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Biomedical subjects

V Ivanov

Publications and source records attributed to V Ivanov.

At least 19 recordsLinked to original sources
PubMed1996

Molecular structure and mechanisms of action of cyclic and linear ion transport antibiotics.

Ionophores are antibiotics that induce ion transport across natural and artificial membranes. The specific function of a given ionophore depends upon its selectivity and the kinetics of ion capture, transport, and release. Systematic studies of complexed and uncomplexed forms of linear and cyclic ionophores provide insight into molecular mechanisms of ion capture and release and the basis for ion selectivity. The cyclic dodecadepsipeptide valinomycin, cyclo[(-L-Val-D-Hyi-D-Val-L-Lac)3-], transports potassium ions across cellular membrane bilayers selectively. The x-ray crystallographic and nmr spectroscopic data concerning the structures of Na+, K+, and Ba+2 complexes are consistent and provide a rationale for the K+ selectivity of valinomycin. Three significantly different conformations of valinomycin are observed in anhydrous crystals, in hydrated crystals grown from dimethylsulfoxide, and in crystals grown from dioxane. Each of these conformations suggests a different mechanism of ion capture. One of the observed conformations has an elliptical structure stabilized by four 4<--1 intramolecular hydrogen bonds and two 5<--1 hydrogen bonds. Ion capture could be readily achieved by disruption of the 5<--1 hydrogen bonds to permit coordination to a potassium ion entering the cavity. The conformation found in crystals obtained from dimethyl sulfoxide is an open flower shape having three petals and three 4<--1 hydrogen bonds. Complexation could proceed by a closing up of the three petals of the flower around the desolvating ion. In the third form, water molecules reside in the central cavity of a bracelet structure having six 4<--1 hydrogen bonds. Two of these bracelets stack over one another with their valine-rich faces surrounding a dioxane molecule. The stacked molecules form a channel approximately 20 A in length, suggesting that under certain circumstances valinomycin might function as a channel. A series of analogues of valinomycin differing in ring composition and size have been synthesized and their transport properties tested. Peptide substitution and chiral variation in the dodecadepsipeptide can result in stabilization or modification of the different conformers. While contraction of the ring size results in loss of ion transport properties, expansion of the ring size permits complexation of larger ions and small positively charged molecules. Gramicidin A is a pentadecapeptide that functions as a transmembrane channel for transporting monovalent cations. Crystal structures of the cesium chloride complex and two uncomplexed forms of gramicidin A have been reported. In all three structures the gramicidin A molecule is a left-handed, antiparallel, double-stranded helical dimer. In the cesium complex the beta 7.2-helix has 6.4 residues per turn with an internal cavity large enough to accommodate cesium ions. In the uncomplexed structures the channel is 31 A long and has 5.6 amino acids per turn. Because the helix is too tightly wound to permit ion transport, ion transport would require breaking and reforming of hydrogen bonds.

Amino Acid Sequence↗
PubMed1994

Regulation of nuclear factor-kappa B and activator protein-1 activities after stimulation of T cells via glycosylphosphatidylinositol-anchored Ly-6A/E.

Cross-linking of glycosylphosphatidylinositol-anchored proteins, including mouse Ly-6A/E, leads to IL-2 secretion and T cell activation, whereas engagement of Ly-6A/E uniquely inhibits IL-2 production induced via TCR. However, little is known concerning the molecular mechanism by which glycosylphosphatidylinositol-anchored proteins regulate IL-2 expression. In this study, we have examined the ability of an anti-Ly-6A/E mAb to regulate transcription factors controlling IL-2 expression. Stimulation of EL4J(Ly-6E).A4 cells with anti-CD3 epsilon or anti-Ly6A/E mAbs induced nuclear factor (NF)-kappa B p65-p50 (RelA/p50) and AP-1 (Fos/Jun) binding activities and increased nuclear factor of activated T cells (NF-AT) activity, whereas octamer-binding factor and NF-Y levels were stable. Cyclic AMP response element binding protein and T cell-specific factor-1 (alpha) activities were selectively enhanced by anti-CD3 epsilon, but not by anti-Ly6A/E, which suggests that signaling via the TCR and Ly-6 were not identical. Costimulation of these cells with both mAbs produced substantially reduced levels of AP-1, NF-AT, and, especially, NF-kappa B p65-p50 whereas cyclic AMP response element binding protein and T cell-specific factor-1(alpha) were induced to a level seen after stimulation by anti-CD3 epsilon. The inducibility of the IL-2 enhancer in vivo and the contribution of individual transcription factors for this induction were assessed with use of reporter chloramphenicol acetyltransferase constructs containing the IL-2 enhancer or oligomerized binding sites for transcription factors. These experiments also demonstrated a key role for NF-kappa B and AP-1 in the transcriptional regulation of the IL-2 gene by TCR- and Ly6A/E-mediated signaling. By using the 2B4.11 T cell hybridoma and a mutated variant, were revealed a crucial role for the zeta-chain in Ly6A/E-mediated activation of NF-kappa B.

Antibodies, Monoclonal↗
PubMed1994

Changes in some parameters of the immune response in rats after cold stress.

The influence of stress upon the endocrine system and its relationship to the immune system have been studied. The simulation of chronic cold stress in rats caused an increase in ACTH, corticosterone, and T3 and T4 levels. The different hormonal background in stressed animals affected the characteristics and the intensity of the immune response, which was traced in dynamics and through the action of 10(9) ovine erythrocytes. A strong suppression of B-lymphocytes and of PFC count was observed. Increased Fc gamma-RFC-suppressing activity was also determined. The PFC suppression changed simultaneously with the rate and dynamics of Fc gamma-RFC-suppressing activity in animals under stress. In control rats (immunized with ovine erythrocytes only), the antigen caused insignificant changes in corticosterone level, which was not typical for stress and was not accompanied by suppressing activity in the immune response.

Adrenocorticotropic Hormone↗
PubMed1994

Source estimation in the human brain from EEG based on the SSB Head Model.

Neuronal activity in the brain cortex is replaced by a small current dipole, and hence the localized electric activities of the cortex are well approximated by equivalent current dipoles. Position and moments of these equivalent dipoles are estimated from the scalp potentials. In this analysis, the head as a conductor is divided into three volumes with uniform electric conductivity: Scalp, Skull and Brain tissue; we call this head model the SSB Head Model. The boundary element method is used in solving this problem and reliability of the final result mainly depends on the arrangement of nodal points on the surfaces of these three regions for numerical calculation, the electric conductivity ratios of the three regions in the model, and the number of electrodes. By means of computer simulation we investigated dependence of errors in the estimated dipole position on these factors. It is found that arrangement of nodal points must be decided adaptively depending on the source position.

Brain↗
PubMed1993

Antioxidant treatment of thymic organ cultures decreases NF-kappa B and TCF1(alpha) transcription factor activities and inhibits alpha beta T cell development.

Using electrophoretic mobility shift assays (EMSA), we have recently shown that nuclear extracts of 14-day mouse fetal thymocytes contain abundant NF-kappa B transcription factor activity. To determine the functional role of NF-kappa B in early thymocyte development, we have exposed fetal thymus organ cultures to inhibitors of NF-kappa B activation, namely the antioxidants N-acetyl-L-cysteine and butylated hydroxyanisole. Both compounds caused a dose-dependent arrest of thymocyte differentiation toward alpha beta, but not gamma delta, T cells. This was associated with a profound decrease in nuclear content of NF-kappa B and TCF1(alpha) transcription factor activity, as determined by EMSA. In contrast, NF-Y was affected less strongly, and cyclic AMP-response-element-binding protein levels remained essentially unchanged by antioxidants. To test the idea that alpha beta T cell development is correlated with NF-kappa B and TCF1(alpha) activity, we conducted additional experiments in a submersion culture system in which the generation of alpha beta T cells can be manipulated. Standard submersion culture supports gamma delta but alpha beta T cell development. Under these conditions, EMSA showed that transcription factor activities were similar to those seen in the presence of antioxidants. Importantly, when the generation of alpha beta T cells in submersion culture was restored by elevating oxygen concentrations, there was a dramatic increase in TCF1(alpha) activity, and both NF-kappa B and NF-Y returned to control levels. Taken together, these results strongly suggest that NF-kappa B and TCF1(alpha), presumably in concert with other transcription factors, play an important role in the development of alpha beta T cells.

Acetylcysteine↗
PubMed1993

Biological monitoring of acrylonitrile exposure through a new analytical approach to hemoglobin and plasma protein adducts and urinary metabolites in rats and humans.

A new, simple and fast procedure of measuring acrylonitrile (ACN) in ACN derived mercapturic acids such as S-(2-cyanoethyl)-L-cysteine(CyEC), and in hemoglobin (Hb) and plasma protein adducts and urinary metabolites in rats and humans exposed to ACN was developed. ACN in mercapturic acids or proteins was analyzed by capillary gas chromatography (GC) by liberating ACN at a high-temperature in the injector port of GC with or without oxidizing sulfur atoms of the ACN-bound cysteines into sulfoxide form by hydrogen peroxide in vitro. At 350 degrees C, more than 90% of ACN in authentic CyEC was recovered by this method. Increasing a single ip dose of ACN from 5 to 50 mg/kg produced proportional increases in ACN bound to Hb 24 hr after the treatment. The alkylation of plasma protein with ACN was about 1/10 as low as that of Hb. After repeated daily ip doses of 1-10 mg/kg, ACN in Hb decreased with a half-life of about 9 days. ACN was also detected in the blood of workers exposed to ACN for 1 to 10 years at a Siberian synthetic rubber factory.

Acetylcysteine↗
PubMed1992

Translational options for the pir gene of plasmid R6K: multiple forms of the replication initiator protein pi.

The autogenously controlled pir gene of plasmid R6K was believed to encode a single polypeptide that plays multiple roles in the plasmid's biology. We have isolated an opal (op) mutant at the 18th codon of the pir coding frame which does not totally abolish translation of pir mRNA. In extracts of cells containing this mutation two translational products (35 kDa and 30.2 kDa) have been detected. We propose that the 35-kDa polypeptide produced by the pir18 op mutation contains Trp substituted for Arg18 as the result of an opal readthrough. Translation, which results in the 30.2-kDa polypeptide, originates downstream from the UGA stop signal created by the mutation. Moreover, we realize now that the 30.2-kDa polypeptide is also produced in cells containing a wild-type (wt) pir gene. The shorter variant of the pi protein lacks replication initiation and inhibition functions, as well as autorepressor activity in vivo. We also show that an in-frame fusion of seven N-terminal codons of the trpE gene with a pir gene lacking the first two codons produces two polypeptides which replace the 35-kDa pi protein and are of similar molecular weight. Thus, at least three options exist in the translation of the wt pir mRNA. Start codons are most likely at codon positions 1, 6 or 7, and 36 or 38. Each of these five AUG codons is preceded by a consensus ribosome-binding site (RBS).

Bacterial Proteins↗
PubMed1992

Transcription factors in mouse fetal thymus development.

T cell precursors from murine fetal liver enter the fetal thymus where they proliferate, differentiate, and mature. These processes are accompanied by changes in the pattern of transcription factors known to control the expression of specific genes. We have monitored the expression of five different transcription factors during mouse fetal thymus ontogeny: nuclear factor (NF)-kappa B, cAMP-response-element binding protein (CREB), NF-IL-2A, msNF-AT1, and hNF-AT1. NF-kappa B binding activity was not detected in extracts from fetal liver but was present in the thymus at day 14 of embryogenesis. Thereafter, NF-kappa B expression was biphasic, being maximal at 14-16 days gestation and in newborn mice, and decreased during the intermediate gestational stages and in the adult. An inverse correlation was observed between NF-kappa B binding activity in the nuclei and levels of its inactive precursor in the cytoplasm of all samples analyzed. In contrast, CREB activity was uniform throughout thymus development. Similarly, NF-IL-2A activity was detected in fetal liver and thymic extracts from different gestational stages, in approximately equivalent amounts. However, band shift experiments revealed three distinct NF-IL-2A-DNA complexes, whose relative abundance is altered during thymic ontogeny. Likewise, NF-AT1 transcription factor appears to be heterogeneous and includes representatives which are differentially (msNF-AT1) or stably (hNF-AT1) expressed during thymic development. These results are discussed in the context of present knowledge about T cell development within the thymus.

Animals↗
PubMed1990

DNA-RNA complexes that might represent transient attachment sites of nuclear DNA to the matrix.

In this study we describe DNA-RNA complexes in matrix DNA of Friend cells. The presence of such unusual structures is confirmed by the following evidence. When a preparation of matrix DNA is electrophoresed in agarose an RNA component always migrates together with DNA. There should be a close interaction between DNA and RNA in such a preparation because the presence of the RNA component causes resistance of DNA to DNase I and Exo III. An intimate, hybrid-type association of part of the RNA component with DNA is indicated also by the fact that about 20% of this RNA is sensitive to RNase H. By specific inhibition of the RNA synthesis with alpha-amanitin and actinomycin D it was shown that the bulk of associated RNA is transcribed by RNA polymerase III. Hybridization experiments showed similarity between the DNA sequences isolated from the complexes and those from the base of dehistonized DNA loops obtained by high-salt extraction of nuclei. This observation suggests that the complexes might represent attachment sites of nuclear DNA to the matrix: possibly, the attachment is mediated via the RNA component. Experiments with induction of erythroid differentiation indicated that a profound reorganization of the nucleus, accompanying terminal differentiation, leads to a striking reduction in the number of complexes and thus in the number of attachment sites. This suggests that the complexes should function as transient attachment sites.

Amanitins↗
PubMed1990

Epitope mapping of the low-molecular-mass subunits of reverse transcriptase in human immunodeficiency virus type 1 by monoclonal antibodies.

With the aid of monoclonal antibodies to the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1), low-molecular-mass subunits (p29, p32, and p40) were identified in HIV-1 RT purified from HIV (HTLV-IIIB) virions by isoelectric focusing. Epitope mapping with synthetic polypeptides from various regions of the pol gene suggests that the low-molecular-mass subunits result from N-terminal cleavage of the p51 subunit. The subunits could be separated only by SDS-polyacrylamide gel electrophoresis and detected by immunoblotting. They could not be separated on chromatographic columns, suggesting that the subunits are complexed or conformationally arranged in such a way that their separation on the basis of molecular mass is not possible. The molecular mass of the active enzyme eluted from a chromatographic column (Sephacryl S-300) loaded with a mixture of the subunits was estimated to be 100 kDa.

Animals↗
PubMed1989

Lipid peroxidation in acrylonitrile-treated rats, evidenced by elevated ethane production.

The intraperitoneal administration of acrylonitrile (greater than 25 mg kg-1) to rats is associated with an increased production of ethane and a rise of the serum activity of the cytosolic enzyme, sorbitol dehydrogenase. These effects are prevented by pretreatment with vitamin E and the microsomal enzyme inhibitor SKF 525A, but are exacerbated by pretreatment with the microsomal enzyme inducer, phenobarbital. Repeated intraperitoneal administration of acrylonitrile (40 mg kg-1) for four weeks also increases ethane production and serum SDH activity, and produces various morphological changes in liver parenchymal cells (necrosis, increased mitotic activity, increased nucleolar size and myelinic figures in mitochondria) and inhibits the growth of the animals. All these effects are prevented by the administration of vitamin E (190 mg kg-1 i.p., daily) during the last two weeks of treatment. A dose of sodium cyanide (2.5 mg kg-1 i.p.) which leads to a urinary excretion of thiocyanate similar to that found after the intraperitoneal administration of 25 mg kg-1 acrylonitrile, does not stimulate ethane production. This study suggests that the hepatoxicity of acrylonitrile may, at least partly, result from a lipoperoxidation process and is linked with its microsomal oxidative biotransformation.

Acrylonitrile↗
PubMed1989

Infection with the intracellular protozoan parasite Theileria parva induces constitutively high levels of NF-kappa B in bovine T lymphocytes.

The intracellular protozoan parasite Theileria parva causes a lymphoproliferative disease of T cells in cattle and uncontrolled lymphocyte proliferation in culture. We have identified and characterized in infected cells the transcriptional activator, NF-kappa B, whose recognition motifs have been identified in several gene enhancers important for lymphocyte-specific gene expression. NF-kappa B is normally constitutively activated in nuclear extracts derived from B cells and can be induced in T cells and nonlymphoid cells by phorbol esters. Theileria-infected lymphocytes contained constitutively high levels of activated NF-kappa B in nuclear fractions and inactive NF-kappa B in cytoplasmic fractions. The inactive cytoplasmic precursor could be activated by treatment of extracts with deoxycholate, which was shown previously to dissociate NF-kappa B from an inhibitor, I kappa B. Treatment of lymphocyte extracts with 3 mM GTP stimulated NF-kappa B binding to its recognition motif in vitro, thereby distinguishing it from a related nuclear factor, H2-TF1. Selective killing of the parasite, which left the host cells intact, resulted in a rapid loss of NF-kappa B from the nuclear fractions and a slower loss from the cytoplasmic fractions. In parasitized cells, NF-kappa B could not be further stimulated by treatment with 12-O-tetradecanoylphorbol-13-acetate whereas in cells treated to remove the parasite, this compound stimulated elevated levels of NF-kappa B. We propose that high levels of activated NF-kappa B are maintained by the presence of the parasite in infected T cells. Similarly, we propose that the high levels of inactive cytoplasmic precursor are a result of increased synthesis due to the presence of the parasite.

Animals↗
PubMed1988

[Cytogenetic effect of thaliblastine in a culture of human peripheral blood lymphocytes].

The mutagenicity of thaliblastine (Bulgarian potential antitumor drug) was investigated in vitro in lymphocytes from healthy donors, and in vivo in lymphocytes of oncological patients after thaliblastine administration. No increase in the rate of chromosome aberrations was noted with increasing thaliblastine concentrations in vitro and in the course of therapy in vivo. Some polyploid metaphases were found in the lymphocytes of the patients treated with thaliblastine, as a result of the statmokinetic effect of the drug. Thaliblastine exerts extraordinarily slight mutagenic effect, as compared with other cytostatics.

Aged↗