[Association of polymorphic variants of hemostatic system genes with the course of COVID-19].

INTRODUCTION:   COVID-19 is characterized by a varied clinical course. The aim of the work was to identify associations of SNPs of hemostatic system genes with COVID-19. MATERIALS AND METHODS: DNA was isolated from patients (n=117) and healthy participants (n=104). All infected patients were divided into 3 groups, depending on disease severity assessment, which was appreciated by NEWS2. Another group consisted of participants, who had asymptomatic infection in the past. Determination of SNPs of the genes FGB (-455 G/A), FII (20210 G/A), FV (1691 G/A), FVII (10976 G/A), FXIIIA1 (103 G/T), ITGA2 (807 C/T), ITGB3 (1565 T/C), SERPINE1 (-675 5G/4G) were performed by PCR using the "Genetics of Hemostasis" kit ("DNA-Technology", Russia). RESULTS: In analyzed SNPs, no significant differences were detected between the group of infected patients and healthy participants. But significant association was revealed in gene SERPINE1 (-675 5G/4G), when patient groups, differing in the disease severity, were analyzed relative to the group of participants with asymptomatic infection (p=0.0381; p=0 .0066; p=0.0009). It was found, that as COVID-19 severity scores increased, the proportion of 5G allele of gene SERPINE1 decreased, and the proportion of the 4G allele increased (p=0.005; p=0.009; p=0.0005). Similar processes were observed for genotypes 5G/5G and 4G/4G. DISCUSSION: The gene SERPINE1 (-675 5G/4G) is associated with the severity of COVID-19. CONCLUSION: For the first time, it was discovered that 5G/5G genotype of gene SERPINE1 (-675 5G/4G) can be a marker of a milder course of COVID-19, and the 4G/4G genotype as a more severe one.

[The analysis of immunoreactivity of individual B-cell epitopes of hepatitis C virus (Flaviviridae: Hepacivirus: Hepatitis С virus) NS4a antigen].

INTRODUCTION: Chronic viral hepatitis C (CHC) is a ubiquitous infectious disease, a significant limitation of which WHO attributes to the use of a new highly effective antiviral therapy. Previously, two B-cell epitopes were identified in NS4a antigen of the hepatitis C virus (HCV). It was shown that certain titers of antibodies (ABs) to the extended C-terminal epitope (1687-1718 a.a.) can predict a high probability of achieving a sustained virological response (SVR) to standard therapy with pegylated interferon-α and ribavirin.The aim of the work was to determine immunoreactivity of two B-cell epitopes (middle and C-terminal) of NS4a antigen, and to estimate a possible association of ABs to them with the achievement of SVR after standard interferon therapy and treatment with direct antiviral drugs (DAAs) daclatasvir and sofosbuvir (velpanat). MATERIALS AND METHODS: Blood serum samples of patients with CHC (n = 113), of which 55 participants received standard interferon therapy, 50 received velpanate treatment, the remaining 8 received no therapy were examined. The middle B-cell epitope (positions 24-34 a.a.) of NS4a was synthesized by the solid-phase method, while the C-terminal epitope (34-54 a.a.) was obtained using genetically engineered techniques. Enzyme immunoassay (ELISA) testing of the sera collected before treatment was performed for the two selected epitopes according to the conventional methods. RESULTS: The antibodies to the C-terminal epitope were detected significantly more frequently than those to the middle one (p = 0.01) when analyzing the blood sera of patients (n = 113). The presence of ABs to the C-terminal epitope in the serum samples of participants who completed standard interferon therapy was associated with the achievement of SVR (p = 0.0245). In the blood sera of participants who completed therapy with velpanate, an association of the presence of ABs to the C-terminal epitope with the achievement of SVR was also established (p < 0.0001). The presence of ABs to the middle B epitope was not associated with the achievement of SVR, regardless of the therapy used. DISCUSSION: The observed difference in the immunoreactivity of the two B-cell determinants may be associated with the localization of the nearest Th-epitopes, the sensitivity of NS4a antigen to proteolytic enzymes, and the peculiarities of epitope presentation by antigen-presenting cells. However, it should be noted that the immunoreactivity of the middle B-epitope is poorly studied. Although the association of ABs to the C-terminal epitope with the achievement of SVR has been shown by several scientific teams, the detailed molecular mechanism of their influence on the effectiveness of therapy is unclear. CONCLUSION: In CHC, ABs to the C-terminal epitope of NS4a are produced more frequently than those to the median epitope. The presence of ABs to the C-terminal epitope is a predictive marker of a high probability of achieving SVR, regardless of the type of therapy and antibody titer.

Analysis of Antitumor Activity of the Liposomal Photosensitizer Lipophthalocyan.

We studied specific antitumor activity of a liposomal drug based on tetra-3-phenylthiophthalocyanine aluminum hydroxide (lipophthalocyan) intended for photodynamic therapy. The optimal dose and protocol for photodynamic therapy with lipophthalocyan were chosen in experiments on mice: single intravenous dose of 6 mg/kg with a 5-h interval between administration and laser exposure and irradiation energy density of 400 J/cm2. A wide spectrum antitumor activity of lipophthalocyan was demonstrated in vivo for various transplantable mouse tumors (Lewis lung epidermoid carcinoma, S37 sarcoma, and colon adenocarcinoma AKATOL). The results show the possibility of using lipophthalocyan for photodynamic therapy of tumors of surface localization (skin and mucosa tumors).

[The mechanism of the pulmoprotective action of carboxytherapy]. / Mekhanizm pul'moprotektornogo deistviia karboksiterapii.

The problem of finding new ways for pharmacological prophylaxis and treatment of respiratory tract diseases is very relevant in public health. To eliminate hypoxia, many European and American clinics use carboxytherapy as an additional or alternative treatment. Carbon dioxide administration eliminates the sequels of oxygen starvation, by producing artificial tissue hypercapnia, which causes an increase in oxygen delivery and carbon dioxide removal from the tissues and then from the body. The main mechanism of action of carboxytherapy is associated with enhanced tissue oxygenation and is due to the Verigo-Bohr effect: the effect of CO2 concentration and pH on the binding of hemoglobin to oxygen and release of the latter from hemoglobin. By affecting the chemoreceptors of the respiratory center, carboxytherapy contributes to increased pulmonary ventilation and reduces bronchial smooth muscle tone and bronchial mucous secretion, as well as inflammation; all this improves breathing. By exerting a direct and reflex effect on the respiratory center, carboxytherapy induces hurried and deeper breathing, which increases pulmonary ventilation, speeds up perfusion and gas exchange in the lung, eliminates dyspnea and bronchospasm; therefore, it is widely used for lung diseases (asthma, pneumosclerosis, and silicosis). Carboxytherapy improves lung function in case of bronchopulmonary diseases, performance, and quality of life and it is used as an additional method in the pharmacotherapy of many diseases in pulmonology.

[Low concentrations of hepatitis C virus RNA in serologically mild infection.]

Occult HCV infection (OCI) provides significant interest recently. HCV RNA in this case can be detected not in plasma, but in blood cells and/or in liver tissue. In case of antibody genesis impairment anti-HCV detection may lead to negative or "uncertain" result. The aim of the study was to estimate infection type in blood donors and patients with hematological diseases by exploration of samples with uncertain anti-HCV detection results. Blood samples of 30 180 potential blood donors' and 4322 patients with hematological diseases were tested. Comparative analysis of wide pattern of HCV markers was performed. 33 blood donors and 42 patients were enrolled in follow-up examination. Uncertain results of Anti-HCV detection in donors' samples were in 0.18% of cases. Follow-up examination of 33 donors provided discordant results using immunochemiluminescence assay and ELISA. 15.2% donors' samples contained HCV RNA in low concentration. Follow-up observation of 42 patients with incomplete antiviral antibody pattern showed HCV RNA presence in 40.5% cases (21.4% high viremia and 19.0% low viremia). Samples with low RNA concentration contained low titers of anti-core antibodies. Samples with high titers of anti-core antibodies contained high HCV RNA level. Uncertain results of anti-HCV in 15.2% of potential blood donors' samples were confirmed by detection of HCV RNA in low concentration. It proved OCI presence in these individuals and called for testing for wide pattern of HCV markers in addition to routine screening. Patients with hematological diseases showed low level of HCV RNA along with low titers of antibodies against one or two viral antigens.

Comparative Analysis of Bioactivity of the Russian-Made Antitumor Substances of the Nitrosourea Group.

We performed an in vivo comparative study of activity of three substances of the nitrosourea group produced in Russia. All substances demonstrated high antitumor activity against various solid and leukemic tumors. Aranosa significantly enhanced life duration in mice with leukemia (by 65-194%) and inhibited the growth of solid tumors (by 49-99.6%). Lisomustine and ormustine showed higher activity than aranose. Single administration of lisomustine increased life span of mice (by 22-114%) and resulted in cure of all animals in four models: lymphoblastic leukemia L-1210, lymphocytic leukemia P-388, Lewis lung carcinoma, and cervical cancer RShM-5. After ormustine treatment, full recovery was observed only in groups with lymphocytic leukemia P-388 and cervical cancer RShM-5. These findings attest to higher activity of lisomustine in the studied models.

[Infant feeding in the rural area of Chuvash Republic and Nizhny Novgorod Region].

The nature of feeding in the first year of life affects the health of the child in all subsequent periods of life. Objective - the article is devoted to the study of the state of feeding the children of the first year of life living in rural areas. The study was carried out on the basis of three children's clinics in the Chuvash Republic and the Nizhny Novgorod region by random sampling using an anonymous questionnaire of women who had children aged 1 to 2 years living in the rural areas of the Chuvash Republic and the Nizhny Novgorod Region (140 and 60 women, respectively). According to our data, the absolute majority of women surveyed (96%) wished to feed their babies for at least 1 year. All infants (100%) after birth were attached to the chest, in the first 30 minutes - 62%, in later terms - 38%. Feeding the artificial formulas before the first application to the breast was used in nutrition of 26% of newborns. Exclusive breastfeeding for less than 4 months received 23% of babies, 4-6 months - 69%, 7 months - 8% of babies. The main reason for transferring infants to mixed and artificial feeding was hypogalactia. At the age of up to 6 months for mixed feeding 49% of babies were transferred, for artificial feeding - 16%. In most cases (69%), when signs of hypogalactia appeared no weighting was carried out, an independent transfer of the child to another type of feeding was observed in the same majority (65%) of cases. The choice of the mixture was carried out by the parents independently in 65% of cases, in other cases - on the recommendation of the pediatrician. In general, in most cases (66%), adapted mixtures were used in feeding the infants, cow milk (22%), goat milk (6%), and sour milk drinks (6%) were used from unadapted products. Before the age of 3 months 5% of babies completed breastfeeding, in the age of 4-6 months - 11%, 7-9 months - 16%, 10-12 months - 5%, more than 6 months continued to be fed with breast milk 84% of babies, more than a year - 63%. The average duration of breastfeeding was 13.8±4.2 months. Half (48%) of children received vegetable puree as the first supplement, in the remaining children porridge (24%), fruit puree (20%), cottage cheese (8%) served as such. Conclusion: breastfeeding of infants in rural areas needs active support from pediatric staff.

IMMUNOGENIC PROPERTIES OF RECOMBINANT MOZAIC PROTEINS BASED ON ANTIGENS NS4A AND NS4B OF HEPATITIS C VIRUS.

The aim of the study was to investigate immunogenic properties of mosaic recombinant proteins constructed on the data of hepatitis C virus NS4A and NS4B antigens. Four mosaic recombinant proteins, containing the T and B epitopes of the NS4A and NS4B antigens, were created by genetic engineering methods in the E. coli system. To enhance the immune response they were linked in different variations to the nucleotide sequences of murine interleukin-2 (IL-2), the Neisseria meningiditis lipopeptide, and the T helper epitope of the core protein of hepatitis C virus. The immunogenic properties of these recombinant proteins were analyzed by immunoblotting, ELISA and ELISpot using sera from immunized mice and patients infected with hepatitis C virus. Recombinant proteins specifically reacted with the sera of immunized mice and infected patients in immunoblotting. According to the ELISA data, the predominant formation of antibodies to NS4B was observed when mice were immunized with the recombinant proteins containing both antigens. Analysis of gamma-interferon production by T-lymphocytes upon contact with activated dendritic cells showed in ELISpot that the maximum production of this cytokine was detected when adjuvant components were located at the N- and C-ends of the recombinant protein. The highest level of gamma-interferon production during stimulation with this drug was detected in lymphocytes from the bone marrow and lymph nodes. The recombinant protein containing the T and B epitopes of NS4A and NS4B, murine IL-2 and the lipopeptide Neisseria meningiditis had the greatest immunostimulate effect among the four constructions. This recombinant protein formed nanoparticles of 100-120 nm in size.

[Prevalence and risk factors for nephrolithiasis among young rural residents].

AIM: To determine the prevalence of nephrolithiasis, assess the significance of known risk factors, and identify new risk factors for nephrolithiasis among young rural residents. MATERIALS AND METHODS: This cross-sectional study was conducted in 5 settlements of the Irkutsk rural district of the Irkutsk region and involved 408 inhabitants (93 men, 315 women) aged from 18 to 47 years. The selection of candidates for the study was randomized. The response of the population was 81.6%. Diagnostic evaluation included clinical examination, renal ultrasound, urinalysis and questionnaires; drinking water hardness was determined. RESULTS: The prevalence of nephrolithiasis was 10.0% (95% CI 7.1-12.9). The men-to-women ratio for the prevalence of nephrolithiasis in was 0.69. No kidney stones were found in participants younger than 20 years, but after 20 years of age, there was a statistically significant (p=0.04) trend in increasing prevalence of nephrolithiasis. Calculi were detected only in participants with acidic urine and were absent in individuals with alkaline urine (p=0.001). A higher prevalence of nephrolithiasis was found in individuals using hard water (12.5%), compared to those using water of medium hardness (1.6%) (p=0.019). The odds ratio of nephrolithiasis in persons using hard water was 8.9 (95% CI 1.2-66.6). DISCUSSION: and conclusions 1.The incidence of nephrolithiasis in the Irkutsk rural area was 10.0%, which is comparable to that in populations of Europe and North America; 2. The primary risk factors for nephrolithiasis were age, acidic urine and the hardness of drinking water, which is consistent with the data of other authors; 3. Difficulties in measuring, grading and controlling subjective risk factors for nephrolithiasis (fluid intake, diet, heredity, and past diseases) make it difficult to analyze them in cross-sectional studies and prevent direct comparison with other studies.

DIAGNOSTIC CAPACITY OF DETECTION OF SPECIFIC ANTIBODIES TO PANDEMIC INFLUENZA A(H1N1)PDM09 VIRUS.

Serologic studies occupy a significant place in influenza diagnosis. The article presents an analysis of the developed experimental version of ELISA test-systems for the detection of specific antibodies to the virus influenza A(H1N1)pdm09, and their dynamics at different stages of infection as compared with those of the traditional HAI method. The study included 20 paired samples of serum from patients hospitalized at different stages of the disease with etiology associated with the influenza virus A(H1N1)pdm09. Two groups were formed on the basis of HAI data, which showed the presence or absence of significant growth of specific antibodies to the influenza virus A(H1N1)pdm09. The control group consisted of 20 serum samples from individuals without influenza but with chronic hepatitis C. To examine the virus specific antibody two types of ELISA test systems were used. The first system was intended for the detection of IgM to the influenza virus A(H1N1)pdm09; the second was used for revealing specific IgG. The study showed the accuracy and specificity of detectable IgM and IgG to the virus influenza A(H1N1)pdm09. The dynamics of specific IgG titers in 15 of the 20 pairs of sera was reliable. The increase in titers was more pronounced than in the HAI. IgM against influenza virus could be detected up to 10 days, although reliable dynamics of these antibodies was not detected in paired samples. The test system was specific for the determination of both IgG and IgM antibodies to the influenza virus A(H1N1)pdm09 and significantly more sensitive than HAI. Using this ELISA test system, it is possible to monitor the dynamics of IgG to this virus even in the absence of diagnostic increases in antibody titers in HAI.

[Primary progressive multiple sclerosis: current issues of timely diagnosis]. / Pervichno-progressiruiushchii rasseiannyi skleroz: sovremennoe sostoianie problemy svoevremennoi postanovki diagnoza.

This article presents a review of international data on primary progressive multiple sclerosis (PPMS) and an analysis of factors influencing timely diagnosis of PPMS in a number of regions of the Russian Federation.

Stepwise injection potentiometric determination of caffeine in saliva using single-drop microextraction combined with solvent exchange.

A flow potentiometric method for determination of caffeine in saliva is suggested. This task is important for non-invasive assessment of drug metabolizing system activity in hepatocytes. In the current study, stepwise injection analysis (SWIA) was successfully combined with single-drop liquid microextraction (SDLME) and solvent exchange procedure. The method is based on the caffeine SDLME with subsequent solvent evaporation and dissolution of analyte in sulfuric acid followed by potentiometric detection using poly(vinyl chloride) membrane electrode containing potassium tetrakis[3,5-bis(trifluoromethyl)phenyl]borate as electroactive component. SDLME was employed for elimination of interfering matrix effects of saliva and caffeine metabolites such as theophylline, theobromine and paraxanthine. A linear range of 10(-5)-10(-2)M was established for caffeine with detection limit at 6 × 10(-6)M. The sample throughput was 6 samples h(-1). The proposed method was successfully applied to the determination of caffeine in saliva and the analytical results agreed well with the results obtained with reference HPLC method.

[Environmental pollution with fluoride compounds and their influence on children health].

Data of hygienic studies of air and soil during last 20 years have confirmed their correlating relationship with the general morbidity and a group of musculoskeletal diseases in children living in cities with the technogenic pollution of the environment. Their bones were established to grow unevenly and disproportionally, in 76% of children there have been violations in the development and growth of bones. The results of X-ray examination of hand bones in children and adolescents in the Irkutsk and Shelekhov cities are presented. Significant differences in morbidity patterns among children and adolescents including an increased incidence of musculoskeletal diseases by 5.6 in children and by 12 in adolescents have been revealed.

[Thyroid gland pathology in children population exposed to the combination of iodine deficiency and fluoride pollution of environment].

The article presents results of study of the impact of iodine deficiency and technogenic fluoride on the state of the thyroid gland in children. On the example of two districts of the city of Bratsk there were executed dynamic investigations (2002 and 2012), including the estimation of the pollution of ambient air and soil by fluorine compounds, levels of iodine intake by the body, the clinical examination of children aged from 5 to 7 years d and interviewing of their parents. In the course of the medical examination there were executed: physical examination by the pediatrician, endocrinologist, ultrasound examination of the thyroid gland, the determination both of serum hormone content by radioimmunoassay and urinary excretion offluorine and iodine. Concentrations of hydrogen fluoride and a solidfluorides in ambient air led to the accumulation offluoride ion in the soil. The iodine entering with drinking water and food, was established to provide only 37.5-50% of the daily requirement of iodine. Increased fluoride ion content in urine and milk teeth in children is associated with the concentrations of the fluorine-containing pollutants in the ambient air and soil. The fluoride pollution against the background of the natural iodine deficiency was established to increase the frequency of functional and morphological disorders of the thyroid gland in children.

[Frequency of detection of different hepatitis C virus subtypes in the Moscow region].

There is a high degree of the hepatitis C virus (HCV) infection in Moscow region. The analysis of different HCV subtypes is caused by the necessity of prognostic estimation of predominant subtypes for the near future because new schemes of antiviral therapy for several HCV genotypes are developed. It is established that subtype 1b (52.1%) prevails in Moscow region. The increase of subtype 3a (37.2%) against the decrease in subtype 1b is observed. In patients infected before 1990 HCV subtype 1b is predominant and liver cirrhosis is detected in 46.7% of cases. The intergenotypic recombinant 2k/1b is registered with the frequency of 0.8% and about 2% of cases of mixed HCV subtypes are detected.

[Hepatitis C virus: therapeutic targets and new drugs].

The viral and patient factors affecting the efficacy of therapy were discussed. The modern data on the molecular mechanism of action of main drugs for hepatitis C therapy were presented. New antiviral drugs (protease inhibitors) and pharmacological substances under development targeted against viral polymerase and protein NS5A were considered.

[Decline of antibodies to hepatitis C virus during antiviral therapy].

The goal of this study was to investigate interrelationship between changes in anti-hepatitis C virus antibody and response to antiviral therapy. The comparative quantitative analysis of antibodies to individual structural and nonstructural viral proteins was done during two years in three patient groups: initial responders, non-responders and a reference group (without therapy). All patients in the treated groups exhibited decrease in the analyzed antibodies to HCV proteins, but with different patterns. The first statistically significant difference in the decrease of virus-specific antibody between initial responders and non-responders was observed within the first three months of therapy beginning. Some treated patients demonstrated decrease in antibody levels to HCV proteins after the end of therapy.

[Humoral response to hepatitis C virus antigens in coinfection with human immunodeficiency virus type 1].

A humoral immune response to individual hepatitis C virus (HCV) antigens was studied in 49 patients at the subclinical stage of HIV-1 infection. These patients, as compared with a group comprising 50 patients with chronic hepatitis C, showed statistically significant higher levels of HCV-specific immunoglobulins G to nucleocapsid protein and the antigens NS3, NS4ab, NS5a. The group of patients with coinfection did not differ from those with chronic HCV monoinfection in detection rates and anti-HCV IgM.

[Antibody responses to consensus sequence of hypervariable region 1 from hepatitis C virus protein E2 in relation to the phase of infection].

Three overlapping peptides containing linear B-cell epitopes and corresponding to the consensus sequence of hypervariable region 1 (HVR1) from protein E2 were studied for their capacity to interact with the sera from patients with acute and chronic hepatitis C. Antibodies to these peptides are detectable with the comparable frequency in both acute and chronic hepatitis C viral infection, other than after viral elimination. A relationship of the outcome of acute Infection to the presence or absence of antibodies to the test peptides has not been established. The level of antibodies against protein E2 HVR1 C-terminus is significantly higher in chronic hepatitis C.

Effect of transplantation of cultured human neural stem and progenitor cells on regeneration of the cornea after chemical burn.

We studied the effect of cultured human fetal (8-12 weeks gestation) neural stem and progenitor cells on regeneration of rabbit cornea after alkaline burn. A single subconjunctival injection of cell culture suspension after burn injury significantly accelerated regeneration of the anterior and posterior epithelium in comparison with the control group.