[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.

[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.

[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.

[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.

[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.

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.

[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.

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.

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.

The stepwise synthesis of oligo(glycosyl phosphates) via glycosyl hydrogenphosphonates. The chemical synthesis of oligomeric fragments from Hansenula capsulata Y-1842 exophosphomannan and from Escherichia coli K51 capsular antigen.

A stepwise approach has been used in the syntheses of pentamannosyl tetraphosphate HO-[-6Man(alpha)-PO4-]4-6Man(alpha)-OMe and tetra(N-acetylglucosaminyl) triphosphate HO-[-3GlcNAc(alpha)-PO4]3- 3GlcNAc(beta)-OC6H4NO2, which are fragments of the yeast and bacteria extracellular phosphoglycans. Elongation of the chain was performed with the use of suitably protected glycosyl hydrogenphosphonate derivatives for successive introduction of glycosyl phosphate residues. Partially protected monosaccharide derivatives and oligomeric blocks served as hydroxylic components.

Significance of long lasting persistence of influenza virus antigens at the portal of infection and in the spleen of mice.

Distribution of virus, its antigens and development of cellular factors of immunity were followed in the course of different forms of acute influenza virus infection in mice. Long term persistence of influenza virus antigens in the portal of entry and spleen were typical for of acute influenza. Lethal effect of influenza infection was caused by massive lesions induced by the virus and due to cell mediated immune response. The fate of infected individual seems to be decided during the first days of post-inoculation and depends on the ability of the virus to modify the cell membranes of the infected individual.

[Fragments of biopolymers containing glycosylphosphates residues. 9. Synthesis of tetra(2-acetamido-2-deoxy-D-glucopyranosyl)triphosphate-- a tetrameric fragment of the capsule antigen from Escherichia coli K51]. / Fragmenty biopolimerov, soderzhashchikh ostatki glikozilfosfatov. 9. Sintez tetra(2-atsetamido-2-dezoksi-D-gliukopiranozil)trifosfata--tetramernogo fragmenta kapsul'nogo antigena Escherichia coli K51.

Linear tetra(N-acetylglucosaminyl)triphosphate GlcNAc(alpha)-P-3GlcNAc (alpha)-P-3GlcNAc(alpha)-P-3GlcNAc(beta)-ONp, a fragment of the capsular antigen of E. coli K51, was synthesized by the step-by-step approach with the use of the H-phosphonate method, starting the chain from p-nitrophenyl 2-acetamido-4,6-di-O-benzoyl-2-deoxy-beta-D-glucopyranoside. The elongation cycle included the coupling of 2-acetamido-4,6-di-O-benzoyl-2-deoxy-3-O-p-methoxybenzyl-alpha-D-gluc opy ranosyl H-phosphonate with a hydroxyl component in the presence of Me3CCOCl followed by oxidation (I2) and de (methoxybenzylation) (Ce(NH4)2(NO3)6). 2-Acetamido-3,4,6-tri-O-benzoyl-2-deoxy-alpha-D-glucopyranosyl H-phosphonate was employed in the final step. After mild debenzoylation the title tetramer was isolated by anion-exchange chromatography. The data of 1H, 13C and 31P NMR spectra of the synthesized oligomers are discussed.

[Synthesis of phosphodiesters of 2-acetamido-2-deoxy-D-glucose--fragments of polymers of capsules from Escherichia coli K51 and Neisseria meningitidis X]. / Sintez fosfodiéfirov 2-atsetamido-2-dezoksi-D-gliukozy--fragmentov polimerov kapsul Escherichia coli K51 i Neisseria meningitidis X.

Hydrogenphosphonate method was used for synthesis of 4-nitrophenyl 2-acetamido-3- and 4-nitrophenyl 2-acetamido-4-(2-acetamido-2-deoxy-alpha-D-glucopyranosyl phosphate)-2-deoxy-beta-D-glucopyranosides. The glycosides, phosphate diester fragments of the title bacteria capsular antigens, were obtained by H-phosphorylation of the suitably protected 2-acetamido-2-deoxy-beta-D-glucopyranosides with 2-acetamido-3,4,6-tri-O-benzoyl-2-deoxy-alpha-D-glucopyranosyl H-phosphonate in the presence of trimethylacetyl chloride followed by oxidation and deprotection.

[Detection of viral antigens in the lungs and spleen of mice infected with influenza virus]. / Vyiavlenie virusnykh antigenov v legkikh i selezenke myshei, infitsirovannykh virusom grippa.

The data are presented on the possibility of using the dot immunosorption technique for identification of influenza viral antigens in the tissues of infected animal in course of infection. The possibility of quantitative evaluation of the viral antigen contents in the tissues of infected animal permits to use the technique for evaluation of dynamics and rate of viral antigens excretion in course of infection. The information might be useful for studying the pathogenicity of influenza as well as for evaluation of efficiency of immunocorrecting and antiviral preparations. Auxiliary analysis of antigen-containing material by immunoblotting technique has confirmed the viral nature of identified polypeptides. In the future the proposed method may be used to study the viral antigenic structure in infected tissues dependent of the location and time of viral antigens isolation.

[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.

[Cytokines--a general system of homeostatic regulation of cell functions]. / Tsitokiny--obshchaia sistema gomeostaticheskoi reguliatsii kletochnykh funktsii.

The paper reviews biological characteristics of cytokines, which are short living molecules exerting pleiotropic and redundant effects on a variety of target cell types, influencing cell activation and differentiation. Significance of cytokines for patho- and immunogenesis of infections and other diseases is analysed, in addition to further perspectives of cytokine employment for prophylaxis and treatment of various diseases.