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S V Komisarenko

Publications and source records attributed to S V Komisarenko.

18 recordsLinked to original sources
PubMed2005

Overexpression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-4 in the human breast and colon malignant tumors.

6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase (PFKFB) is a bifunctional enzyme, which is responsible for maintaining the cellular level of fructose-2,6-bisphosphate, a powerful allosteric activator of glycolysis. We describe herein the overexpression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-4 (PFKFB-4) isozyme in the human breast and colon malignant tumors as compared to corresponding non-malignant tissue counterparts. We have shown also that breast malignant cell line MCF7 constitutively express PFKFB-4 mRNA and that the expression of this gene is highly induced by hypoxia. Overexpression of PFKFB-4 transcript levels in breast and colon malignant tumors correlates with enhanced expression of PFKFB-3, hypoxia-inducible factor (HIF)-1alpha and known HIF-1 dependent genes glucose transporter 1 (Glut1) and vascular endothelial growth factor (VEGF). Thus, our data clearly demonstrates overexpression of PFKFB-4 mRNA and protein in the breast and colon malignant tumors.

Breast↗
PubMed2004

Two monoclonal antibodies to D-dimer-specific inhibitors of fibrin polymerization.

D-dimer of human fibrin was used as antigen to obtain monoclonal antibodies (mAbs). We have obtained 16 hybridomas producing mAbs of different specificity. Only two of these mAbs inhibited fibrin polymerization. They are of the IgG-class. One mAb (II-4d) inhibited fibrin polymerization to 100% and another (II-3b) to 60% at a molar ratio mAb/fibrin=1.0. Fab-fragments of these mAbs inhibited fibrin polymerization completely at the same molar ratio. The epitopes for the mAbs studied are situated in the NH2-terminal part of the gamma-chain in fibrin D-domain. Electron microscopy showed that fibrin was in monomeric form in the presence of these mAbs or their Fab-fragments. Thus, these mAbs stop the initial step of fibrin polymerization, i.e. protofibril formation. Only one site of protofibril formation is known now in COOH-terminal half of the D-domain gamma chain named "a" site, which is complementary to the "A" site in the central E-domain of fibrin molecule. Our experiment with immobilized GPRP showed that the "a" site in fibrin D-fragment preserved its binding activity to GPRP when the D-fragment was complexed with mAbs-inhibitors of fibrin polymerization. Thus, these two mAbs inhibit fibrin polymerization not by blocking the sites "a" but either by blocking another (not "a") specific site in D-domain or by steric hindrance of highly organized fibrin polymerization process.

Antibodies, Monoclonal↗
PubMed2002

A neoantigenic determinant in the D-dimer fragment of fibrin.

Monoclonal antibody (mAb) III-3b binds D-dimer with K(d)=1.4 x 10(-10) M without cross-reaction with fibrin(ogen). The epitope for this mAb is in Bbeta134-190, presumably in Bbeta155-160. The latter site is buried in the coiled coil structure of fibrin(ogen) but it is exposed as a neoantigenic determinant in D-dimer upon plasmic lysis of fibrin. mAb III-3b may be used as a tool for immunodiagnostic quantification of D-dimer in blood plasma.

Antibodies, Monoclonal↗
PubMed2002

Polymerization sites in the D-domain of human fibrin(ogen).

The present work deals with localization of previously unknown polymerization sites of the fibrin DD-fragment. D-dimer we obtained has a pronounced inhibitory effect on fibrin polymerization (IC50 = 0.06 microM). The inhibitory effect of the DD-fragment disappeared after reduction and carboxymethylation. However, polypeptide chains betaDD (Bbeta134-461) and gammagammaDD (gamma63-411)2 of the DD-fragment, isolated by preparative electrophoresis, displayed their inhibitory activity. For instance, the rates of fibrin protofibril lateral association were decreased twice in the presence of betaDD and gammagammaDD chains at their molar ratios to fibrin of 0.40 and 0.15, respectively. The IC50 values for betaDD and gammagammaDD were 0.24 and 0.10 microM, respectively. Highly specific inhibition of protofibril lateral association suggests that the protofibril lateral association sites are located in Bbeta134-461 and gamma63-411 regions of the fibrin D-domain. Our data confirm those reported by Doolittle et al. regarding the gamma-chain and a hypothesis about beta-chain of fibrin D-domain (Yang, Z., Mochalkin, I., and Doolittle, R. F. (2000) Biochemistry, 97, 14156-14161).

Biopolymers↗
PubMed2000

[Monoclonal antibodies as an instrument to study fibrin polymerization].

Examples of using monoclonal antibodies (MAb) for studying the fibrin polymerization mechanism are considered. MAb with epitopes situated in the fibrin polymerization sites or in the recognition sites of enzymes thrombin, plasminogen, and factor XIII, which are the functional partners of fibrin, are primarily discussed. The MAb to epitopes in various regions of A alpha, B beta, and gamma polypeptide chains of the functionally important E, D, and alpha C domains of fibrin are successively described.

Amino Acid Sequence↗
PubMedDate not supplied

[O.V. Palladin Institute of Biochemistry of the Ukrainian National Academy of Sciences--75 years].

The article deals with a short record including the information about the history of creation and main developmental stages of the first and presently the single in Ukraine Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine. The record contains the main achievements of the Institute scientific subdivisions for the 75 years period of their creative work in the field of the most urgent problems of fundamental and applied biochemistry. As well it displays the information about comprehensive and extensive publishing and educational activity conducted by the staff,, the work on training the scientific manpower of the highest qualification--Doctors of Science (D.Sc.) and Philosophy Doctors (Ph. D.), active participation of the research scientists in attracting to science the talented young people representing the students and school-children taking interest in the biological chemistry. The article shows the great role of the Institute famous scientists-biochemists in forming some scientific schools on functional biochemistry acknowledged in the world biochemistry. There are also some data about the each of nine scientific departments of the Institute for the period of last 20-25 years of their activity in the field of the most urgent problems of the contemporary biochemistry. These are the following Departments: Neurochemistry, Metabolism Regulation, Muscular Biochemistry, Protein Structure Functions, Coenzymes Biochemistry, Enzymes Chemistry and Biochemistry, Molecular Immunology, Biochemistry of Lipids, Sensor and Regulator Systems Biochemistry. There is also a short review regarding the history of foundation, development and scientific achievements of the Institute division in L'viv including four scientific departments: Biochemistry of Cellular Differentiation, Biochemical Genetics, Regulation of Low Molecular Compounds Synthesis, Regulation of Cells Differentiation.

Academies and Institutes↗
PubMedDate not supplied

[Department of Molecular Immunology--25 years].

Department of Molecular Immunology of Palladin Institute of Biochemistry was founded 25 years ago as the Laboratory of Immunochemistry uniting, at the beginning, a small group of scientists. In 1982 the Laboratory was transformed into the Department of Molecular Immunology. The first research was made on studding the biological effect and the mechanisms of action of phosphonates and bisphosphonates--phosphororganic analogues of inorganic pyrophosphate or closely related in structure molecules having P-C bond instead of P-O bond. As a result the immuno-modulatory activity of methylene-bis-phosphonic acid was discovered and a set of immuno-vector, immuno-toxin and anti-tumour constructs were designed. The researchers of the Department were the first in the USSR who started the biochemistry of activated lymphocytes, immunochemical analysis of peptides and proteins, implemented immunoenzyme methods and cytofluorimetry to study immunocompetent cells. In particular, monoclonal and polyclonal antibodies were obtained and used to study the antigenic structure of neurotoxin apamin, cytochrome c, neurospecific proteins S-100 and 14-3-2, nicotinic acetylcholine receptors, pertusis and diphtheria toxins, different antigens of Mycobacterium bovis and of fibrin(ogen) and its fragments and others. The new site of fibrin polymerization was localized. The novel information about the nature of polyreactive immunoglobulins and their influence on the development of the humoral immune response was obtained. A range of reagents for immunodiagnostic tests were developed as the outcome of this research work. The negative effect of small doses of radiation on natural immunity in people working during the cleaning of Chernobyl nuclear plant was first demonstrated by the researchers of the Department in co-operation with their colleagues from a couple of Kiev immunology laboratories. In particular the suppression of NK cells function and structure was shown.

Academies and Institutes↗
PubMedDate not supplied

[Complement-binding and immuno-modulating properties of polyreactive immunoglobulins].

New data concerning biological properties of polyreactive immunoglobulins (PRIG) were obtained as a result of treatment of mouse serum immunoglobulins by 4 M KSCN and are presented in the paper. In particular, the capacity of PRIG to bind C1q, the subunit of the first component of complement was studied. It was shown that PRIG's binding capacity to C1q is similar to that of intact immunoglobulins. Intravenous administration of PRIG into mice together with either sheep red blood cells or heat-inactivated staphylococcal bacteria did not affect the immune response to these antigens. Meanwhile, the same administration of PRIG together with the purified protein derivate of tuberculin resulted in 10-fold increase of mouse antibody response to PPD. These results demonstrate that PRIG can have some immuno-modulating properties concerning low-immunogenic antigens.

Adjuvants, Immunologic↗
PubMedDate not supplied

The influence of high ambient glucose level on the production of pericellular glycosaminoglycans by cultured endothelial cells.

The 14C-acetate metabolic labeling of glycosaminoglycans (GAGs) was used to investigate the effect of high glucose level on the production of hyaluronic acid (HA), heparan sulphate (HS), chondroitin sulphate (CS) and dermatan sulphate (DS) by human immortalized umbilical vein endothelial cells. It is demonstrated that 30 mM glucose decreased the accumulation of HS and increased the accumulation of CS and DS in the cell layer, pericellular matrix and conditioned medium in 48 h of incubation. The modulation of the overall metabolism of sulphated GAGs by high glucose is in contrast to the observed redistribution of HA from the conditioned medium to the pericellular matrix of endothelial cells. The preincubation at 30 mM glucose increased also the attachment of hyaluronidase-treated endothelial cells to HA-coated surface and had no effect on the cell attachment to poly-D-lysine, indicating the alterations of CD44 binding to immobilized HA. The treatment of endothelial cells with p-nitrophenyl-beta-D-xylopyranoside, which inhibits the coupling of CS to the core protein, attenuated high glucose-induced pericellular HA accumulation and decreased cell attachment to HA-coated surface. It is supposed the implication of CD44-related CS in the accumulation of pericellular HA by endothelial cells exposed to high glucose level.

Cell Adhesion↗
PubMedDate not supplied

[Thrombospondin-1-dependent mechanism of transforming growth factor beta-1 activation in cultured human endothelial cells].

Using two immortal human umbilical vein endothelial-derived cell lines we investigated the thrombospondin-1 (TSP-1)-dependent mechanism of the transforming growth factor beta-1 (TGFbeta1) activation. Exogenous human platelet TSP-1 was demonstrated to increase (1.7-2.0-folds) the concentration of active TGFbeta1 produced by both cell lines, but did not influence the concentration of total TGFbeta1. The process of TGFbeta1 activation is known to be regulated by G418GWSHW423 fragment of TSP-1 molecule. The simultaneous addition of TSP-1 and 100-fold molar excess (about TSP-1) of the synthetic GGWSHW peptide abolished TSP-1-induced TGFbeta1 activation. GGWSHW peptide inhibited the basal activation of TGFbeta1 in endothelial cells, whereas the secretion of total TGFbeta1 and TSP-1 was not altered. We conclude that TSP-1-dependent mechanism of endogenous TGFbeta1 activation is operation in human endothelial cells.

Cell Culture Techniques↗
PubMedDate not supplied

[The role of nicotine in regulation of lymphocyte proliferation].

The effect of nicotine on both the expression of nicotinic acetylcholine receptors (nAChRs) and proliferation of hybridoma cells and normal mouse lymphocytes has been investigated. By means of immunoenzyme assay, nicotine was shown to regulate the number of nAChRs in both hybridoma cells and normal rat splenocytes. According to the data of triazolyl blue inclusion and ELISpot assay, nicotine stimulated proliferation of both hybridoma cells and normal plasma cells generated in the course of immune response in vivo. The cell sensitivity to nicotine depended on the number of nAChRs expressed on the membrane, as well as on their functional activity affected, in particular, by adhesive contacts. The use of the open channel blocker benzohexonium revealed that proliferative signal through nAChR in hybridoma cells was mediated by ion channel opening. The data obtained demonstrate the proproliferative role of nicotine for B lymphocytes, and may account for the development of lymphoproliferative disorders in tobacco smokers.

Animals↗
PubMedDate not supplied

Novel and simple ELISA-based method for antibody affinity determination.

New coordinates for antibody affinity determination by ELISA are suggested. The suggested approach is very simple but at the same time it is more convenient and is deprived of the drawbacks inhered in the earlier suggested methods. The examples of antibody affinity determination by the suggested methods for both simulative and experimental binding curves are considered. It was demonstrated that the suggested methods allow getting more precise values of antibody affinity.

Animals↗
PubMedDate not supplied

[Quantification of D-dimer and soluble fibrin in blood plasma of people with ischemic heart disease and hypertension].

The method of D-dimer quantification in the human blood plasma has been developed using monoclonal antibodies 111-3b and II-4d. The method has been verified on the blood plasma of the patients with ischemic heart disease with and without stenocardia and with hypertension. The results showed that at ischemic heart disease with and without stenocardia and at hypertension the quantities of D-dimer in the blood plasma were generally less than the highest normal level 500 ng/ml (64.3%, 76.2% and 95%, correspondingly). The semiquantitative measurements of soluble fibrin levels in blood plasmas of the patients with ischemic heart disease and hypertension have been performed. It has been shown that the quantity of soluble fibrin at these diseases range greatly from < 0.03 mg/ml to 0.15 mg/ml. There was no correlation between the quantities of D-dimer and soluble fibrin in blood plasmas of the patients. Electrophoresis in PAAG with SDS showed that the soluble fibrin at these diseases had the mo- lecular mass of the fibrin (ogen). Thus the soluble fibrin in blood plasmas analysed consisted mainly of fibrin desAA oligomers (may be with fibrinogen incorporation) which are not stabilized by the factor XIIIa.

Blood Protein Electrophoresis↗
PubMedDate not supplied

[HIF-1alpha, HIF-2alpha and VHL mRNA expression in different cell lines during hypoxia].

Hypoxia inducible factor-1alpha (HIF-1alpha) mRNA expression is significantly decreased under hypoxia in different cell lines exposed directly to hypoxia or treated with dimethyloxalylglycine which mimics hypoxic effects under normoxic conditions. However, the decreased expression of HIF-1alpha mRNA is accompanied by an increase of HIF-1alpha protein (pHIF-1alpha) level as well as by overexpression of known HIF-dependent genes (VEGF, Glut1, PFKFB-3 and PFKFB-4) under hypoxic conditions or with the use of dimethyloxalylglycine. Expression of HIF-1alpha mRNA also depends on iron because desferrioxamine and cobalt chloride produce similar to hypoxia effects on the levels of this mRNA. It was shown that HIF-1alpha mRNA expression did not change significantly in some cell lines (SKBR3, MDA-MB468 and BT549) under hypoxia. However, in these cell lines hypoxia decreases expression of HIF-2alpha mRNA, another member of HIF-alpha gene family, as a result of cell specific regulation of HIF-alpha genes under hypoxia. Moreover, hypoxia slightly induces expression of PFKFB-4 mRNA in SKBR3, MDA-MB468 and BT549 as compared to other cell lines where this effect of hypoxia was much stronger and adaptation to hypoxia is controlled by HIF-1alpha. Hypoxia slightly reduces expression of tumor suppressor VHL which targets HIF-1alpha for ubiquitination. Thus, our results clearly demonstrated down regulation of HIF-1alpha or HIF-2alpha in different cell lines by hypoxia.

Amino Acids, Dicarboxylic↗
PubMedDate not supplied

Specific identification of Mycobacterium bovis by monoclonal antibody-based enzyme immunoassay.

To improve identification of M. bovis, rabbit immune sera and mouse monoclonal antibodies against M. bovis-secreted protein antigens were used in the enzyme-linked assay (ELISA). In western blot analysis, M. bovis-specific epitopes within culture filtrate proteins were demonstrated to be mostly concentrated on the immunodominant 35-38 kDa antigen. Polyclonal and cross-reactive monoclonal antibodies were shown to be able to bind to all mycobacterial strains tested in ELISA (M. tuberculosis, M. bovis, M. kansasii, M. marinum, M. avium, M. smegmatis), but not to bacteria of the other genera. Among the monoclonal antibodies against M. bovis specific antigen, 2-6B was found to be the only one which could evidently react with whole cells of M. bovis in ELISA, but not with those of the other mycobacteria including M. tuberculosis.

Animals↗