Primary and Secondary micro-RNA Modulation the Extrinsic Pathway of Apoptosis in Hepatocellular Carcinoma.

Abstract-One of the most common malignant liver diseases is hepatocellular carcinoma, which has a high recurrence rate and a low five-year survival rate. It is very heterogeneous both in structure and between patients, which complicates the diagnosis, prognosis and response to treatment. In this regard, an individualized, patient-centered approach becomes important, in which the use of mimetics and hsa-miRNA inhibitors involved in the pathogenesis of the disease may be determinative. From this point of view hsa-miRNAs are of interest, their aberrant expression is associated with poor prognosis for patients and is associated with tumor progression due to dysregulation of programmed cell death (apoptosis). However, the effect of hsa-miRNA on tumor development depends not only on its direct effect on expression of genes, the primary targets, but also on secondary targets mediated by regulatory pathways. While the former are actively studied, the role of secondary targets of these hsa-miRNAs in modulating apoptosis is still unclear. The present work summarizes data on hsa-miRNAs whose primary targets are key genes of the extrinsic pathway of apoptosis. Their aberrant expression is associated with early disease relapse and poor patient outcome. For these hsa-miRNAs, using the software package ANDSystem, we reconstructed the regulation of the expression of secondary targets and analyzed their impact on the activity of the extrinsic pathway of apoptosis. The potential effect of hsa-miRNAs mediated by action on secondary targets is shown to negatively correlate with the number of primary targets. It is also shown that hsa-miR-373, hsa-miR-106b and hsa-miR-96 have the highest priority as markers of hepatocellular carcinoma, whose action on secondary targets enhances their anti-apoptotic effect.

[Primary and Secondary micro-RNA Modulation the Extrinsic Pathway of Apoptosis in Hepatocellular Carcinoma].

One of the most common malignant liver diseases is hepatocellular carcinoma, which has a high recurrence rate and a low five-year survival rate. It is very heterogeneous both in structure and between patients, which complicates the diagnosis, prognosis and response to treatment. In this regard, an individualized, patient-centered approach becomes important, in which the use of mimetics and hsa-miRNA inhibitors involved in the pathogenesis of the disease may be determinative. From this point of view hsa-miRNAs are of interest, their aberrant expression is associated with poor prognosis for patients and is associated with tumor progression due to dysregulation of programmed cell death (apoptosis). However, the effect of hsa-miRNA on tumor development depends not only on its direct effect on expression of genes, the primary targets, but also on secondary targets mediated by regulatory pathways. While the former are actively studied, the role of secondary targets of these hsa-miRNAs in modulating apoptosis is still unclear. The present work summarizes data on hsa-miRNAs whose primary targets are key genes of the extrinsic pathway of apoptosis. Their aberrant expression is associated with early disease relapse and poor patient outcome. For these hsa-miRNAs, using the software package ANDSystem, we reconstructed the regulation of the expression of secondary targets and analyzed their impact on the activity of the extrinsic pathway of apoptosis. The potential effect of hsa-miRNAs mediated by action on secondary targets is shown to negatively correlate with the number of primary targets. It is also shown that hsa-miR-373, hsa-miR-106b and hsa-miR-96 have the highest priority as markers of hepatocellular carcinoma, whose action on secondary targets enhances their anti-apoptotic effect.

Reconstruction and analysis of the gene regulatory network for cell wall function in Arabidopsis thaliana L. leaves in response to water deficit.

The plant cell wall represents the outer compartment of the plant cell, which provides a physical barrier and triggers signaling cascades under the influence of biotic and abiotic stressors. Drought is a factor that negatively affects both plant growth and development. Cell wall proteins (CWP) play an important role in the plant response to water deficit. The adaptation mechanisms of the cell wall to water loss are of interest for identifying important genetic factors determining plant drought resistance and provide valuable information on biomarkers for further selection aimed at increasing the yield of crop plants. Using ANDSystem, a gene network describing the regulation of CWPs under water restriction conditions was reconstructed. The analysis of the gene network and the transcriptome data analysis allowed prioritizing transcription factors (TF) based on their enrichment of differentially expressed genes regulated by them. As a result, scores were calculated, acting as indicators of the association of TFs with water deficit. On the basis of the score values, eight most significant TFs were selected. The highest priority was given to the TF GBF3. CWPs were prioritized according to the criterion of summing up the scores of transcription factors regulating these genes. Among the most prioritized CWPs were the AT5G03350 gene encoding a lectin-like protein, AT4G20860 encoding BBE-like 22 required for the oxidation of cellulose degradation products, and AT4G37800 encoding xyloglucan endotransglucosylase/ hydrolase 7. Overall, the implemented algorithm could be used for prediction of regulatory interactions between transcription factors and target genes encoding cell wall proteins in plants.

Prioritization of potential pharmacological targets for the development of anti-hepatocarcinoma drugs modulating the extrinsic apoptosis pathway: the reconstruction and analysis of associative gene networks help.

Hepatocellular carcinoma (HCC) is a common severe type of liver cancer characterized by an extremely aggressive course and low survival rates. It is known that disruptions in the regulation of apoptosis activation are some of the key features inherent in most cancer cells, which determines the pharmacological induction of apoptosis as an important strategy for cancer therapy. The computer design of chemical compounds capable of specifically regulating the external signaling pathway of apoptosis induction represents a promising approach for creating new effective ways of therapy for liver cancer and other oncological diseases. However, at present, most of the studies are devoted to pharmacological effects on the internal (mitochondrial) apoptosis pathway. In contrast, the external pathway induced via cell death receptors remains out of focus. Aberrant gene methylation, along with hepatitis C virus (HCV) infection, are important risk factors for the development of hepatocellular carcinoma. The reconstruction of gene networks describing the molecular mechanisms of interaction of aberrantly methylated genes with key participants of the extrinsic apoptosis pathway and their regulation by HCV proteins can provide important information when searching for pharmacological targets. In the present study, 13 criteria were proposed for prioritizing potential pharmacological targets for developing anti-hepatocarcinoma drugs modulating the extrinsic apoptosis pathway. The criteria are based on indicators of the structural and functional organization of reconstructed gene networks of hepatocarcinoma, the extrinsic apoptosis pathway, and regulatory pathways of virus-extrinsic apoptosis pathway interaction and aberrant gene methylation-extrinsic apoptosis pathway interaction using ANDSystem. The list of the top 100 gene targets ranked according to the prioritization rating was statistically significantly (p-value = 0.0002) enriched for known pharmacological targets approved by the FDA, indicating the correctness of the prioritization method. Among the promising potential pharmacological targets, six highly ranked genes (JUN, IL10, STAT3, MYC, TLR4, and KHDRBS1) are likely to deserve close attention.

Molecular-genetic pathways of hepatitis C virus regulation of the expression of cellular factors PREB and PLA2G4C, which play an important role in virus replication.

The participants of Hepatitis C virus (HCV) replication are both viral and host proteins. Therapeutic approaches based on activity inhibition of viral non-structural proteins NS3, NS5A, and NS5B are undergoing clinical trials. However, rapid mutation processes in the viral genome and acquisition of drug resistance to the existing drugs remain the main obstacles to fighting HCV. Identifying the host factors, exploring their role in HCV RNA replication, and studying viral effects on their expression is essential for understanding the mechanisms of viral replication and developing novel, effective curative approaches. It is known that the host factors PREB (prolactin regulatory element binding) and PLA2G4C (cytosolic phospholipase A2 gamma) are important for the functioning of the viral replicase complex and the formation of the platforms of HCV genome replication. The expression of PREB and PLA2G4C was significantly elevated in the presence of the HCV genome. However, the mechanisms of its regulation by HCV remain unknown. In this paper, using a text-mining technology provided by ANDSystem, we reconstructed and analyzed gene networks describing regulatory effects on the expression of PREB and PLA2G4C by HCV proteins. On the basis of the gene network analysis performed, we put forward hypotheses about the modulation of the host factors functions resulting from protein-protein interaction with HCV proteins. Among the viral proteins, NS3 showed the greatest number of regulatory linkages. We assumed that NS3 could inhibit the function of host transcription factor (TF) NOTCH1 by protein-protein interaction, leading to upregulation of PREB and PLA2G4C. Analysis of the gene networks and data on differential gene expression in HCV-infected cells allowed us to hypothesize further how HCV could regulate the expression of TFs, the binding sites of which are localized within PREB and PLA2G4C gene regions. The results obtained can be used for planning studies of the molecular-genetic mechanisms of viral-host interaction and searching for potential targets for anti-HCV therapy.

The impact of short-term confinement on human innate immunity.

During space missions cosmonauts are exposed to a myriad of distinct stressors such as radiation, overloads, weightlessness, radiation, isolation in artificial environmental conditions, which causes changes in immune system. During space flights it is very difficult to determine the particular factor associated with the observed immunological responses. This makes ground-based experiments examining the effect of each space flight associated factor along of particular value. Determining mechanisms causing alterations in cosmonauts' immunity can lead to potential targets for different countermeasures. In the current article we present the study of the early period of adaptation of human innate immunity of 6 healthy test-subjects, 4 males and 2 females aged 25 through 40, to isolation factors (hypodynamia, psychological stress, artificial environment). We measured multiple parameters characterizing innate immunity status in blood samples at chosen time points before, during and after the mission. In the experiment, highly enhanced cytokine responses were observed upon ex vivo antigen stimulations in comparison to baseline values. For cellular parameters we found multidirectional dynamics with a persistent prevalence of increasing TLRs+ monocytes as well as TLRs expression. Our study provides evidence that even a short-term confinement leads to immune changes in healthy humans that may trigger aberrant immune response.

Computer analysis of regulation of hepatocarcinoma marker genes hypermethylated by HCV proteins.

Hepatitis C virus (HCV) is a risk factor that leads to hepatocellular carcinoma (HCC) development. Epigenetic changes are known to play an important role in the molecular genetic mechanisms of virus-induced oncogenesis. Aberrant DNA methylation is a mediator of epigenetic changes that are closely associated with the HCC pathogenesis and considered a biomarker for its early diagnosis. The ANDSystem software package was used to reconstruct and evaluate the statistical significance of the pathways HCV could potentially use to regulate 32 hypermethylated genes in HCC, including both oncosuppressor and protumorigenic ones identified by genome-wide analysis of DNA methylation. The reconstructed pathways included those affecting protein-protein interactions (PPI), gene expression, protein activity, stability, and transport regulations, the expression regulation pathways being statistically significant. It has been shown that 8 out of 10 HCV proteins were involved in these pathways, the HCV NS3 protein being implicated in the largest number of regulatory pathways. NS3 was associated with the regulation of 5 tumor-suppressor genes, which may be the evidence of its central role in HCC pathogenesis. Analysis of the reconstructed pathways has demonstrated that following the transcription factor inhibition caused by binding to viral proteins, the expression of a number of oncosuppressors (WT1, MGMT, SOCS1, P53) was suppressed, while the expression of others (RASF1, RUNX3, WIF1, DAPK1) was activated. Thus, the performed gene-network reconstruction has shown that HCV proteins can influence not only the methylation status of oncosuppressor genes, but also their transcriptional regulation. The results obtained can be used in the search for pharmacological targets to develop new drugs against HCV-induced HCC.

[Non-delusional hypochondriac disorders in old age]. / Nebredovye ipokhondricheskie rasstroistva v pozdnem vozraste.

This review aims to present main concepts of hypochondria and 'hypochondriac mood' in old age. Attention is paid to historical and modern positions of old age hypochondria as a meta-syndromic pathology and as an independent mental disorder. Relationships of hypochondriac manifestations with affective variations and anxiety disorders, as well as somatic diseases, organic degenerative brain diseases and vascular disorders are discussed.

[TOLL-LIKE RECEPTORS IN COSMONAUT'S PERIPHERAL BLOOD CELLS AFTER LONG-DURATION MISSIONS TO THE INTERNATIONAL SPACE STATION].

Studies of Toll-like receptors (TLR) in 20 cosmonauts-members of long-duration (124-199-day) missions to the International space station evidenced changes in relative and absolute counts of peripheral blood monocytes with TLR2, TLR4 and TLR6 on the surface, expression of TLR2 and TLR6 genes, and genes of molecules involved in the TLR signaling pathway and TLR-related NF-KB-, JNK/p38- and IRF pathways on the day of return to Earth. The observed changes displayed individual variability.

[Status of the osteoclast-activating system in cosmonauts after long-duration missions to the International Space Station].

The results of studying the system of osteoprotegerin/ receptor activator of nuclear factor kappa-B ligand (OPG/RANKL) in 22 cosmonauts after long-duration (124 to 199 days) ISS missions are presented. Immediately on return to 1 g, changes were observed in OPG and RANKL serum levels and the ability to produce unstimulated and stimulated PGA of peripheral blood mononuclear cells in vitro. Individual variability of these changes was noticed. Our findings suggest that the cytokine OPG/RANKL-system is involved in bone remodeling in members of long-duration space missions.

[Immunological aspects of a piloted mission to Mars].

The paper deals with the results of the effects of 520-day isolation and confinement modeling some elements of a mission to Mars on the immune system. Longitudinal analyses revealed that the mechanisms of adaptive response of the human immune system to the conditions of extremely long isolation led to a change of the parameters, characterizing innate and adaptive immunity. Among them the most important are: changes in the signaling PRRs--TLR, manifested in the reduction of the percentage of circulating monocytes and granulocytes expressed on its own surfaces TLR2, TLR4 and TLR6, decreases early NK cell activation potential, increases in the percentage T- and B-lymphocytes, that expressed early activation marker CD69 after adequate stimulation, and in production of cytokines in response to PHA stimulation. The active mobilization of the mechanisms of adaptive immunity, the implementation of the function of the level of immunity to a qualitatively different level, apparently, should be taken as a sign of adaptive adjustment of an organism in response to the complex influence of unfavorable factors, aimed at the preservation of immune homeostasis.

[Congenital human immunity during 5-day dry immersion].

The system of congenital immunity was studied in 12 essentially healthy males 18 to 26 years of age subjected to 5-day dry immersion without use of countermeasures. Peripheral blood was analyzed for monocytes, granulocytes and lymphocytes expressing the TLR2+, TLR4+, TLR6+, CD11b+, CD14+, CD16+, CD18+, CD24+, CD36+, CD54+, CD56+ and CD206+ receptors. Expression of early activation marker CD69 on lymphocytes-natural killers was studied in unstimulated and interleukin-2 activated mononuclear cell cultures. The negative shifts in the congenital immunity system in some volunteers at the end of immersion and during recovery can be considered as warnings about depletion of the system reserve and increase of the risk of infectious diseases such as caused by normal microflora which typically does not provoke pathological reactions of the host.

[Analysis of the relationships between the psychophysiological status and system of adaptive immunity in the conditions of 5-day dry immersion].

Relationships of the T- and B-components of adaptive immunity and the psychophysiological status were studied in 14 volunteers for the experiment with 5-d dry immersion (DI) w/o countermeasures. Comparison of frequency of deviations in immunity parameters of psychologically different subjects demonstrated the highest frequency in non-anxious and extravert individuals on day-5 in DI. These differences in immune reactions as a function of psychological type and temperament point to existence of a neuroimmune typology and, therefore, the necessity of concurrent immunologic and psychological investigations in order to develop separate measures of rehabilitation from and prevention of stress in people with polar psychological status.

[Indicators of innate and adaptive immunity of cosmonauts after long-term space flight to international space station].

Results of innate and adaptive immunity indicators research at 12 cosmonauts who took part in long (128-215 days) expeditions to the International space station (ISS) are presented. It is shown that a space flight can lead to deflection of deviations in human immune system. These shifts occurred in decrease of phagocytes, NK, T-lymphocytes functional activity and also in abilities of immunocompetent cells to synthesize cytokines. Significant individual changes are noted in reaction of immune system to the long term space flight conditions specifying on individual predisposition to development of immune reactance infringements in the conditions of varying gravitational influences.

[Immune system parameters of healthy humans in a chamber experiment with hyperbaric oxygen-nitrogen-argon air].

The investigation into the immune effects of 9-d exposure to elevated pressures of normoxic and hypoxic O2-N-Ar atmosphere showed changes in a number of indices of genetic and adaptive immunity in normal humans. Dynamics and depth of the functional shifts under the influence of the experimental factors appear to be connected with genetically coded mechanisms of immunoreactivity. Monitoring of immune homeostasis is basic to the prevention of immune adaptation failure.

[Human immunity system status during 7-day dry immersion].

Subpopulations of lymphocytes, activation potential of T-, B- and NK-cells as well as cytokines production by immunocompetent cells in peripheral blood were studied in five volunteers for 7-day dry immersion without use of countermeasures. Results of the investigation revealed several negative shifts in the immunity and cytokines systems instigated by the experimental conditions. The considerable variability of the immune reactions to immersion suggests individual predisposition to immunological breaks in a changed gravity environment.

[The activation processes of the immune system during low intensity exercise without relaxation].

The activation processes of T-, B- and NK-cells were studied during 8-week low intensity strength training without relaxation. After the long-term training, no changes occurred in the peripheral blood contents of the main subpopulations of immunocompetent cells (CD3+, CD4+, CD8+, CD19+ and CD16+/CD56+ lymphocytes) and serum levels of immunoglobulin A, M, and G. At the same time, training was accompanied by positive activation of the immunocompetent cells which was evident from increased percentage of CD3+, CD19+ and /CD56+ cells expressing CD69 after activation (PHA, PW and II.2, respectively). The final period of the training course was also associated with a decrease in the level of lymphocyte apoptosis and increase of proliferative responses of lymphocytes.

[Immunoglobulin e and allergen-specific IgE antibodies in cosmonauts before and after long-duration missions on the International Space Station].

Immunoglobulins (IgG, IgA, IgM, IgE), specific IgE-antibodies, and interleukin-4 (IL-4) were investigated in blood serum of nine cosmonauts before and after 128- to 195-day ISS missions. It was shown that long-duration space flight does not change significantly the content of serum immunoglobulins, allergen-specific IgE-antibodies or IL-4. Analysis for probable ratios of the total IgE and IL-4 contents in the pre- and post-flight periods did not reveal any linear correlation of these parameters.