Alla Vladimirovna Shevchenko

In pseudoexfoliative syndrome (PES), extracellular material accumulates in the eye tissues, which can lead to the clinical features of primary open-angle glaucoma (POAG), with the development of pseudoexfoliative glaucoma (PEG). The genetic predisposition to the development of PES has been shown. Aim of the study was, in the simultaneous multicenter study, to conduct a clinical and genetic analysis of toll-like receptor genes TLR2, TLR3, TLR4, TLR6  polymorphism in groups of patients with simple primary glaucoma (SPG) and pseudoexfoliative glaucoma living in different geographical regions of the Russian Federation with an assessment of the possible prognostic significance of the identified differences. Material and methods . 648 patients with POAG were examined – 199 people with PEG and 350 with SPG from six S. Fyodorov Eye Microsurgery Federal State Institution branches (99 people were not described for this trait and were not included in the analysis when dividing patients into PEG/PP subgroups): Irkutsk – 50 people, Novosibirsk – 199 people, Cheboksary, Orenburg, Saint Petersburg, Khabarovsk – 100 people each. TLR2  (rs5743708), TLR3  (rs3775291), TLR4  (rs4986790, rs4986791), TLR6  (rs5743810) were genotyped using real time PCR with the Cyber Green (Litech, Russia) on a single instrument and reagent base in the Laboratory of Clinical Immunogenetics of Research Institute of Clinical and Experimental Lymphology – Branch of the Federal Research Center Institute of Cytology and Genetics SB RAS (Novosibirsk). Results and discussion.  The deviation from the Hardy‒Weinberg distribution in the TLR4 299 was found among patients in Saint Petersburg, associated with a decrease in heterozygosity. For patients from Cheboksary, Saint Petersburg, Khabarovsk and Orenburg, in contrast to Novosibirsk, a significant (including after applying the Bonferroni correction) increase in the frequency (up to 100%) of the TLR2  753 ArgArg variant was found. For individual TLR  complexes, the values of the odds ratio indicators for predicting the development of glaucoma with PES are significant, but they are offset by the Bonferroni correction. However, this may indicate the possible involvement of these constitutional factors of innate immunity in the formation of PES. Conclusions . The results of the study outline new directions in the development of approaches to the diagnosis and treatment of glaucoma.

Previous studies have revealed a potential role of gene variants in the main histocompatibility complex, which regulates the expression of human leukocyte antigen molecules ( HLA ) in the course of coronavirus infection (COVID-19). The aim of our study was to evaluate the relationships between HLA polymorphism and COVID-19, as well as the severity of the infection process in patients from the Western Siberia. 260 patients with COVID-19 were examined two months after their convalescence. The comparison group included 372 residents from the Novosibirsk Region. Genotyping of the alleles and allele groups was performed using PCR test-systems (DNA-Technology, Moscow, Russia). The IBM SPSS Statistics 23 and Harlequine 3.5.2 were used for statistical processing. An increase in DRB1*15; DQA1*01:02 alleles and decreased frequency of DRB1*03; DQA1*01:03; DQB1*05:03 were detected in COVID-19 patients versus control group. After Bonferroni correction, the mentioned differences retained only for DQA1*01:02 . Three haplotypes were identified, which differed between the groups without Bonferroni correction. DRB1*07,15; DQA1*01:02,0201 and DQB1*02,06:02-08 frequency prevailed in patients versus control group. DRB1*07,15-DQB1*02,06:02-08; DQA1*01:02,02:01-DQB1*02,06-02-08 , and DRB1*07,15DQA1*01:02,02:01-DQB1*02,06:02-08 combinations prevailed in COVID-19 patients against control group. The DRB1*07,11; DRB1*11,15 genotypes, and several allelic combinations ( DRB1*07,11-DQA1*02:01,05:01; DRB1*11,15-DQA1*01:02,05:01; DRB1*11,15-DQB1*03:01,06:02-08, DRB1*11,15-DQA1*01:02,05:01DQB1*03:01,06:02-08 ) were present only in patients with severe disease. Prevalence of DRB1*11, DRB1*07,11 and DRB1*07,11-DQA1*02:01,05:01 was higher in the combined group (severe plus moderate course) of the disease versus the group of patients with mild disease. The results obtained make a definite contribution to clarifying the role of HLA gene polymorphism in COVID-19 not only in Western Siberia, but also in the entire Russian Federation. Further research is required, with increasing the sample size, thus reaching statistical significance, when correcting for the multiplicity of alleles.

VEGF-A is significant cytokine associated with angiogenesis. In SARS-CoV-2 infection (COVID-19), an increased level of VEGF-A was detected in serum being associated with severity and mortality of the disease. A number of polymorphic sites have been identified in regulatory regions of this gene that are associated with VEGF production. The VEGF-2758 (rs699947) is located in promoter region of this gene and the VEGF-2578 CC genotype is associated with higher production rates. The VEGF 936 (rs3025039) is located at the 3' untranslated region of VEGF gene, and its T allele is associated with a reduced plasma protein level. The aim of the study was to analyze the association of polymorphic positions of the regulatory regions of the VEGF gene (rs699947 and rs3025039) with clinical severity of the disease and cardiovascular problems in patients from the West Siberian region of Russia who previously suffered with COVID-19. The study included 260 former COVID-19 patients with varying degrees of severity. The examination took into account the previous history of cardiovascular diseases (CVD) and those with first clinical CVD signs occuring after the infection. VEGF rs699947 and VEGF rs3025039 were genotyped using TagMan probes. The significance of distribution differences in the studied genetic features was determined using a two-way version of the exact Fisher test. We did not find any differences in distribution of the genotype frequencies, both for single polymorphic positions and the complex VEGF-2578/VEGF+936 between the groups with varying degrees of the disease severity (severe, moderate, and mild), both in general group of patients, and in the subgroup of patients with CVD history. Moreover, there were no significant differences revealed between patients with newly emerged CVD after infection compared to patients without similar complications, both for single genotypes and in VEGF-2578/VEGF+936 complexes. According to our data, the functional polymorphism of the VEGF gene at these gene locuses is not associated with either COVID-19 severity, or with cardiovascular disorders accompanying the disease. Changes in VEGF levels may be due to various factors affecting it, thus requiring additional studies.

The presence of preceding cardiovascular disease (CVD) is an important risk factor for the severe clinical course of COVID-19. In addition, COVID-19 is often aggravated by cardiovascular complications. The relationship between COVID-19 and the cardiovascular system is very complex and has not been studied. When infected with SARS-CoV-2, the innate immune response is activated through the toll-like receptors (TLRs) family. And TLR role in the pathogenesis of cardiovascular diseases is important. The aim of the study was to conduct the comprehensive comparative analysis by assessing toll-like receptor TLR2 (rs5743708), TLR4 (rs4986790, rs4986791), TLR6 (rs5743810), TLR6 (rs5743810) gene polymorphism in COVID-19 convalescent patients to identify markers for disease susceptibility, severity of the course and development of cardiovascular complications. 260 patients with COVID-19 of varying severity degrees were examined. Groups with mild, moderate, and severe disease, groups with history of cardiovascular issues, and COVID-19 convalescent patients newly diagnosed with them were identified. Single nucleotide polymorphisms TLR2 (rs5743708), TLR4 (rs4986790, rs4986791), TLR6 (rs5743810) were analyzed by real-time PCR. Statistical was carried out by using SPSS 23.0 software. Gene allele and genotype rates were assessed by using a two-way Fisher criterion, and in cases of multiple comparisons, the Bonferroni correction. An increase of TLR2G and TLR2GG was revealed in COVID-19 patients. Heterozygosity in this position was significantly reduced in the group of patients. No differences in the frequencies of genotypes between groups with different disease severity was observed. TLR2-753 ArgArg:TLR4-299 AspGly:TLR4-399 ThrThr were decreased in patients with a combined moderate-severe vs. mild COVID-19. CVD patients with TLR4-299 AspAsp, TLR4-299 AspAsp:TLR4-399ThrThr were significantly more likely to suffer from severe COVID-19. The complex TLR4-299 AspGly:TLR4-399 ThrThr and TLR2-753 ArgArg:TLR4-299 AspGly:TLR4-399 ThrThr are associated with a milder disease course. Six complexes were identified, the frequency of which is significantly higher in COVID-19 convalescent patients with cardiovascular complications. These data confirm that the TLR polymorphism affects COVID-19 development and clinical diversity.

Background. Primary open-angle glaucoma (POAG) is optic neuropathy, the etiology of which is associated with genetic and non-genetic factors. IL-10 and MMP2 SNPs are associated with POAG, but the nature of the association depends on the ethnic profile of the population. For the Russian Federation, whose population includes 190 nationalities, this issue is relevant.Aim. To perform a multicenter comparative analysis of the IL-10 and MMP2 SNPs as potential factors for predicting the development of POAG in patients in four regions of the Russian Federation: the Novosibirsk, Leningrad, and Orenburg Regions, and the Chuvash Republic.Materials and мethods. We examined 499 POAG patients from 4 branches of S. Fyodorov Eye Microsurgery Federal State Institution (main group), 530 people without visual pathology (control 1), and 100 patients with cataracts (control 2). Genotyping of IL10 (rs1800896 and rs1800872 SNPs) and MMP-2 (rs243865) was performed by real-time polymerase chain reaction (RT-PCR) according to the manufacturer’s instructions (Lytex and Syntol, Russia). The differences were considered statistically significant at the Bonferroni-corrected p < 0.05.Results. An increase in the incidence of IL-10-1082 A was revealed in POAG patients compared to patients with cataract and healthy individuals. An increase in the incidence of IL-10 AA in both regions and a decrease in the frequency of MMP2-1306 TT were found. Similar patterns were established for interlocus IL-10 and MMP2 genotypes. The group of patients in the Leningrad Region differed the most compared to other regions, which may be due to their long-term residence together with the indigenous Finno – Ugric peoples.Conclusion. The data obtained should be taken into account when developing additional criteria for predicting predisposition to POAG, which is important in case of POAG in the family history.

One of the development factors of primary open-angle glaucoma (POAG) is genetic predisposition to its occurrence, associated with the peculiarity of cytokine genes. However, mechanisms for the implementation of this phenomenon for different geographic regions remain unknown. The aim of the study analyze the distribution features of the single nucleotide polymorphisms of IL10 gene (rs1800896 and rs1800872) in groups of patients with POAG from four regions of the Russian Federation: Novosibirsk (NSR), Leningrad (LEN), Orenburg (ORE) regions and the Chuvash Republic (CHVSh).Material and methods. The comparative multicenter genetic study on 499 patients with POAG from 4 branches of the S. Fyodorov Eye Microsurgery Federal State Institution (Novosibirsk – 199 people, Cheboksary – 100 people, Orenburg – 100 people, St. Petersburg – 100 people) was conducted. The control group – 530 individuals without this disease. Genotyping was performed by TaqMan (Syntol, Russia). Differences were considered statistically significant at p < 0.05, with Bonferroni correction.Results and discussion. Statistically significant multidirectional interregional differences in the distribution of IL10 gene polymorphisms in POAG patients were revealed. The frequency of IL10-592 C allele in patients from the LEN region have increased to 87.00 % relative to patients from NSO (72.86 %, odds ratio (OR) = 2.50, p_cor = 0.0006) and from ORE (75.50 %, OR = 2.17, p_cor = 0.0276), as well as IL10-592 CC genotype. LEN region patients with this genotype have 3.57-fold higher chance of POAG developing than NSO patients (p_cor = 0.0006) and 2.86-fold higher chance – relative to ORE residents (p_cor = 0.0084). In LEN region patients the frequencies of IL10-592 A, IL10-592 CA and IL10-592CA / IL10-1082AA complex genotype were reduced relative to patients from the ORE (24.00 %, OR = 6.00, p_cor = 0,0016). This complex is, on the one hand, protective against POAG disease for residents of the LEN region, and on the other hand, a risk factor for the development of POAG in residents of the NSR and the ORE.Conclusions. We have identified new factors of genetic predisposition to the POAG development in Russian Federation and identified both positive and negative associative links with this ophthalmopathology. The results can be used to develop polygenic risk indicators for the POAG development, taking into account regional ethnogeographic specifics.

Modern research shows that innate immunity plays an important role in the pathogenesis of primary open-angle glaucoma (POAG). An increase in the content of toll-like receptors (TLR) in the glaucomatous retina of the human eye was revealed. TLRs can modulate the immune response in glaucoma; provide early recognition of damaging agents, activation of signaling pathways and effector mechanisms of the nonspecific immune defense system aimed at restoring homeostasis. The TLR-encoding genes’ polymorphism alters the amino acid structure of the receptors, which leads to changes in their immune functions: expression level, ligand-binding and coreceptor functions, transport and signal transmission.   The aim was to analyze the association of the TLR2 (rs5743708), TLR3 (rs3775291), TLR4 (rs4986790, rs4986791) and TLR6 (rs5743810) polymorphisms with primary open-angle glaucoma in patients of Western Siberia.   Methods: 99 patients (52 men and 47 women) with a diagnosis of primary open-angle glaucoma were examined. The comparison group consisted of 100 people (81 women and 19 men). TLR2 (rs5743708), TLR3 (rs3775291), TLR4 (rs4986790, rs4986791) and TLR6 (rs5743810) polymorphisms were analyzed by RT-PCR using test systems with Syber Green (Lytex, Russia). Statistical analysis was performed using the software package SPSS 23.0 and Arlequin 3.5.2.2.   Results: the distribution of genotypes in the patient group and in the control group corresponded to the Hardy–Weinberg equilibrium. The genotype frequencies did not significantly differ between the two analyzed groups. The frequency of TLR2-753 ArgArg:TLR6-249 ProPro was increased in the group of patients with POAG. The linkage disequilibrium between two polymorphic positions of the TLR4 gene was revealed. In addition, the linkage disequilibrium between TLR2-TLR6 gene for the glaucoma group and the control group was revealed.   Conclusion: an increase in certain genotypes in the patient group relative to the control group may indirectly indicate the involvement of infectious factors in the initiation of POAG. However, despite the proven importance of the participation of their protein products in the pathogenesis of glaucoma, the relationship of TLR polymorphism requires additional research taking into account the ethnic characteristics of patients and intergenic interactions for a better understanding of the complex mechanisms of disease development. This will help carry out early diagnosis and develop the necessary therapeutic strategy.

Uterine leiomyomas (UL) are benign uterine tumors. Hypertrophic increase in muscle mass in LM is accompanied by development of vascular networks, which are regulated by the balance of pro-angiogenic factors, e.g., VEGF-A. Moreover, a number of inflammatory molecules exert pro-angiogenic effects, especially, IL-1β, IL-8, etc. The spectrum of their activity may overlap, being regulated by other cytokines. The aim of our work was to assess serum concentrations of cytokines actively involved in vascular network growth in the patients with multiple uterine fibroids, as compared with data obtained in conditionally healthy women. The survey included 178 females: 89 women (23-60 years old) with uterine fibroids, and 89 conditionally healthy age-matched women (22-61 years old). The levels of TNFα, IL-1β, IL-4, IL-6, IL-8, IL-10 and VEGF-A were detected by ELISA technique (Vector-Best, Russia). Statistical analysis was carried out using the IBM SPSS Statistics 23 (USA). Serum levels of IL-6, TNF and IL-8, were higher among patients compared with healthy women. The dependence of VEGF level on the number of myoma nodes has been established: VEGF serum level was higher in patients with multiple tumor nodes. In healthy women, an increase in TNFα level showed direct correlation with higher serum level of IL-6. Correlation with VEGF level was weakly negative. In leiomyoma, these relationships persist for IL-6, IL-8, VEGF levels. The obtained data are of practical importance not only as potential prognostic criteria for development of the uterine myoma at preclinical stage, but also as additional laboratory indexes for differential diagnostics, in particular when discerning uterine leiomyoma, the most common benign myomatous tumor of uterus, from malignant uterine leiomyosarcomas.

Cells of various organs and systems perform their functions and intercellular interactions not in an inert environment, but in the microenvironment of tissue fluids. Violations of the normal drainage of tissue fluids accompany lymphedema. An important mechanism of angiogenesis and vasculogenesis regulation in tissue fluids is the production and reception of vascular endothelial growth factors in combination with the regulation of matrix metalloproteinases. The aim of the work was to perform: a comparative analysis of some polymorphisms of vascular endothelial growth factor and their receptors and the genes encoding matrix metalloproteinases in two forms of lymphedema; an analysis of the relationship of these genes’ polymorphisms with the levels of vascular endothelial growth factor and matrix metalloproteinases and their inhibitors in serum and affected tissues. Polymorphism of VEGF (rs699947, rs3025039), KDR (rs10020464, rs11133360), NRP2 (rs849530, rs849563, rs16837641), matrix metalloproteinases MMP2 (rs2438650), MMP3 (rs3025058), MMP9 (rs3918242), Timp1 (rs6609533) and their combinations were analyzed by the Restriction Fragment Length Polymorphism method and TaqMan RT­PCR. The serum and tissue fluid levels were determined using the ELISA test system. Changes in the frequency distribution of MMP2 genotypes in primary and MMP3 in secondary lymphedema are shown. Significant frequency differences in NRP2 genotypes were revealed by comparing primary and secondary lymphedema. Features of the distribution of complex genotypes in primary and secondary lymphedema were revealed. The correlation analysis revealed the interdependence of the concentrations of the MMP, TIMP and VEGF products and differences in the structure of the correlation matrices of patients with both forms of lymphedema. It was shown that, in primary lymphedema, genotypes associated with low MMP2 and TIMP2 in serum and tissue fluid are detected, while in secondary lymphedema, other associations of the production levels with combined genetic traits are observed.