Features of Cytokine and VEGFA Gene Expression Modified with SARS-CoV-2 Virus in an In Vitro Experiment (Using the Example of the SARS-CoV-2 Vaccine Antigen).

The influence of SARS-CoV-2 antigen on the cytokine-producing function of immune cells was studied. We observed suppression of the production of proinflammatory cytokines by 11-46% relative to the spontaneous level under the influence of SARS-CoV-2 antigen vaccine simulator, as well as when it was co-administered with cortisol (IL-6 by 1.8 times and IFNγ by 1.57 times) compared with control samples. IL-8 production was reduced by 1.72 times relative to its spontaneous level. IL-8 production was reduced by 1.72 times relative to its spontaneous level. Under conditions of SARS-CoV-2 stimulation with the vaccine antigen in vitro, an increase in the relative scaled expression of the VEGFA gene by 2.16 times relative to the spontaneous level was observed, which can be regarded as a model "cytokine storm" scenario. The obtained experimental data verify the ideas about the pathogenetic mechanisms of the COVID-19 and can contribute to the development of new approaches to the correction of its complications.

Indicators of Immune and Neurohumoral Profile in Women of Fertile Age with Functional Disorders of the Autonomic Nervous System Associated with Polymorphic Variants of the HTR2A (rs7997012) and TP53 (rs1042522) Genes.

In women of fertile age with functional disorders of the autonomic nervous system (ANS), a complex of indicators of the immune and neurohumoral profile associated with polymorphic variants of the HTR2A (rs7997012) and TP53 (rs1042522) genes was revealed. In patients with the diagnosis "G90.8. Other disorders of the autonomic nervous system", the neurohumoral profile is characterized by excessive content of cortisol and serotonin (p<0.05), which indicates the development of the hypersympathicotonic variant of autonomic regulation disorders. The cellular immune profile in the examined individuals was characterized by a significant decrease in the content of CD3+CD4+ and CD3+CD8+ lymphocytes (p<0.05). At the same time hyperactivation of the humoral immune response was observed. In particular, we revealed enhanced production of IgG and IgM antibodies accompanied by increased count of CD19+ lymphocytes (p<0.05), which characterized clinical and laboratory manifestations of the asthenic syndrome. The women with ANS disorders had increased frequency of the G allele (OR=3.00; 95%CI 1.20-7.47) and GG genotype (OR=3.91; 95%CI 1.00-15.24) of the HTR2A (rs7997012) serotonin receptor gene, as well as the G allele (OR=1.93; 95%CI 1.04-3.57), CG genotype (OR=2.38; 95%CI 1.02-5.53) and the GG genotype (OR=1.48; 95%CI 0.42-5.24) of the TP53 (rs1042522) oncosuppressor protein gene (p<0.05). The polymorphic G allele and GG genotype variants of candidate genes (HTR2A (rs7997012) and TP53 (rs1042522) genes) in women with ANS pathology are associated with an imbalance of the neurohumoral (excess of cortisol and serotonin) and immune regulation (deficiency of the CD3+CD4+ and CD3+CD8+, excess of CD19+, IgG and IgM). These parameters form a complex of the immune, neurohumoral, and genetic profile indicators in women of fertile age that characterize functional disorders of ANS manifestations by hypersympathicotonic type with an asthenic component.

Comparative Evaluation of the Parameters of Sperm Apoptosis of Young and Middle-Aged Men by Flow Cytometry.

A comparative analysis of the parameters characterizing sperm apoptosis of young (27-42 years) and middle-aged (44-51 years) men was performed by flow cytometry. Irrespective of age, activity of caspase-3 and p53-mediated controlling the transmission of apoptogenic signal transmission in gametes remained stable with the formation of germ cells with delayed (p<0.05) cell death according to the Annexin V-FITC+PI- criterion (predominantly in middle-aged men). Inhibition of the transmission of a proapoptogenic stimulus mediated by membrane cell death receptors (FAS) was also observed in this group. Comparison of indicators of sperm apoptosis showed age-related features of cell death, in particular, inhibition of membrane reception triggering FAS-dependent apoptosis, which is associated with insufficient phosphatidylserine production in middle-aged men, excessive life cycle duration, and aging of spermatozoa.

The Endocrine Function of Testes in 12- and 18-Month-Old Boars of Different Breeds.

Testes have several primary functions, such as male gametes production (spermatozoa) and secretion of several endocrine factors, including the production of steroid and protein hormones which facilitate elements in the healthy reproductive function of mammals. The potential of an animal functional endocrine reservoir could be an interesting finding in animal reproduction management. Therefore, the current study aimed to analyze the functional endocrine reserves of testes in 12- and 18-month-old boars of four different boar breeds, namely Large White, Landrace, Duroc, and Tempo, as the animal model in this research (n=10). To determine the functional endocrine reserves of the testes at 12 and 18 months of age, boars received injections of human chorionic gonadotropin (hCG) 3 times every 72 hours. Testosterone was measured in blood samples (10 mL) taken from the jugular vein before the administration of hCG as well as 2, 12, 24, 48, and 72 hours after administration of each of three hCG injections. The index of endocrine activity of the testes was determined by the formula: Ita = T1 -T0 / T0 , where Ita is the index of endocrine activity of the testes, T0 is the testosterone concentration before the administration of hCG, and T1 is the maximum testosterone concentration after the third administration of hCG. The effects of the hCG injection on boar testes at different age periods indicated that the testes of Large White and Duroc boars have higher potential endocrine reserves compared with the Landrace and Tempo breeds.

Oxidized Cell-Free DNA Role in the Antioxidant Defense Mechanisms under Stress.

The present study focuses on the investigation of the oxidized cell-free DNA (cfDNA) properties in several experimental models, including cultured cerebellum cells, peripheral blood lymphocytes (PBL), plasma, and hippocampus under an acute and chronic unpredictable stress model in rats. Firstly, our study shows that Spectrum Green fluorescence-labeled oxidized cfDNA fragments were transferred into the cytoplasm of 80% of the cerebellum culture cells; meanwhile, the nonoxidized cfDNA fragments do not pass into the cells. Oxidized cfDNA stimulates the antioxidant mechanisms and induction of transcription factor NRF2 expression, followed by an activation of NRF2 signaling pathway genes-rise of Nrf2 and Hmox1 gene expression and consequently NRF2 protein synthesis. Secondly, we showed that stress increases plasma cfDNA concentration in rats corresponding with the duration of the stress exposure. At the same time, our study did not reveal any significant changes of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) level in PBL of rats under acute or chronic stress, probably due to the significantly increased Nrf2 expression, that we found in such conditions. 8-oxodG is one of the most reliable markers of DNA oxidation. We also found an increased level of 8-oxodG in the hippocampal homogenates and hippocampal dentate gyrus in rats subjected to acute and chronic stress. Taken together, our data shows that oxidized cfDNA may play a significant role in systemic and neuronal physiological mechanisms of stress and adaptation.

ROS-Induced DNA Damage Associates with Abundance of Mitochondrial DNA in White Blood Cells of the Untreated Schizophrenic Patients.

OBJECTIVE: The aim of this study was (1) to examine the leukocyte mtDNA copy number (CN) in unmedicated (SZ (m-)) and medicated (SZ (m+)) male patients with paranoid schizophrenia (SZ) in comparison with the healthy male controls (HC) and (2) to compare the leukocyte mtDNA CN with the content of an oxidation marker 8-oxodG in lymphocytes of the SZ (m-) patients. METHODS: We evaluated leukocyte mtDNA CN of 110 subjects with SZ in comparison with 60 male HC by the method qPCR (ratio mtDNA/nDNA (gene B2M) was detected). SZ patients were divided into two subgroups. The patients of the subgroups SZ (m+) (N = 55) were treated with standard antipsychotic medications in the hospital. The patients of the subgroup SZ (m-) (N = 55) were not treated before venous blood was sampled. To evaluate oxidative DNA damage, we quantified the levels of 8-oxodG in lymphocytes (flow cytometry) of SZ (m-) patients (N = 55) and HC (N = 30). RESULTS: The leukocyte mtDNA CN showed no significant difference in SZ (m+) patients and HC. The mtDNA CN in the unmedicated subgroup SZ (m-) was significantly higher than that in the SZ (m+) subgroup or in HC group. The level of 8-oxodG in the subgroup SZ (m-) was significantly higher than that in HC group. CONCLUSION: The leukocytes of the unmedicated SZ male patients with acute psychosis contain more mtDNA than the leukocytes of the male SZ patients treated with antipsychotic medications or the healthy controls. MtDNA content positively correlates with the level of 8-oxodG in the unmedicated SZ patients.

Analysis of the Role of Carriership of Polymorphic Genotypes of ESR1, eNOS, and APOE4 Genes in the Development of Arterial Hypertension in Men.

We studied the role of the carrier status for polymorphic loci of genes encoding estrogen receptors (ESR1), endothelial NO synthase (eNOS), and apolipoprotein E (APOE4) and products of their expression nitrogen oxide (NO) and apolipoprotein (ApoE) in the development of arterial hypertension in men. Conventionally healthy volunteers and 149 men with clinical manifestations of stage I-II arterial hypertension were examined. In men with arterial hypertension, the frequency of minor allele A of ESR1 gene was higher (27.5 vs. 9.5% in the reference group; χ2=4.43, p=0.04). The level of NO in the peripheral blood was also higher in the main group (χ2=3.93, p=0.047). The increase in NO concentration did not depend on the presence of polymorphic genotypes (GG and GT) of eNOS gene, but the decrease in ApoE level in blood serum was associated with TC genotype of APOE4 gene (p=0.04). Our results suggest that minor allele A of ESR1 gene is associated with the development of arterial hypertension in men. Reduced content of ApoE in blood serum of men with arterial hypertension was associated with APOE4 gene polymorphism. However, increased level of NO did not depend on polymorphic genotypes GG and GT of eNOS gene. These polymorphisms are of specific interest as additional markers of genetic predisposition to the development of arterial hypertension in middle-age men.

Diagnostics of Early Changes in the Immune System Due to Low Concentration of N-Nitrosamines in the Blood.

The concentration of N-nitrosamines (N-nitrosodimethylamine and N-nitrosodiethylamine) was measured in blood samples from children after consumption of drinking water with high content of nitrates (main group) or water meeting health standards (reference group). N-nitrosodimethylamine level in the blood from children of the main group differed from that in the reference group by 2.6 times (0.00026±0.00012 and 0.0001±0.00092 mg/dm3, respectively; p<0.05). The specific immune response to N-nitrosodimethylamine exposure was manifested in an increase in the level of specific serum IgG (2 times higher than that in the reference group). An increase in the specific sensitivity to N-nitrosodimethylamine (by the criterion of IgG) was observed in 60.7% subjects. A correlation was found between an increase in the level of IgG to N-nitrosodimethylamine and rise in the concentration of N-nitrosodimethylamine in the blood (R 2 =0.35; p=0.021). Under these conditions the spontaneous and induced production of arachidonic acid metabolites (leukotrienes) increased by 2.1 times, while the expression of p53 transcription factor (responsible for oncosuppression) decreased by 1.9 times as compared to those in the reference group (p<0.05).

Low-Dose Ionizing Radiation Affects Mesenchymal Stem Cells via Extracellular Oxidized Cell-Free DNA: A Possible Mediator of Bystander Effect and Adaptive Response.

We have hypothesized that the adaptive response to low doses of ionizing radiation (IR) is mediated by oxidized cell-free DNA (cfDNA) fragments. Here, we summarize our experimental evidence for this model. Studies involving measurements of ROS, expression of the NOX (superoxide radical production), induction of apoptosis and DNA double-strand breaks, antiapoptotic gene expression and cell cycle inhibition confirm this hypothesis. We have demonstrated that treatment of mesenchymal stem cells (MSCs) with low doses of IR (10 cGy) leads to cell death of part of cell population and release of oxidized cfDNA. cfDNA has the ability to penetrate into the cytoplasm of other cells. Oxidized cfDNA, like low doses of IR, induces oxidative stress, ROS production, ROS-induced oxidative modifications of nuclear DNA, DNA breaks, arrest of the cell cycle, activation of DNA reparation and antioxidant response, and inhibition of apoptosis. The MSCs pretreated with low dose of irradiation or oxidized cfDNA were equally effective in induction of adaptive response to challenge further dose of radiation. Our studies suggest that oxidized cfDNA is a signaling molecule in the stress signaling that mediates radiation-induced bystander effects and that it is an important component of the development of radioadaptive responses to low doses of IR.

[The significance of interleukin-6 and polymorphism of its gene (C174G) under viral, alcoholic and mixed cirrhosis of liver].

The study was carried out to evaluate significance of interleukin-6 (IL-6) and polymorphism of gene IL-6 (C174G) under liver cirrhosis of viral, alcoholic and mixed etiology. The blood of patients with liver cirrhosis of viral (n=48), alcoholic (n=11) and mixed (n=10) genesis was used for analyzing indices, level of IL-6 and polymorphism of gene IL-6 (C174G). In patients with liver cirrhosis independently of etiology increasing of concentration of IL-6 was established as compared with control group (p<0.001). The gene reliably correlated with marker of cytolysis (r=0.32; p=0.009), direct bilirubin (r=0.26; p=0.04), gamma-glutamyltranspeptidase (r=0.36; p=0.004), albumin (r=0.59; p<0.001) and degree of severity of liver cirrhosis according the Child-Pugh scale (r=0.76; r<0.001). The concentration of IL-6 was significantly higher in patients with alcoholic liver cirrhosis tahn in patients with viral and mixed genesis of disease (p=0.02 and p=0.02 correspondingly). The analysis of prevalence of genotypes and alleles of polymorphism of gene IL-6 (C174G) in groups of donors and patients with liver cirrhosis in the Permskii Kraii established no reliable differences in rate of occurrence of minor allele G and its association with production of IL-6 in examined cohorts (p=0.82 and p=0.29 correspondingly). This result indicates absence of relationship of probability of immune disorders and their genetic determination. The development of liver cirrhosis independently of etiologic factor is characterized by hyper-production of anti-inflammatory cytokine reflecting severity of lesion of hepatocytes, intensity of inflammatory process and progression of disease. The more intensive production of IL-6 was observed in case of alcoholic genesis of disease that conditioned fast progression of cirrhosis.

Quantification of cell-free DNA in blood plasma and DNA damage degree in lymphocytes to evaluate dysregulation of apoptosis in schizophrenia patients.

Oxidative DNA damage has been proposed as one of the causes of schizophrenia (SZ), and post mortem data indicate a dysregulation of apoptosis in SZ patients. To evaluate apoptosis in vivo we quantified the concentration of plasma cell-free DNA (cfDNA index, determined using fluorescence), the levels of 8-oxodG in cfDNA (immunoassay) and lymphocytes (FL1-8-oxodG index, flow cytometry) of male patients with acute psychotic disorders: paranoid SZ (total N = 58), schizophreniform (N = 11) and alcohol-induced (N = 14) psychotic disorder, and 30 healthy males. CfDNA in SZ (N = 58) does not change compared with controls. In SZ patients. Elevated levels of 8-oxodG were found in cfDNA (N = 58) and lymphocytes (n = 45). The main sources of cfDNA are dying cells with oxidized DNA. Thus, the cfDNA/FL1-8-oxodG ratio shows the level of apoptosis in damaged cells. Two subgroups were identified among the SZ patients (n = 45). For SZ-1 (31%) and SZ-2 (69%) median values of cfDNA/FL1-8-oxodG index are related as 1:6 (p < 0.0000001). For the patients with other psychotic disorders and healthy controls, cfDNA/FL1-8-oxodG values were within the range of the values in SZ-2. Thus, apoptosis is impaired in approximately one-third of SZ patients. This leads to an increase in the number of cells with damaged DNA in the patient's body tissues and may be a contributing cause of acute psychotic disorder.

[Genetically determined lipid metabolism disorders due to oral intake of technogenic hyperchlorination products].

The study covered genetically determined lipid metabolism disorders due to oral intake of technogenic hyperchlorination drinkable water products. Findings are that overweight and obese children in a main group appeared to have serum chloroform level 2.3 times higher than that in a reference group. In oral intake of hyperchlorination drinkable water products, the study revealed main genes having polymorphism associated with endocrine disorders: overweight and obesity--APOE, PPARG, HTR2A, characterizing antioxidant system state--SOD2 and detoxication--SULTA. Polymorphism of candidate genes HTR2A and SOD2 was characterized by increased occurrence of mutant homo-- and heterozygous genotype, relative risk of pathologic allele presence in population exceeded the refrence group values. Probability of increased serum serotonin and lower Cu/Zn in children with mutant homozygous genotype HTR2A and SOD2 is 1.2-1.3 times higher than in those with heterozygous and normal homozygous genotypes.

[FEATURES OF THE IMMUNE GENETIC PARAMETERS IN WORKERS IN NON-FERROUS METAL INDUSTRY].

There was performed a comparative analysis of immunogenetic indices in non-ferrous metallurgy employees under the exposure to different combinations of harmful occupational factors. The combined effect of chlorine and vanadium fumes, noise, overall vibration appeared to be associated with the gene polymorphism of cytokine regulation--VEGF and TNF (p < 0.05). In workers the combination offactors such as dust containing silicon dioxide, noise, elevated environmental thermal load was associated with cytochrome p450 gene polymorphism, allele variation ofwhich is formed owing to the homozygous genotype. At the same time there was observed an excess production of specific antibodies to vanadium and silicon, significantly differed from that of the indices in the reference group. There are proposed genetic (CYP1A1, VEGF TNFalfa) and immunological (IgG to vanadium and silicon) indices as markers of susceptibility and effect in health risk assessment of different combinations of harmful occupational factors, which will allow to increase the availability of laboratory control during surveillance activities at the objects.

[Immune and genetic changes in workers exposed to industrial noise and dust].

Comparative analysis covered immune genetic parameters in nonferrous metallurgy workers. The authors demonstrated differences in immune genetic profile between male and female workers, under exposure to combination of occupational hazards: noise and dust. Analysis of gene polymorphism revealed predominant disorders in the female workers, concerning criterion of prevalence for minor allele of genes in 1 and 2 phases of detoxication, neuro-endocrine regulation, lipid and energy metabolism, genes of immune regulation and apoptosis.

[Immune and genetic markers revealed in women working in technical rubber goods production].

Medical examination covered female workers engaged into chemical production of technical rubber goods, under combined exposure to chemicals (nitrogen oxides, carbon oxide) and noise exceeding MAC by 3.7-27.5%. Findings are increased phagocytary activity, lower level of serum IgM and IgG, suppressed expression of CD25 and CD95 T-cell receptors, and slower apoptosis induced via Fas receptor--all those indicators associated with work conditions. The disorders revealed are actualized on a basis of genetic variability features of the females, that is characterized by prevalence of mutant allels of FAS and TNFR genes with minor homozygous phenotype.

[Development of lipids and carbohydrates metabolism disorders caused by drinkable water with high content of chlorine organic compounds].

Evaluation of effects caused by environmental peroral exposure to chlorine organic compounds revealed that individuals with AG variation of HTR2A gene are a community with increased sensitivity to chloroform and a risk group for lipid and carbohydrates metabolism disorders. Individual risk of endocrine disorders (ICD: E67.8 excessive nutrition and E66.0 obesity) in these individuals is higher than in general population exposed to chloroform at residence (HQ1.72). Serum serotonin level, that is functionally connected with HTR2A gene, is 1.3 times lower vs. the reference group value.

[Characteristics of regulatory system in children exposed to the environmental chemical factors].

In children residing in areas with a high content of a number of aromatic hydrocarbons in ambient air and organochlorine compounds in drinking water there were studied the blood levels of these compounds, as well as the assessment of the indices of the immune and neuroendocrine systems was performed. The higher blood content of phenol and formaldehyde has been established and there was identified an array of organochlorine and aromatic compounds not detected in the control group children. In the blood of the children of a study group there was found an imbalance of indices of cellular components of innate and adaptive immunity, pro- and anti-inflammatory cytokines, as well as increased concentrations of free thyroxine and serotonin in the blood serum, which indicates to a change in the functions of regulatory systems in children exposed to organochlorine and aromatic compounds.

[Justifying genetic and immune markers of efficiency and sensitivity under combined exposure to risk factors in mining industry workers].

The authors evaluated and justified immunologic and genetic markers under combined exposure to risk factors in mining industry workers. Analysis covered polymorphism features of 29 genes with variant alleles possibly participating in occupationally conditioned diseases formation and serving as sensitivity markers of these diseases risk. The genes association selected demonstrates reliably changed polymorphism vs. the reference group (SOD2 superoxidedismutase gene, ANKK1 dophamine receptor gene, SULT1A1 sulphtransaminase gene, MTHFR methylene tetrahydrofolate reductase gene, VEGF endothelial growth factor gene, TNF-alpha tumor necrosis factor gene). Under combined exposure to occupational hazards (sylvinite dust, noise) in mining industry, this association can serve as adequate marking complex of sensitivity to development of occupationally conditioned diseases. Increased-production of immune cytokine regulation markers: tumor necrosis factor and vascular endothelial growth factor. Genes SOD2, ANKK1, SULT1A1, VEGF, TNFalpha are recommended as sensitivity markers, and the coded cytokines (tumor necrosis factor and endothelial growth factor) are proposed as effect markers in evaluation of health risk for workers in mining industry.

[Hygienic evaluation of immune and endocrine systems and modifications of their relationship in reproductive-age women working under exposure to chemical factors in activated carbon emissions].

The article presents results of the evaluation the changes in the relationships between immune and endocrine systems in reproductive-age women, working under exposure to chemical factors from activated carbon production. A significant increase of some chemical elements and compounds was found in blood that was associated with changes in the endocrine and immune status, as well as the presence of features in correlation parameters of these systems in reproductive-age women, working under exposure to chemical factors.

[Immunogenetic features of apoptosis in workers engaged in methanol production].

Disordered cellular death process with inhibition is seen during stress states including occupational activities associated with increased concentrations of methanol in biologic media. Workers exposed to methanol demonstrate prevalence of homozygous type of tumor necrosis factor (TNF) gene and heterozygous variant of cytochrome 450 gene, reliably increased vs reference group. Genetic changes in the main group are associated with reliable decrease in occurrence of transcription protein p53 and TNF receptor, and with altered ratio of intracellular apoptosis factors (bax and bcl-2) towards slower programmed cellular death in chemical production.