47 research outputs found
Morphofunctional characteristics of blood cells in reconvalescents after suffering COVID-19
Complexity and multifactorial nature of potential pathogenic consequences of SARS-CoV-2 infection in human body, discovery of new virus-induced mechanisms triggering a cascade of pathological responses in the cells of host organism leading to development of multiple organ failure elicited increasing interest in morpho-functional state of blood cells in reconvalescent persons after COVID-19 infection. The aim of the present work is to characterize morphofunctional pattern of blood cells at different periods of recovery, depending on the severity of COVID-19. We examined 55 convalescents after bearing COVID-19 infection: Group I included the convalescents 30 days after the disease (n = 39); Group II consisted of the persons 60 days after recovery (n = 16); Group III included clinically healthy volunteers with no history of clinical SARS-CoV-2 infection (n = 11). The cells were examined by means of Olympus CX41 microscope (Olympus, Japan), and VZ-C31S digital videocamera (VideoZavr, Russia) using the VideoZavr software (version 1.5). Assessment of neutrophil populations in the whole blood samples was performed with BD Accuri C6 Plus flow cytometer (USA) with automatic differentiation of cells between lymphocytes and monocytes, according to the degree of granularity. Cytokine production was determined using commercial kits for detection of IFNγ, TNFα, IL-4, IL-8, IL-10 (JSC Vector-Best, Russia), IL-17A (eBioscience, Austria) was assayed with automatic enzyme immunoassay analyzer “LAZURIT” (Dynex Technologies, USA). Among the convalescents who suffered the moderate-degree COVID-19 (45.5% and 50% of cases, respectively) on days +30 and +60 after clinical recovery, a significantly increased ratio of morphologically altered forms of erythrocytes (echinocytes, ovalocytes, dacryocytes, codocytes) was noted as compared with group III (p = 0.00001 and p = 0.001, respectively). Regardless of clinical severity of the disease; a mean of 40.6% convalescents from groups I and II had moderate disturbances in the neutrophil morphology (cytoplasmic vacuolization, chromatin decondensation at the pre-netosis stage, transformation of cells by the netosis type), and, in 27.4% of cases, the areas of neutrophilplatelet aggregation were seen. In blood supernates from recovered patients, we have revealed a significantly decreased content of IFNγ (P = 0.02), TNFα (p = 0.03), IL-10 (p = 0.04) and IL-17A (p = 0.02). The revealed morphological and functional changes in blood cells in the persons who underwent COVID-19 infection suggest long-term maintenance of toxic damage to erythrocytes, neutrophils and lymphocytes over the recovery period. The effects of the detected morphological and functional disorders of blood cells following COVID-19 recovery leading to increase in blood viscosity and microcirculation, formation of neutrophil-platelet aggregates, may cause higher risks of thrombotic complications at the long-range period as well as decreased levels of regulatory cytokines, thus confirming slow recovery of the lymphocyte populations (Th1, Th2, Th17) of the immune system
EFFECT OF IMMUNOMODULATORS AND ANTI-TULAREMIA VACCINATION ON APOPTOSIS AND LYSIS OF SPLENOCYTES INTERACTING WITH TULYARIN IN VITRO
Objective of the study is to evaluate the effect of immunomodulators on the intensity of the post-apoptotic lysis of sensitized organism leukocytes in the presence of specific antigens of tularemia microbe in vitro.Materials and methods. Flow cytometry method was used to determine the relative content of apoptotic and proliferating splenocytes obtained from mice, immunized against tularemia against the background of immunomodulation.Results and conclusions. Obtained is the evidence that is consistent with modern data on the massive leukocyte apoptosis and post-apoptotic leukocyte autolysis (secondary necrosis) in case of tularemia infection. Given the important role of secondary necrosis in the systemic inflammatory response development, the use of immunomodulators suppressing macrophage apoptosis and dead leukocyte lysis, emerging in the course of interaction with Francisella tularensis antigens, may be promising in order to reduce the live tularemia vaccine reactogenicity
Zymography: developing of the enzyme soil activity visualization method
The enzymes produced by the soil biota are a key link in the regulation of biochemical processes. The soil enzyme activity can be visualized with zymography, a method based on using fluorescent substrates and obtaining two-dimensional images (zymograms). A variant of a zymographic measuring system has been proposed. Characteristics of lighting, photographic equipment and shooting modes, reagents preparation and calibration are presented. Preparing and analyzing soil samples of different texture (sand and clay loam) and processing the study results have been described. The ways of introducing the substrate are considered in this study, namely pipetting, short-time dipping, and saturation. An analysis of the kinetics of incubation of samples was carried out. The possibilities and disadvantages of the method were also considered and options for solving possible methodological problems during the analysis were proposed. The zymography is a promising method that allows comparing data with the results of other methods. The use of neural network technologies makes it possible to obtain the volumetric distribution of soil enzymes with high reliability. The soil zymography requires qualitative preparatory work and extreme accuracy during the analysis. It is necessary to ensure maximum contact between the substrate and the soil, as this is one of the key factors determining the quality of the results. The most optimal way to introduce the substrate is to saturate the membranes with substrate solution for 60 minutes. At this stage of the development of the method, it is not possible to establish a universal sample incubation time, since this depends on characteristics of both the studied soils and the experiment conditions. Also, it is necessary to document the conditions in detail for discussion the study results
IDENTIFICATION OF HLA II CLASS GENE ASSOCIATIONS OF THE MAIN HISTOCOMPATIBILITY COMPLEX WITH PECULIARITIES OF IMMUNE RESPONSE IN PERSONS VACCINATED WITH LIVE PLAGUE VACCINE IN THE REPUBLIC OF KALMYKIA
Objective of the study was to determine allelic variants of HLA II class haplotypes in persons living in the Republic of Kalmykia in the territory of Pre-Caspian sandy natural focus of the plague, immunized for epidemic reasons with live plague vaccine and search for associations of HLA class II haplotypes with peculiarities of post-vaccinal immunity development.Materials and methods. 20 individuals took part in the study. HLA typing was performed applying multiplex PCR. Production of immune-regulatory cytokines and antibody titers to fraction 1 of the plague microbe was determined using enzyme immunoassay. Statistical processing of the results was carried out using standard programs.Results and conclusions. Allelic variants of haplotypes HLA-DQA1, HLA-DQB1 and HLA-DRB1 class II of the main histocompatibility complex of 20 persons residing in Lagansky and Chernozemelsky districts of the Republic of Kalmykia have been identified. Determined have been the differences in the ratio of allelic variants of HLA-DQA1 and cytokine production INF-γ, TNF-α, IL-4 and IL-10 by the areas of residence. Association of the HLA-DRB1*01 allele with a high level of spontaneous and induced IL-10 cytokine production has been revealed at various times after booster vaccination. Further study of genes that regulate the development of immunity, along with immunological methods will make it possible to personalize the use of the existing vaccine against plague, and predict the immunogenicity and effectiveness of preventive drugs under development
Evaluation of Immunobiological Properties of Cholera Vibrio O1 and O139 Antigens
Evaluated are immunobiological properties of experimental preparation on the basis of
Vibrio cholerae O-antigen, O1 and O139 serogroups on the mouse model: determination of toxicity, analysis of morphological changes in organs, and flow-cytometric monitoring of cell cycle of leukocytes, splenocytes and bone marrow cells. It is demonstrated that O-antigen 100 µg dose per mouse does not initiate either significant changes in parenchymatous organs, or imbalance of apoptosis and proliferation of immune-competent cells. Comprehensive assessment of immunobiological properties of the preparations allows to get an insight about their effect on macroorganism, possibility for further studies and their application as chemical vaccine components
Comparative Study of Some Physical-Chemical and Immunochemical Properties of Plague Microbe Lipopolysaccharide Preparations Obtained with the Help of Different Techniques
, and degraded polysaccharide (PS) are easily soluble in water and in 0,9 % NaCl solution. They are homogenous and characterized by an adequate degree of purity. Aside from that, it is demonstrated that potentially PS is the most productive molecule fragment of LPS for the construction of plague immunodiagnostic preparation, since despite its decreased cytotoxocity PS retains identity of chemical composition and immunechemical specificity of endotoxin
Immunobiological Characteristics of Cholera Toxin B-Subunit
Carried out was immunobiological evaluation of cholera toxin B subunit preparations obtained using different experimental and production methods. It was demonstrated that B subunit preparations were non-toxic for biomodels and did not cause significant pathological alterations in their organs and tissues. They also did not alter the condition of immunocompetent cells at different stages of their cycle, and promote anti-toxic antibodies production. Thus, B subunit preparations under study can be considered as promising components of cholera vaccine
Egg retention and dispersal activity in the parasitoid wasp, Trichogramma principium
Effects of egg retention on movement and dispersal activity of Trichogramma principium (Hymenoptera, Trichogrammatidae) females were investigated under laboratory conditions. Individual females were observed during one minute in the absence of hosts. Movement activity and dispersal rate were estimated by the length of the track and by the distance from the start point, respectively. Before the test, all wasps during 2 – 4 days were presented with a possibility to parasitize a factitious laboratory host, Sitotroga cerealella Oliv. (Lepidoptera, Gelechiidae). Wasps that had parasitized before the test show significant reduction of spontaneous walking activity and dispersal rate when compared with females that refused to parasitize the non-preferred host (i.e. manifested egg retention). This effect cannot be considered as a direct arrestment reaction to the host because during the test period, no hosts were provided. Thus, egg retention results not only in temporal spread, but also in more intensive spatial dispersal of a group of simultaneously emerged females
Experimental Preventive Anti-Tularemia Preparation
Designed is an experimental preparation of a prototype chemical tularemia vaccine (PCTV). It is composed of protective antigenic complex (PAC) of tularemia microbe and S-layer protein (Slp) of plague microbe. Determined is optimum ratio of these components in the preparation and schedule of its administration. Displayed are the results of its testing as regards physical-chemical properties, reactogenicity, specific activity and impact on the immune system of laboratory animals. It is found out that preparation of the prototype is non-toxic for white mice and Guinea pigs and has no damaging effect on their immune systems. Single-stage subcutaneous immunization with PCTV induces elaboration of high-level adaptive immunity in laboratory animals within 14–21 days: specific antibody generation and stimulation of immune system cell component. PCTV protective index for white mice in case of experimental tularemia, caused by
Francisella tularensis subsp. holarctica, is 87,5 % on average; in case of infecting with
F. tularensis subsp. nearctica – 50 %; and high-level immunity in both cases. High potency of the experimental preparation against tularemia caused by subsp. holarctica (protective index is 75 %) and high-grade immunity persistence is verified on the model of Guinea pigs too