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The efficacy of methotrexate (MT) in patients with rheumatoid arthritis (RA) may be determined by genetic factors.Objective: to evaluate the isolated and combined effects of single nucleotide polymorphisms (SNPs) of membrane transporter proteins (RFC1 80G>A and MDR1 3435C>T) and the GGH -401C>T gamma-glutamyl hydrolase enzyme genes on the efficacy of MT in patients with RA.Material and methods. The study group consisted of 85 patients with a confirmed diagnosis of RA, who received therapy with MT starting at 10 mg/week and increasing in dose to a maximum of 25 mg/week. Efficacy was assessed after six months of treatment using the dynamics of the DAS28 index, identifying patients who responded and those who did not respond to MT therapy.Genotyping of RFC1, MDR1 and GGH gene polymorphisms was performed by real-time polymerase chain reaction. Three different approaches were used to analyze the results: 1) analysis for each of the genes; 2) logistic regression; and 3) multifactor dimensionality reduction (MDR).Results and discussion. Single gene analysis was used to determine the most likely predictors of non-response to therapy: 1) for GGH-401C>T, TT genotype (odds ratio, OR 5.09; 95% confidence interval, C11.11—23.3); 2) forMDR13435C>T, the TT genotype (OR 2.38; 95% CI0.89-6.37); 3) for RFC180G>A, not - AA genotype (OR 1.87; 95% CI 0.93-3.76).The logistic regression model showed a significant effect of homozygous genotype GGH -401TT on the efficacy of MT with low sensitivity of the method. The multifactorial dimensionality reduction results show a significant synergistic effect of the MT transport genes (MDR1, RFC1) and the GGH enzyme encoding the conversion of MT to the elimination form.Conclusion. Using various statistical methods, the following results were obtained: Single gene analysis revealed the most likely predictors of nonresponse to MT therapy: GGH -401C>T - TT genotype, MDR1 3435C>T - TT genotype, RFC1 80G>A - not-AA genotype; the method of multiple logistic regression allowed to determine the significant effect of GGH -401ТТ genotype on the effect of the drug with a low sensitivity of the method; the isolated effect of polymorphisms is probably less pronounced than their combined effect on the effectiveness of MT. SNP synergism is a major contributor to the development of treatment resistance. MDR is a promising method that can be used in the future to assess the impact of SNPs.

The MIC genes are located on chromosome 6 in the class I major histocompatibility complex (MHC) region and encode a membrane-bound stress-inducible protein that acts as a ligand to stimulate the NKG2D activating receptor expressed on the surface of the most natural killer cells (NK). Currently, 7 MIC loci are known, of which only MICA and MICB encode proteins and show a significant allelic polymorphism. The MIC gene polymorphism and their location in the HLA region suggests presence of some ethnic and populational differences for the gene frequencies, linkage disequilibrium of distinct loci, and distribution of HLA-MIC haplotypes, thus making it possible to get information on genetic relationship of human populations. The aim of our study was to assess immunogenetic profile of Russian population in Chelyabinsk Region based on the non-classical HLA loci, i.e., MICA and MICB, in the context of worldwide population data. Methods of the study included immunogenetic typing of 100 donors identifying themselves as Russians, taken from the Registry of Stem Cell Donors at the Chelyabinsk Regional Blood Transfusion Station. The 2 loci (MICA and MICB) were typed at basic resolution, using PCR technique with sequence-specific primers (SSP-PCR). Gene frequencies (GF) were calculated using programs for immunogenetic research (Arlequin 3.5).Among Russian population from Chelyabinsk Region, the following characteristics of the MICA gene distributions were found: MICA *008, *002, *010, *009, frequency of > 7%; average frequencies, for MICA *004, *007, *018, *017; whereas MICA *027, *011, *006, *009:02, *049, *012, *016 was registered at a frequency of < 3.5%. MICB gene profile was as follows: MICB *005:02, *004, *002, *008 at a frequency of > 6%; at a frequency of 4% MICB *003, *005:03; MICB *005:01, * 005:04, * 009N, MICB *013, *014 at a frequency of0.5%. As based on calculated genetic distances (according to Ney) for the MICA locus, the dendrogram and scatter plot were designed by means of multidimensional scaling (MDS) method, presenting location of 30 world populations, including data on Russians in Chelyabinsk Region. The smallest genetic distances between the population of Russians from Chelyabinsk Region and other world populations were found between the population of Slovenia, as well as the USA population of European origin. As based on scatterplot obtained by the MDS approach for MICA gene frequencies, using the data of cluster analysis, we have found that the population of Russians from Chelyabinsk Region belongs to a cluster of typical European populations.The obtained patterns could be used for practical purposes to create a registry of stem cell donors in Russia. In addition, the data may be used as a control group for further research in the area of HLA-disease association, and could be also demanded by the specialists in population ethnogenesis.

Tuberculosis is a widespread infectious disease caused by M. tuberculosis, which is one of the leading causes of death in the world. According to numerous literature data, this is a genetically determined disease, and genetical polymorphism is a mechanism that leads to progression from infection to clinical manifestation. Susceptibility to infection correlates with different genes at several loci, and each individual gene plays a unique role. It is known, that the analysis of individual polymorphic variants of genes does not provide a sufficiently complete picture of the mechanisms of formation of a predisposition to multifactorial pathologies, such as tuberculosis, since their development is based on complex intergenic and gene-environmental interactions, which must be taken into account when predicting the risk of developing active forms of the disease and its severity. The concept of the functioning of cytokines as biomarkers of tuberculosis suggests that their products and interactions play an important role in the immunopathogenesis of the disease, because they form a cytokine chain with unique functions, where the removal of any link in the chain disrupts the entire mechanism of the immuno-inflammatory process. IL-6, together with TNFα and IL-1β, initiate early pro-inflammatory reactions in tuberculosis, stimulating local and systemic inflammatory reactions under participation of all common pro-inflammatory mechanisms with further transition to activation of acquired immunity. Earlier, we carried out a set of studies to evaluate the association of alleles and genotypes of these cytokine genes with a predisposition/resistance to pulmonary tuberculosis in Russians of the Chelyabinsk region. These studies have resulted into assessment of certain distribution patterns of IL-1β, TNFα, IL-6 alleles and their genotypes in pulmonary tuberculosis and its various clinical forms. The following methods were used: isolation of DNA samples from whole blood, genotyping of the studied gene polymorphisms using PCR and RFLP techniques. In this study, we analyzed the intergenic interactions of the genes for the pro-inflammatory cytokines IL-1β, TNFα, IL-6 using the method of reducing multifactor dimension in patients with pulmonary tuberculosis. The program designs optimal models of combinations for the studied genes and their interactions in tuberculosis patients. As a result of this study, a three-locus model IL-6 (-174)*С – IL-1β (+3953)*Т – IL-1β (+3953)*С was established, which was characterized by 100% reproducibility and prediction accuracy of 72%. Among the analyzed polymorphisms, the IL-6 (-174)*C polymorphism possessed the highest predictive potential with 15.27%.

Сomplex intergenic interactions should be taken into account when predicting the risk of adverse course in a pathological process. This presumption is at the heart of evolving multifactorial diseases (ulcerative colitis and irritable bowel syndrome). A single genetic polymorphism seems to be a weak risk factor when predicting the disease evolution and it could not be used as a prognostic model for development of multifactorial pathologies, especially in cases of rare alleles. However, it is well known that the combination of unfavorable alleles of several genes with an additive effect is dangerous. Therefore, identification of such polymorphisms is very important.Previously we conducted a test panel, in order to evaluate associations of alleles and genotypes of some cytokine genes (IL1β, TNFα, IL1rа, IL10, IL6) with predisposal, or resistance for ulcerative colitis and irritable bowel syndrome in the Russian Chelyabinsk region. Distribution of alleles and genotypes of these genes have been assessed in irritation bowel disease (IBD) and ulcerative colitis (UC). The following methods were used: isolation of DNA samples from whole blood, genotyping of the studied gene polymorphisms with PCR, RFLP. A comparative analysis of intergenic interactions between the cytokine genes IL1β, TNFα, IL1ra, IL10, IL6 in the IBD and UC patients was carried out by the Multifactor Dimensionality Reduction method. The analysis is based on the use of polymorphic loci of IL1β, TNFα, IL1ra, IL10, IL6 combinations chosen for analysis of intergenic interactions, with respect to the risk for IBS and UC predisposition. As a result of this study, a fourlocus model IL1β(+3953)*Т/TNFα(-308)*A/IL10(-1082)*G/IL6(-174)*G was identified for IBS, which was characterized by 90% repeatability and prediction accuracy of 74.5%. This model of gene interactions between the cytokine IL1β, TNFα, IL1ra, IL10, IL6 genes had the greatest prediction potential (p < 0.001) in IBS, whereas the model for UC was not statistically significant. The following types of genetic interactions were established: synergism between the IL1β(+3953)*Т and TNFα(-308)*A loci, whereas IL6(-174)*G и IL10(-1082)*G, TNFα(-308)*A seem to be in antagonistic relationships. The study made it possible to establish that the -174G/C IL-6 polymorphism may play a central role and provide intergenic interactions with SNPs of IL1β, TNFα, IL10 for predisposal to irritable bowel syndrome in Chelyabinsk Region of Russia.

Abstract. Common polymorphisms of interleukin-1β, receptor antagonist interleukin-1, and interleukin-4 genes have been studied among the patients with rheumatoid arthritis of Bashkir origin, living in Chelyabinsk Region. As a result of study, an increased frequency of interleukin-1β homozygosity for a high-producer allele was found. An association has been revealed between the homozygosity for 2-repeat allele of interleukin-4, and the age of disease onset.

Aim. Evaluate the fraction of various TLR1-TLR6 haplotypes in populations of Russians, Bashkir and Nagaybak of Chelyabinsk Region. Materials and methods. Potential donors of stem cells from Chelyabinsk Region Station of Blood Transfusion registry were included into the study and split into 3 populations: Russians (81), Bashkir (78) and Nagaybak (84). Genotyping by 2 polymorphisms of TLR1 and TLR6 genes was carried out in all the 3 groups. Point polymorphism of TLR1 gene 1805T>G was determined by polymorphism analysis of length of restriction fragments, and polymorphism of TLR6 gene 745C>T - by PCR using sequence-specific primers. Results. TLR1 1805*G-TLR6 745*T haplotype occurs in population of Russians (42%) and relatively rare - among Bashkir (17%). An inverse picture is observed for TLR1 1805*T-TLR6 745*C haplotype: a more frequent spread among Bashkir (65%) and relatively rare occurrence in Russians (23%). Frequencies of the mentioned haplotypes, that occupy intermediate position compared with corresponding parameters for populations ofRussians and Bashkir, were detected for Nagaybak, that, probably, reflects complex pathways of settling of their ancestors and effects of other nonadaptation factors. Conclusion. Frequencies of TLR1-TLR6 two-locus haplotypes in major populations of South Urals were determined for the first time. Further studies in this field will allow better understanding of features of immune response and sensitivity to infections in various populations.