[THE APPLICATION OF DOT-TECHNIQUE FOR DETECTING ANTIGENS OF ADENOVIRUS IN CLINICAL SAMPLES].

The article substantiates possibility of application of point enzyme-linked immunosorbent assay (dot-technique) for detecting viral antigens in samples from patients. To diagnose adenovirus infection conjugate of virus-specific monoclonal antibodies and peroxidase of horse-radish were used The chromatographic rectification of conjugate from free peroxidase permits diminishing background coloring of nitrocellulose membrane and therefore to increase sensitivity. The application of direct conjugates on the basis of virus-specific monoclonal antibodies increases specifcity of dot-technique and significantly shortens time period of analysis. As in case of application of direct conjugates on the basis of polyclonal serum, samples from patients require preliminary processing with detergent for preventing non-specific reactions. The dot-technique demonstrates good coincidence with data of polymerase chain reaction and after clinical trials it can be used in diagnostic of human viral infections.

Dawn simulation vs. bright light in seasonal affective disorder: Treatment effects and subjective preference.

BACKGROUND: Studies comparing the efficacy of dawn simulation to conventional bright light for the treatment of seasonal affective disorder (in parallel groups) have yielded conflicting results. This crossover study investigated treatment outcomes and long-term treatment preference. METHODS: Forty winter depressives were treated for a week with bright light (4.300lx for 30-45min shortly after awakening) or dawn simulation (gradually increasing light during the last 30min of sleep achieving 100lx before alarm beep, with the dawn simulator placed closer to the open eyes for a further 15min: 250lx). The depression level was self-rated using SIGH-SAD-SR. RESULTS: Depression scores reduced similarly following bright light and dawn simulation: for 43.8% and 42.2% (medians), respectively; efficacy ratio was 23:17. The preference was also similar (21:19). Among those who preferred bright light, the most common reason was that they perceived the bright light to be more effective (19/21; it was more effective, p=0.0096; this subgroup tended to have more severe depression) and ease of use (6/21). Among those who preferred the dawn simulator, the reasons were a more "natural" action (9/19), device compactness and/or time-saving (10/19) and in 4 cases where bright light caused eyestrain. LIMITATIONS: Not overhead naturalistic light for dawn simulation, self-rating of depression. CONCLUSIONS: Dawn simulation is similarly effective to bright light in the treatment of winter depression. Patients with more severe depression tended to report greater improvement with bright light; in such cases, this would outweigh the non-clinical advantages of dawn simulation.

[SOME ASPECTS OF NON-SPECIFIC PROPHYLAXIS AND THERAPY OF ESPECIALLY DANGEROUS INFECTIONS].

Recently, due to spread of dangerous and especially dangerous infections much attention is given to development of complex approaches to their prophylaxis and therapy. Data on use of immune modulators, cytokines, probiotics, preparations of plant origin for non-specific prophylaxis of especially dangerous infections are analyzed in the review, and expediency of their combined use with specific and emergency prophlaxis of these diseases is evaluated.

[Mechanisms of formation of post-vaccinal immune response in children immunized with APDT and ADT-M preparations].

AIM: Study the mechanisms of formation of cell and humoral immunity against pertussis, diphtheria and tetanus in children immunized with immunobiological preparations (APDT vaccine and ADT anatoxin). MATERIALS AND METHODS: 30 practically healthy children (6 - 9 years of age) immunized with APDT and ADT-M preparations had TLR2, TLR4 expression determined in mononuclear cells (MNC). Vaccine preparations (APDT, ADT-M, AD-M, AT) and Corynebacterium diphtheriae gravis tox+, C. diphtheriae mitis tox- and Bordetella pertussis 345 were used as ligands. Cytokine production was determined in EIA. Content of anti-diphtheria, anti-tetanus and anti-pertussis antibodies--by PHA reaction and EIA. RESULTS: During stimulation with vaccines and B. pertussis 345 strain MNC were characterized by an increase (p < 0.05) of expression level of TLR2 and TLR4 and did not respond to stimulation with C. diphtheriae gravis tox+ and C. diphtheriae mitis tox- strains. Similar results were obtained during study of cytokine production (TNFalpha, IL-1, IL-6). A direct correlation between levels of antitoxic antibodies against diphtheria and tetanus (R = 0.486), antibacterial antibodies against pertussis and diphtheria was detected (R = 0.529). CONCLUSION: Analysis of cytokine production profile and determination of surface TLR expression can be used during evaluation of functional status of innate immunity cells and intensity of post-vaccinal immunity.

FER kinase promotes breast cancer metastasis by regulating α6- and ß1-integrin-dependent cell adhesion and anoikis resistance.

Metastatic breast cancer cannot be treated successfully. Currently, the targeted therapies for metastatic disease are limited to human epidermal growth factor receptor 2 and hormone receptor antagonists. Understanding the mechanisms of breast cancer growth and metastasis is therefore crucial for the development of new intervention strategies. Here, we show that FER kinase (FER) controls migration and metastasis of invasive human breast cancer cell lines by regulating α6- and ß1-integrin-dependent adhesion. Conversely, the overexpression of FER in non-metastatic breast cancer cells induces pro-invasive features. FER drives anoikis resistance, regulates tumour growth and is necessary for metastasis in a mouse model of human breast cancer. In human invasive breast cancer, high FER expression is an independent prognostic factor that correlates with high-grade basal/triple-negative tumours and worse overall survival, especially in lymph node-negative patients. These findings establish FER as a promising target for the prevention and inhibition of metastatic breast cancer.

Fibroblast growth factor receptors in breast cancer: expression, downstream effects, and possible drug targets.

Cancer treatments are increasingly focusing on the molecular mechanisms underlying the oncogenic processes present in tumors of individual patients. Fibroblast growth factor receptors (FGFRs) are among the many molecules that are involved in oncogenesis and are currently under investigation for their potential as drug targets in breast cancer patients. These receptor tyrosine kinases play a role in several processes including proliferation, angiogenesis, and migration. Alterations in these basal processes can contribute to the development and progression of tumors. Among breast cancer patients, several subgroups have been shown to harbor genetic aberrations in FGFRs, including amplifications of FGFR1, FGFR2, and FGFR4 and mutations in FGFR2 and FGFR4. Here, we review in vitro and in vivo models that have partly elucidated the molecular implications of these different genetic aberrations, the resulting tumor characteristics, and the potential of FGFRs as therapeutic targets for breast cancer treatment.

[Effect of neutrophilokines on functional activity of macrophages during formation of immunity against cholera].

AIM: To study effect of neutrophilokines on functional activity of macrophages (Mph) during formation of immunity against cholera. MATERIALS AND METHODS: In order to obtain peritoneal neutrophils (Nph), 2 ml of 0.1% glycogen solution in buffered with phosphates sodium chloride solution was administered intraperitoneally to 100 outbred mice. Vibrio cholerae 1130 in dose 10 microbial cells/Nph and cholera toxin (CT) in dose 1 or 10 mcg/ ml were used as inducers of neutrophilokines synthesis. Obtained neutrophilokines were assessed on their effect on phagocytic activity of Mph, resistance of these cells to cytotoxic and apoptogenic effects of Vibrio cholerae and CT as well as effect on lysosomal apparatus of Mph. RESULTS: It was established that neutrophilokines induced by Vibrio cholerae and CT stimulate killer activity of Mph and lability of their lysosomal membranes, and suppress programmed death of these cells. CONCLUSION: Results of studies revealed immunoregulatory activity of neutrophilokines relative to Mph and demonstrated ability for cooperation between mono- and polynuclear phagocytes mediated by cytokines and, in particular, neutrophilokines.

[Evaluation of the immunoregulatory activity of neutrophilokine fractions in a macrophage model].

A procedure was proposed to evaluate the immunoregulatory activity of neutrophilokine fractions on a model of macrophages. It was established that all the fractions studied did not affect the absorptive capacity of these cells in both primary and secondary immune responses. At the same time, the majority of neutrophilokine fractions modulated the killer activity of macrophages: they potentiated or inhibited it. The proposed procedure for evaluating the regulatory effect of individual neutrophilokine fractions on a model of studying the killer activity makes it possible not only to characterize their activity, but also to identify helper and suppressor fractions, which discloses approaches to correcting an immune response by means of these fractions.

[Influence of neutrophilokines on population and subpopulation repertoire of lymphocytes and their functional activity during immune response against plague].

Results of experimental study of regulatory effect of nutrophilokines induced by Yersinia pestis EV strain on population and subpopulation repertoire of lymphocytes and their functional activity during immune response against plague infection are presented. It was established that these neutrophilokines stimulate CD4+ and suppress CD8+ lymphocytes. Helper effect of neutrophilokines on functional activity of lymphocytes was more pronounced during secondary than during primary immune response.

[Role of cholera toxin-induced apoptosis in alteration of macrophages in mice].

AIM: To study the role of active programmed cell death induced by Vibrio cholerae antigens in alteration of peritoneal macrophages of experimental animals. MATERIALS AND METHODS: Apoptosis was assessed by cytofluorometric analysis with propidium iodide using cytofluorometer "Coulter" as well as on characteristic morphological changes of cells in stained histological preparations. RESULTS: Performed experiments carried out by both methods provide evidence that V. cholerae and its antigens (cholera toxin, neuraminidase, chitinase, and lypopolysaccharide) cause apoptosis of mice peritoneal macrophages, which leads to their alteration. CONCLUSION: Our results demonstrate that programmed cell death of phagocytes is one of the causes of cytotoxic effect of V.cholerae and its antigens. Performed experiments show the role of apoptosis of macrophages in formation of postimmunization immunosuppression after vaccination against cholera.

Phosphorylation by p38 MAP kinase is required for E2F1 degradation and keratinocyte differentiation.

The transcription factor E2F1 plays key roles in skin homeostasis, and is essential for normal keratinocyte proliferation and epidermal regeneration after injury. We have previously established that, in differentiating keratinocytes, E2F1 activity is controlled by nuclear export and subsequent degradation. These events are triggered by differentiation-induced stimulation of protein kinase C and p38 mitogen-activated protein kinase (MAPK). However, the mechanisms that induce E2F1 export from the nucleus and the role of p38 MAPK in this process are poorly understood. We now describe a novel regulatory pathway for E2F1, which involves phosphorylation by p38. We demonstrate that E2F1 forms complexes with active p38 through regions that exclude the N-terminus of this transcription factor, and that p38 activity is a major contributor to the phosphorylation status of E2F1 in keratinocytes. Using in vitro kinase assays, we identified Ser403 and Thr433 as the residues phosphorylated by p38. The biological significance of these observations is underscored by the inability of E2F1 mutants lacking one or both of these residues to be exported from the nucleus and degraded when keratinocytes receive differentiation stimuli, which results in impaired keratinocyte maturation.

[Preparation of fractions and characteristic of neutrophilokines complex induced by Yersinia pestis EV].

Biochemical and immunobiologic characteristics of fractions of neutrophilokines during primary and secondary immune response against plague infection are presented. Fractions were obtained using gel chromatography from neutrophilokines complex induced by vaccine strain of Yersinia pestis. It was revealed that fractions of neutrophilokines regulate IL-2 synthesis by Th1-helpers, IL-4 and IL-5 synthesis by Th2-helpers and also expression of IL-2 receptors by immunocompetent cells. Helper effect of neutrophilokines' fractions was more pronounced during secondary immune response.

Activation of p38- and CRM1-dependent nuclear export promotes E2F1 degradation during keratinocyte differentiation.

E2F factors modulate a plethora of cell functions, including proliferation, differentiation, DNA repair and apoptosis. We have shown that differentiation in primary epidermal keratinocytes leads to E2F1 downregulation via activation of protein kinase C and p38 mitogen-activated protein kinase. We now demonstrate that E2F1 downregulation in differentiating keratinocytes involves its ubiquitination, as well as proteasomal degradation subsequent to CRM1-dependent nuclear export. E2F1 nuclear export specifically in response to differentiation requires regions adjacent to the cyclin A-binding domain in the N-terminus of this protein. Significantly, inhibition of p38 interferes with nuclear export and degradation of E2F1 during differentiation, but has no effect on E2F1 in undifferentiated cells. Thus, induction of differentiation in epidermal keratinocytes activates a specific program for post-transcriptional downregulation of E2F1, which involves signaling through p38 and activation of nuclear export pathways.

E2F1 stability is regulated by a novel-PKC/p38beta MAP kinase signaling pathway during keratinocyte differentiation.

E2F transcription factors regulate proliferation, differentiation, DNA repair and apoptosis. Tight E2F regulation is crucial for epidermal formation and regeneration. However, virtually nothing is known about the molecular events modulating E2F during epidermal keratinocyte differentiation. Elucidation of these events is essential to understand epidermal morphogenesis, transformation and repair. Here we show that, in differentiating keratinocytes, Ca(2+)-induced protein kinase C (PKC) activation downregulates E2F1 protein levels. Further, we have identified PKC delta and eta as those isoforms specifically involved in induction of E2F1 proteasomal degradation. We also demonstrate that E2F1 downregulation by novel PKC isozymes requires activation of p38beta mitogen-activated protein kinase (MAPK). This is the first example of regulation in the E2F transcription factor family by activation of PKC and MAPK in the context of biologically significant differentiation stimuli in epithelia.

[Development of an automated microbiological analyzer].

An automated hematological analyzer, prototype of a quick microbiological diagnostics system, was tested positively for the ability to differentiate microbial cells (six test-strains with different morphological and tinctorial properties) by geometry and dye intensity.

[Influence of neutrophilokines on the synthesis of lymphokines in the process of the formation of immunity to plague].

The evaluation of the complex of neutrophilokines whose synthesis was induced by Yersinia pestis vaccine strain EV on the production of lymphokines in the process of the formation of primary and secondary immunity to plague is presented. As revealed in this study, neutrophilokines regulate the synthesis of IL-2 by T helpers of type 1, IL-4 and IL-5 by T helpers of type 2, IL-1 by B lymphocytes, as well as the expression of receptors IL-2 by immunocompetent cells. The helper effect of neutrophilokines is more pronounced in the secondary immune response.

[Anticholinergic Effect of a New Antiarrhythmic Class III Drug RG-2]. / Kholinoliticheskaia aktivnost' antiaritmicheskogo preparata III klassa RG-2.

In experiments on isolated rat and rabbit right atrium, a new class III antiarrhythmic drug RG-2 (0,01-1 microM) was shown to have anticholinergic action competing with 0.2-1 mM carbachol. RG-2 (0.1-1 microM) produced dose-dependent increase of APD90% in rat and rabbit atrial cells and had no effects on other action potential parameters. In presence of carbachol RG-2 produced significantly greater increase of APD90% and caused significant increase of APD50%. Thus RG-2 exerts anticholinergic action, which can take important part in RG-2 antiarrhythmic activity.

[Use of detergents for elimination of non-specific reactions of monoclonal antibodies]. / Primenenie detergentov dlia ustraneniia nespetsificheskogo reagirovaniia monoklonal'nykh antitel.

The purpose of the case study was elaboration of methods for removing the false-positive results in the detection of virus antigens that cause acute respiratory lesions both in the plane and dot-membrane variations of immune-enzyme assays of monoclonal antibodies. Modeled experiments showed that the non-specific staining of biological samples, in which the above viral antigen are looked for, is preconditioned by a direct reaction of the substrate with its own oxidizing enzymes and by the binding of monoclonal antibodies with biological macromolecules free of any viral adherence and sorbated in the solid phase. Approaches to solving such issues are described in the paper. They comprise the inhibition of the own oxidizing activity of wash-outs by the substrate and the application of detergents. Different detergents were comparatively analyzed and the optimal compounds were chosen for each variation of immune-enzyme test-systems. The described techniques ensure more reliable results in the viral antigen detection by immune-enzyme assay.

[Comparative evaluation of mono- and neutrophilokins inducing activity of Yersinia pestis antigens]. / Sravnitel'naia otsenka mono- i neitrofilokinindutsiruiushchei aktivnosti antigenov Yersinia pestis.

The results of the comparative analysis of the cytokine inducing activity of Yersinia pestis EV antigens are presented. Y. pestis fraction 1A (F1A) and lipopolysaccharide (LPS) were shown to induce mono- and neutrophilokines, regulating cooperative interaction of phagocytes in the process of immunity formation to plague. Neutrophilokines and monokines exceed in their capacity for inducing F1A such acknowledged inductor as Escherichia coli LPS. As revealed by the comparative evaluation of Y. pestis EV LPS and E. coli LPS, neutrophilokines synthesized under the action of the former preparation, have greater influence on the inhibition of the macrophage migration from the infection focus as well as on digestive activity of these cells (in secondary immune response) and on the labilization of the lysosome membranes of macrophages than neutrophilokines induced by E. coli LPS. At the same time they produce a lesser modulating effect on the killer and chemotactic activity of neutrophils, as well as on the expression of FC receptors (FcR) on their surface in comparison with monokines, synthesized under the influence of E. coli LPS.

[Effect of monokines induced by Yersinia pestis EV on the functional activity of neutrophils during development of antiplague immunity]. / Vliianie kompleksa monokinov, indutsirovannykh Yersinia pestis EV, na funktsional'nuiu aktivnost' neitrofilov pri formirovanii protivochumnogo immuniteta.

A study was made of the Yersinia pestis EV induced monokine complex influence on neutrophil (Nph) functional activity in the course of antiplague immunity formation. The obtained monokines essentially enhance Nph killing, chemotactic activities and Fc-receptor expression, and stimulate lysosome membrane labilization of these cells. The helper effect of the Y. pestis EV induced monokines on Nph functional activity is more pronounced during the secondary immune response, than in the course of the primary one.