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Biomedical subjects

I V Guzhova

Publications and source records attributed to I V Guzhova.

At least 19 recordsLinked to original sources
PubMed2006

Production of heat shock proteins, cytokines, and nitric oxide in toxic stress.

Expression of heat shock proteins Hsp27, Hsp90, and Hsp70 and production of tumor necrosis factors (TNF-alpha, TNF-beta), interferon-gamma (IFN-gamma), interleukin-2, -3, -6, and nitric oxide (NO) were studied under conditions of acute and chronic intoxication of animals with lipopolysaccharides. Injection of endotoxin increased expression of heat shock proteins Hsp70 and Hsp90-alpha in mouse cells. Acute toxic stress also provoked a sharp increase in the production of TNF-alpha, TNF-beta, and NO in mouse cells. The production of other cytokines (interleukins and IFN-gamma) was changed insignificantly. In the model of chronic toxic stress, changes in the production of Hsp70, Hsp90, TNF, and NO were followed during 11 days after the beginning of the toxin injections. The expression of Hsp70 and Hsp90 in acute stress was significantly higher than at the final stage of the chronic exposure. The changes in the TNF and NO productions, on one hand, and the production of heat shock proteins, on the other hand, were synchronous. The findings indicate that repeated injections of increasing endotoxin doses result in a decreased ability of the body cells to respond to stress by overproduction of heat shock proteins, TNF, and NO.

Animals↗
PubMed2005

[Hsp70 chaperone and the prospects of its application in anticancer therapy].

Major stress protein Hsp70 is known to possess two important properties: ATP-dependent activity and protective activity; these two are thought to play a significant role in anticancer therapy. Many malignant tumors contain high amounts of intracellular Hsp70. Moreover, many anticancer drugs themselves are able to elevate Hsp70 expression in tumor cells. Since Hsp70 was found to disturb many signal pathways of apoptosis in many points, the high chaperone expression may lead to an increased resistance of tumor cells to anticancer drugs. On the other hand, when overexpressed by a certain mechanism, Hsp70 is able to emerge at the cell surface by itself or together with tumor antigens and present these to immune cells T-lymphocytes and natural killers, in such a manner that makes cancer cell recognized and abolished. These properties make Hsp70 very promising instrument in designing some novel anticancer vaccines.

Animals↗
PubMed2005

[The balance between Hsp70 and its cochaperones Hdj1 and Bag1 determines its substrate-binding activity].

Heat shock protein Hsp70 presents one of the most effective cell protective systems. Its protective activity is mostly due to the fact that Hsp70 is able to restore native conformation of newly synthesized or damaged proteins. Two other proteins. Hdj and Bag 1, are involved in the process, allowing Hsp70 to perform binding-release cyclec of target proteins. The aim of this study was to investigate interactions between cochaperones Hdj 1 and Bag 1, and the major cell chaperone Hsp in vitro. The accumulation of Hsp70 and Hdj 1 in human erythroleukemia K562 cells was stimulated by heat stress (43 degrees C, 60 min). Cells were collected at certain time periods after heat stress, and amounts of cell chaperones were measured using Western blotting and ELISA assay. The level of Hsp70 chaperone activity in cell extracts was estimated using original technique. The effects of exogenous cochaperones and of their parts on this activity were also investigated. The results of the study indicate that Hsp70 chaperone activity is regulated by the level of its cochaperones, especially Hdj 1. At the same time the amount of ATP appears to be critical for functional activity of Hsp70. Hdj 1 and Bag 1 peptides, which bind to Hsp70 with high affinity, are able to significally reduce its chaperone activity. This finding confirms the possibility of using peptide approach for regulation of Hsp70 function at the cellular and organismal levels.

Adenosine Triphosphate↗
PubMed2004

[Expression of Hsp70 and Hdj1 chaperone proteins in human tumor cells].

Heat shock proteins (Hsp), or "stress proteins", play an important role in maintenance of cellular homeostasis both under normal conditions and during cellular stress. We tested malignant female urogenital and breast tumors for Hsp70 and Hdj1 (Hsp40) chaperones. Immunoenzyme procedure (based on Hsp70 and ATP interaction) was used to assay Hsp70 levels. Hsp70 and Hdj1 expression was higher in malignant cells than in benign ones. We were the first to demonstrate the feasibility of using Hdj1 expression as a novel prognostic factor for neoplastic disease.

Biomarkers, Tumor↗
PubMed2004

[The role of Hsp70 chaperone in the reaction of human leukemic cells to anticancer drugs].

Most of anti-tumor factors are designed to kill selectively cancer cells; in most cases this action is related to the ability of the above substances to induce apoptosis. One of potent anti-apoptotic mechanisms is based on Hsp70 protein. Since the level of this protein is often higher in malignant tumors than in normal tissues, the aim of this study was to establish whether the elevated Hsp70 content may influence the process of apoptosis induced by anti-tumor drugs in cancer cells population. The increase of intracellular content of Hsp70 in human leukemia U-937 cells was attained by a mild heat stress or by transfection of cells with the human hsp70 gene. The elevation of Hsp70 quantity, irrespective of the way it was performed, leads to the inhibition of apoptosis in cells treated with two substances, etoposide or adriamycin. The inhibition of apoptosis was accompanied with the reducing of the share of cells with fragmented nuclei and with the delay in caspase activation. It is suggested that in addition to the previously discovered targets, whose activity is suppressed by the Hsp70 chaperone, this protein can inhibit the activity of caspase-3 and -7; this delays the onset of apoptosis in part of a cancer cell population.

Antibiotics, Antineoplastic↗
PubMed2004

[Dynamics of chaperone complex Hdj1-Hsp70-Bag1 as a response of erythroleukemia K562 cells to heat stress].

Heat shock protein Hsp70 is known to play an important role in cell protection against a variety of harmful factors. This property, at least in part, is due to Hsp70 ability to restore the native conformation of newly synthetized or damaged proteins. In this activity Hsp70 is accompanied by two proteins, Hdj1 and Bag1, that enable Hsp70 to peform cycles of binding-release of target proteins. The aim of this study was to investigate interactions of Hdj1 and Bag1 co-chaperones with Hsp70 in vivo. The accumulation of Hsp70 was stimulated by heat stress, and later, at certain periods following the stress, cell probes were collected for biochemical and microscopic analysis. The data of Western blotting showed that within 24 h after heat shock amounts of Hsp70 and Hdj1 raised to remain at the elevated level for nearly 48 h. Several time points within this period were chosen for analysis of the complexes between Hsp70 and co-chaperones. The data of reciprocal immunoprecipitation/immunoblotting and confocal microscopy showed that Hsp70-Hdj1 complexes were detected primarily at early stage after heat shock, then Hsp70 was preferably bound to Bag1. The dynamics of chaperone complex formation and changes in their intracellular localization are discussed in terms of cell reaction to stress.

Chaperonins↗
PubMed2003

The use of HSP70 for prevention of consequences of unavoidable stress in rats.

We studied whether intranasal treatment with HSP70 can be used for the correction of maladaptive behavior in rats after unavoidable stress. After 3 intranasal injection of HSP70 in a dose of 3.25 microg we observed normalization of adaptive behavior after unavoidable stress and improvement of physiological reserves of the organism.

Administration, Intranasal↗
PubMed2000

[Stress proteins in eukaryotic cells].

The review concerns properties and function of heat shock proteins (hsp) in the eukaryotic cell. The factors inducing Hsps' expression are considered with the emphasis on the fact that most of these agents are able to affect the function of peptide-synthesizing and protein-modifying machineries. The common outcome of this effect is accumulation of wrongly assembled protein structures which results in activation of heat shock transcription factors. The major property of proteins belonging to Hsp70 and Hsp90 is their chaperonic activity or the ability to bind newly synthesized or damaged polypeptides followed by restoration of the native structure of the latter. In this activity Hsp70 and Hsp90 are assisted by other proteins, and the work of such chaperonic systems is described. The function of the major stress protein Hsp70 in the eukaryotic cell was analysed, and on the base of a huge amount of data in has been resumed that proteins of this family constitute one of the most abundant systems of cell protection against cytotoxic factors. In the concluding part of the review we present some ways of application of our knowledge of Hsp in ecology, for analysing biological pollution, and in medicine, for increasing tissue or organismal resistance to injuring factors.

Bacteria↗
PubMed2000

[Induction and accumulation of HSP70 leads to formation of its complexes with other cell proteins].

The expression of a major stress protein Hsp70 may be induced by a great number of cytotoxic agents and also by factors known to cause process of cell proliferation or apoptosis. After induction the synthesis of Hsp70 is reduced to its basic level within first 2-6 hours after inducer application, while the high content of the protein may persist up to several days. The aim of this study was to understand the reason of such a long storage of Hsp70 in U-937 cells induced with three distinct factors. The data of immunoblotting showed that the mild heat shock at 43 degrees C for 60 min, phorbol ester and tumor necrosis factor (the two latter known to induce differentiation and apoptosis, respectively) caused Hsp70 accumulation, the protein level being high within 48 hours and then slowly declined to the basal level. According to the results of chromatography on anti-Hsp70 antibody-Sepharose gel, approximately 50% of the protein was retarded by the gel, while the other part was not recognized by immunoglobulins. Among proteins bound to the Hsp70, three polypeptides were identified as actin, p65 and c-Rel, two latter being constituents of NF-kappa B regulatory complex. The data demonstrate that besides its traditional target, i.e. newly synthesized or abnormal polypeptides, Hsp70 is able to bind mature, active proteins.

Chromatography, Affinity↗
PubMed2000

[Effect of heat shock on cell differentiation and apoptosis in U-937 cells].

One of the most abundant cell protective systems is based on an inducible member of Hsp70 family of stress proteins. Proteins belonging to the family are known to participate in all processes of cell physiology including differentiation and apoptosis. Here data are presented concerning effect of heat shock accompanied by a high-level accumulation of Hsp70 on the phorbol ester-induced expression of surface antigens and on TNF-alpha mediated apoptosis. The data showed that heat shock at 43 degrees C for 60 min reduced the expression of CD11c and CD23 surface markers pre-established by phorbol ester; the latter is known to induce macrophage-like phenotype by 70-80% of the original level. Heating in the same conditions was also shown to markedly delay the outcome of apoptosis stimulated by TNF-alpha. Suggesting that Hsp70 by its binding transcription activators of NF-kappa B complex might transiently suppress both the processes, we determined the amount of p65 and c-Rel in nuclear fractions of cells subjected to various stimuli. It was found that both proteins were retarded in the cytoplasm and were not transported to nuclei in cells heated before the administration of PMA or TNF. It is concluded that Hsp70 accumulating in higher amounts is able to transiently protect cells of TNF-mediated cytotoxic effect by physical association with the proteins that serve as modulators of apoptosis.

Apoptosis↗
PubMed2000

[Induction of apoptosis in murine myeloma cells NS0/1, transfected with the gene for the basic heat shock protein HSP70i].

Murine myelomas are rare cell variants deficient in inducible isoform of Hsp70 that protects cells from injury. In these cells Hsp70 is absent and is not induced under stress conditions. In this study myeloma cells NS0/1 were transfected with hsp70, and their susceptibility to apoptosis was challenged by serum deprivation or hydrogen peroxide. Expression of Hsp70 in NS0/1 cells made them more resistant to apoptosis in serum-free medium but did not affect their response to hydrogen peroxide. Hsp70 involvement in the protection of myeloma cells from apoptosis caused by different agents is discussed.

Animals↗
PubMed1998

Effects of exogenous stress protein 70 on the functional properties of human promonocytes through binding to cell surface and internalization.

The presence of antibodies against the major stress protein, Hsp70, in patients with autoimmune diseases led us to hypothesize that Hsp70 may occur extracellularly, and could exert chaperoning and regulatory effects on various cells. We examined the action of pure Hsp/Hsc70 on the main physiological functions of human promonocytic U-937 cells. The protein was isolated from calf muscle and was shown to be a mixture of inducible Hsp70 (60%) and constitutive Hsc70 (40%) isoforms. It was observed that the addition of the protein up-regulated two major monocyte/macrophage differentiation markers, CD11c and CD23, by 20-35%, while it had no effect on CD14. The experiments performed to investigate the influence of Hsp/Hsc70 on the reaction of U-937 cells to differentiation stimuli demonstrated that the addition of the protein prior to PMA was able to inhibit binding of proper transcription factors to double-symmetry and cAMP-response elements of the c-fos early response gene promoter. Administration of exogenous Hsp/Hsc70 prior to treatment with the tumor necrosis factor-alpha significantly lowered the number of apoptotic and necrotic cells. In no case did the control protein, ovalbumin, taken in the same concentration give a comparable effect on U-937 cells. Since the Hsp/Hsc70 effects occurred within the first hour of co-incubation, and therefore they might be explained by its interaction with the cell surface, we assayed binding of the biotinylated protein to U-937 cells by immunoenzyme assay, flow cytometry and indirect immunofluorescence. Using these three techniques we were able to detect Hsp/Hsc70 bound to cells after a 20 min incubation. According to flow cytometry data, at this time 32% of cells were positively stained with streptavidin-FITC. Immunofluorescence studies demonstrated Hsp/Hsc70 bound to the cell surface after a 20 min incubation followed by induction of patch and cap-like structures. One hour later, the majority of the protein had been internalized by U-937 cells.

Animals↗
PubMed1997

Accumulation of major stress protein 70kDa protects myeloid and lymphoid cells from death by apoptosis.

The major heat shock protein, hsp70, is known to contribute to the mechanisms of cell protection against a variety of stress and cytotoxic factors, providing an increase of cell survival. Whether hsp70 could be implicated in the rescue of cells from stress-induced death proceeding on apoptotic pathway is not well established. Here we report that susceptibility of myeloid and lymphoid cell lines to apoptosis induced by heat shock or ethanol coincides with hsp70 content and can be modulated by changes in expression of this protein. Cells of lymphoid and myeloid lines differing in basal and inducible level of the protein were tested. The cells containing higher amounts of hsp70 (U937, Jurkat, Molt4) were more resistant to the apoptosis-inducing stimuli then cells which accumu-late lower amounts of the protein (HL60) and especially those lacking the protein (NSO). Inhibition of hsp70 accumulation by quercetin made cells more susceptible to the same apoptotic inducer. Enhancement of hsp70 expression by previous heating or by liposomal delivery of the exogenic protein to the cells lacking hsp70 made them more resistant to apoptosis. The possible mechanisms of the hsp70 protective effect in apoptosis are discussed.

Journal Article↗
PubMed1997

Major stress protein Hsp70 interacts with NF-kB regulatory complex in human T-lymphoma cells.

Polypeptides belonging to the Hsp70 major stress protein family and to the NF-kB/Rel multi-functional regulatory complex are known to be involved in cellular defense mechanisms. It was suggested that both systems may interact in cells that respond to injuring stimuli. To check this, Molt4 human lymphoma cells were heated at 43 degrees C for 15 min and, after a 6 h post-shock recovery period, the cells were activated with phorbol ester or bacterial lipopolysaccharide. It was found that mild heat shock caused a substantial increase of the intracellular Hsp70 content with the concomitant suppression of NF-kB complexes, though the latter was properly activated in non-stressed cells. After a 24 h period of being inactive the complex fully recovered its activity and p65 and c-Rel subunits migrated to the nucleus. This new active period lasted even longer than that in non-heated control cells. As this suggested the existence of a Hsp70-related mechanism of NF-kB/Rel complex retention in cytoplasm, we carried out immunoprecipitation with the use of anti-Hsp70 and anti-Rel antibodies. All three Rel family members p65, c-Rel, p50, but not their precursors and IkB alpha inhibitory protein were shown to co-precipitate with the stress protein and anti-Hsp70 antibodies from both heated and non-heated cells. We conclude that the Hsp70 stress protein may confer a new mechanism of NF-kB regulation in cells affected by elevated temperature or other factors related to the cellular response to stress.

Blotting, Western↗
PubMed1992

Antibody against p58 surface antigen of RA-2 rat rhabdomyosarcoma cells inhibits their metastatic activity.

Antibodies generated against the whole membrane preparation isolated from rhabdomyosarcoma RA-2 cells were shown by immunoblotting and immunoaffinity chromatography to recognize 58-kDa polypeptide, p58. The latter was confirmed to be a surface molecule in a test of radioiodination of RA-2 membrane by lodogen. The antibodies added to a suspension of RA-2 cells before their inoculation into rats decreased metastatic activity 50-fold without any noticeable influence on RA-2 proliferation level and viability. The data indicate that masking of p58 surface antigen by antibodies could make RA-2 cells unable to form experimental metastases in lung. We suggest that p58 may participate in the specific recognition by RA-2 of lung endothelial cells.

Animals↗
PubMed1992

[Interclonal and interpopulational differences in the frequency of spontaneous karyotypic changes in cell populations of transplantable rat rhabdomyosarcoma RA-2].

Rat organospecific transplantable RA-2 rhabdomyosarcoma substrains RA-2H (high metastatic potential), RA-2L (low metastatic potential) and RA-2T (thermal resistance) were investigated using single cell cloning technique for frequency of micronucleated cells (FMC). Significant interclonal and interpopulational differences were established. Average FMC for RA-2H was 2.96 +/- 0.13%; 5.94 +/- 0.24% for RA-2L and 1.89 +/- 0.12% for RA-2T. Clones showed 5-10 times differences in FMC in each substrain. FMC versus growth time and clone size was studied in RA-2H: average FMC dropped from 2.7% on day 9 to 0.5% on day 20 after administration and the lowest values were recorded in larger clones at each stage. Perspectives of FMC studies in tumor cell populations are discussed.

Animals↗
PubMed1990

[Clonal analysis of karyotypic instability and metastatic potential in tumor cell populations].

Rat organospecific transplantable RA-2 rhabdomyosarcoma strains RA-2H and RA-2L were investigated using single cell cloning technique for "Micronuclear containing cell frequency" (MCF) and "Metastatic potential" (MP). The RA-2H was obtained from RA-2 during selection for increasing metastatic potential, whereas the RA-2L was the result of selection in opposite direction. MP was assessed by lung colony formation technique and in the case of RA-2H, it was one hundred times greater than in the case of RA-2L (2490 +/- 280 and 22 +/- 11 lung colonies i.v. after injection of 10 cells, respectively). Average MCE for RA-2H was 3 and for RA-2L 5.3%. Also, significant differences were observed between these substrains in percentage of anaphase and telophase cells with bridges and fragments (24 +/- 4 for RA-2H and 56 +/- 5% for RA-2L, P less than 0.05). The selection of RA-2H clones with high level of MCF was successful. In the population obtained MCF was increased up to 7.8% and MP decreased (79 +/- 44). So, the selection for decreasing of MP led to significant increase in genome instability characteristics and the selection for increasing of MCF--to the abrupt decline of MP, which proved the existence of negative correlation between these two characters in populations studied. It is concluded that correlation between malignancy and genome instability in tumor populations with different levels of malignancy is not, as a rule, positive.

Animals↗