1, eIF1A, eIF4A, eIF4B and eIF4F to bind to a messenger RNA and to scan to the initiation codon. eIF5 stimulates hydrolysis of eIF2-bound GTP and eIF2 is released from the 48S complex formed at the initiation codon before it is joined by a 60S subunit to form an active 80S ribosome. Here we show that hydrolysis of eIF2-bound GTP induced by eIF5 in 48S complexes is necessary but not sufficient for the subunits to join. A second factor termed eIF5B (relative molecular mass 175,000) is essential for this process. It is a homologue of the prokaryotic initiation factor IF2 (re and, like it, mediates joining of subunits and has a ribosome-dependent GTPase activity that is essential for its function.

[Cloning preprolactin cDNA from Pacific salmon in Escherichia coli]. / Klonirovanie kDNK preprolaktina tikhookeanskogo lososia v Escherichia coli.

Prolactin coding mRNA was shown to be a prevalent part of chum salmon (Oncorhynchus keta) pituitary poly(A)-RNA during the spawning period. Clone lambda gtPrk12 was selected from the pituitary cDNA library by means of hybridization with the prolactin probe, and a nucleotide sequence of the insertion was determined and compared to the prolactin coding sequences from rainbow trout and Pacific chinook salmon, which had been published earlier. The sequences compared exhibited a significant homology. The deduced amino acid sequence of the chum salmon prolactin differed from a sequence determined directly in a single position. The prolactin-coding sequence can be used for constructing the bacterial strain producing prolactin.

[Use of cranio-cerebral hypothermia among the complex resuscitation measures in mental disorders]. / Primenenie kraniotserebral'noi gipotermii v komplekse reanimatsionnykh meropriiatii pri psikhicheskikh zabolevaniiakh.

Craniocerebral hypothermia was employed in multimodality treatment of 10 patients with hypertoxic schizophrenia and 60 patients with intoxication psychoses, the clinical picture of which was characterized by increasing hypoxia and edema-swelling of the brain. Hypothermia was made to a cerebral temperature of 28-30 degrees C for 4-6 hours in the presence of the neurovegetative blockade. The data obtained attest to a high therapeutic efficacy of craniocerebral hypothermia.

[Cloning and structural analysis of cDNA coding for human prointerleukin-1 alpha and prointerleukin-1 beta]. / Klonirovanie i strukturnyi analiz kDNK, kodiruiushchikh prointerleikin-1 alpha i prointerleikin-1 beta cheloveka.

The data are presented on the cloning and structural analysis of the cDNA coding for human prointerleukin-1 alpha and prointerleukin-1 beta (proIL-1 alpha and proIL-1 beta). The nucleotide sequences of proIL-1 alpha and proIL-1 beta cDNAs have been compared with the sequences published earlier. The nucleotide changes resulting in the aminoacid changes of the protein were not found. Some nucleotide changes were identified within the 3'-nontranslated region of the proIL-1 beta cDNA. The existence of the allelic variants for interleukin genes registered only on the gene level has been supposed.

[Nucleotide sequence of cDNA of glicinin A3 subunits: variability of different species detected by comparative analysis]. / Nukleotidnye posledovatel'nosti kDNK sub''edinits glitsinina podsemeistva A3: razlichnaia vnutrividovaia variabel'nost', vyiavliaemaia pri sravnitel'nom analize.

Several cDNA cloned which code for subfamily A3 subunits (A3B4 and A5A4B3) of soybean storage protein glycinin were analysed by means of restriction mapping and hybrid - selection. The comparison of A3B4 and A5A4B3 subunits cDNA sequences reveals the 90% homology. The nucleotide sequences obtained in the process of this work were compared with those reported about elsewhere, in order to study the interaspecific variability of homologous but not identical storage protein genes of subfamily A3 glycinin subunit. Nucleotide replacements were found to occur 6 times more frequently in A3B4 subunit gene, as compared to A5A4B3 subunit gene (48 replacements against 8).