Репозиторий Университета

© 2019 by the authors. Licensee MDPI, Basel, Switzerland. Background: Covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is the major cause of viral persistence in patients with chronic HBV infection. Understanding the mechanisms underlying stability and persistence of HBV cccDNA in hepatocytes is critical for developing novel therapeutics and managing chronic hepatitis B. In this study, we observed an unexpected increase in HBV cccDNA levels upon suppression of transcription by de novo DNA methyltransferase DNMT3A and uncovered additional mechanisms potentially involved in HBV cccDNA maintenance. Methods: HBV-expressing cell lines were transfected with a DNMT3A-expressing plasmid. Real-time PCR and HBsAg assays were used to assess the HBV replication rate. Cell cycling was analyzed by fluorescent cell sorting. CRISPR/Cas9 was utilized to abrogate expression of APOBEC3A and APOBEC3B. Alterations in the expression of target genes were measured by real-time PCR. Results: Similar to previous studies, HBV replication induced DNMT3A expression, which in turn, led to reduced HBV transcription but elevated HBV cccDNA levels (4-to 6-fold increase). Increased levels of HBV cccDNA were not related to cell cycling, as DNMT3A accelerated proliferation of infected cells and could not contribute to HBV cccDNA expansion by arresting cells in a quiescent state. At the same time, DNMT3A suppressed transcription of innate immunity factors including cytidine deaminases APOBEC3A and APOBEC3B. CRISPR/Cas9-mediated silencing of APOBEC3A and APOBEC3B transcription had minor effects on HBV transcription, but significantly increased HBV cccDNA levels, similar to DNMT3A. In an attempt to further analyze the detrimental effects of HBV and DNMT3A on infected cells, we visualized γ-H2AX foci and demonstrated that HBV inflicts and DNMT3A aggravates DNA damage, possibly by downregulating DNA damage response factors. Additionally, suppression of HBV replication by DNMT3A may be related to reduced ATM/ATR expression. Conclusion: Formation and maintenance of HBV cccDNA pools may be partially suppressed by the baseline expression of host inhibitory factors including APOBEC3A and APOBEC3B. HBV inflicts DNA damage both directly and by inducing DNMT3A expression.

© 2017 Elsevier B.V. Background: A major role of the innate immunity in the defence of mucosal tissue is well established. However, a balance between the main components of the immunity such as toll-like receptors (TLRs) and defensins in the pathology of upper respiratory tract in children has not been addressed yet. Our aim was to investigate the gene expression of some TLRs as well as alpha and beta-defensins in children suffered from adenoid hyperthrophy in comparison with healthy children. Methods: Samples (nasal epithelium and adenoids) from patients with hypertrophic adenoids (n = 77) and control group (n = 33) were investigated. Quantification of HBD-1 and 2 mRNA, alpha-defensin-HNP1 and toll-like receptors (TLR) 2, 4 and 9 mRNA expression was performed by real-time polymerase chain reaction (PCR). The detection of TLR4 and TLR9 was performed by immunohistochemistry. Results: The main finding of the study is a dramatic up-regulation of TLR2 and TLR4 expression (but down-regulation of TLR9) along with a significant reduction in the expression of the defensins in children with adenoid hyperthrophy. Conclusion: The data suggest that one of the mechanisms of mucosal involvement in the pathogenesis of upper respiratory tract infection might by a disbalance between TLRs and defensins revealed in our study.

© 2018, SPb RAACI. The aim of this work was to study features of gene expression TLR2, TLR4, and TLR9 in the course of acute respiratory infection, depending on the time elapsing since the contamination, and dose of infection. The studies of in vivo models of acute respiratory infections caused by Gram-negative Klebsiella pneumoniae showed that, at infection dose of 104 CFU/ml, the TLR4 gene expression levels in epithelium of upper respiratory tract at 1, 3, 10 days were increased 30 times and more, complete elimination of the pathogen was observed at 3 days. At the dose of infection of 107 CFU/ml, persistence of the pathogen in upper respiratory tract was observed within a few days, accompanied by a significant increase in the level of TLR9 expression in epithelium of upper respiratory tract, and TLR4 levels in the lungs 1 day after infection, in parallel to elimination of the pathogen from the lower respiratory tract. Thus, the characteristic features of TLR4 and TLR9 gene expression in the upper respiratory tract may be considered a potential diagnostic and prognostic factors in evaluation of the course of acute respiratory infections caused by Klebsiella pneumoniae.

© 2018 Dynasty Publishing House. All rights reserved. The systematic review provides a justification for the value of innate immunity as an initial, necessary and determinant stage in the development of adaptive immunity. The participation of TLRs as a leading component of PRRs-system in maintaining natural congenital anti-infection resistance and homeostasis of the organism, in launching and dynamics of development of adaptive immunity to pathogens of infectious and non-infectious genesis was studied in detail. The importance of the influence of these pathogens on the homeostasis of the organism, on the formation of disturbances in anti-infective resistance at the organism and local levels, revealing new pathophysiological and immunological pathogenetic mechanisms of the development of these pathological processes is established. The colossal gap between fundamental studies of the biology and morphology of microorganisms and clinical studies of the diseases they cause is shortening. In an accessible form, explanations are provided for the absence of symptoms, the possibility of atypical manifestations, and the asymptomatic course of infection. There are new wide opportunities to improve and enhance the information content and personalization methods of diagnosis, treatment and prevention, as well as the creation of pharmaceuticals that act detrimental to all forms of cycle of development of pathogens, and new immunomodulatory drugs for the most effective treatment and prevention of diseases.

© 2018, SPb RAACI. The theory of polyetiological tumorigenesis is one of the most important theories of carcinogenesis. A great place in this theory is given to the role of inflammatory component, which is implemented via the factors of innate immunity. I.e., toll-like receptors (TLRs), chemokines and their receptors are related to innate immunity. Activation of TLRs may lead to regress or progression of cancer process. It is known that TLR3, TLR5, TLR7, TLR9 have the greatest anti-Tumor effect due to the dendritic cells (DCs)-mediated activation of type I T helpers, activation of M1-Type macrophages and Treg inhibition. Stimulation of TLR2 and TLR4 exerts an activating effect upon the tumor, by the MyD88 hyperactivation and secretion of IL-6 and TNFα, but exact mechanisms are not fully understood. In addition to TLRs, chemokines and their receptors have a great influence on the cancer development. It is shown that CCL2, CCL4, CCL17, CCL22 and CXCL12, which are secreted by cancer microenviroment, activate chemotaxis of tumor cells. It is also known that the chemokines activate CXCR4 and CCR7 (expressed by tumor cells) thus leading to metastasis. It is shown that there is an association between some gene polymorphisms of TLRs', chemokines and their receptors, and development of cancer. Thus, we may conclude that the role of TLRs and chemokines is important in oncogenesis. Further study of innate immunity factors influencing tumorigenesis are important for finding new approaches to cancer therapy and new potential vaccines against cancer.