EFFECT OF SIRT1 GENE POLYMORPHISM (rs2273773) ON CARDIOVASCULAR RISK MARKERS IN OVERWEIGHT PATIENTS WITH TYPE 2 DIABETES MELLITUS OF THE EAST UKRAINIAN POPULATION

There is evidence that sirtuins as energy sensors contribute to the pathogenesis of obesity and related metabolic and cardiovascular diseases in humans. The single nucleotide polymorphism (SNP) C > T (rs2273773) of the SIRT1 gene is a so-called «silent» mutation in exon 5 and may be linked to other mutations that affect both sirtuin-1 protein levels and its activity. The aim of the study was to assess the association of polymorphic variants rs2273773 of the sirtuin-1 gene with characteristics of insulin resistance and hormonal activity of adipose tissue in overweight patients with type 2 diabetes from the East Ukrainian population. Materials and methods. The study was conducted in accordance with international and domestic ethical and legal requirements and was approved by the Medical Ethics Committee at the IEPP. All examined patients underwent inpatient treatment at the institute's clinic. A retrospective analysis of clinical and biochemical parameters obtained for patients whose samples are included in the DNA collection of patients with type 2 diabetes mellitus (DM) was conducted. 61 patients with type 2 DM (male/female: 27/34) aged 53.35 ± 1.38 years, with diabetes duration of 5.33 ± 0.67 years, glycosylated hemoglobin level of 7.74 ± 0.19%, with body mass index of 33.28 ± 0.89 kg/m2, and with waist-to-hip ratio of 0.99 ± 0.01 were selected. Antidiabetic therapy included sulfonamides, biguanides, or their combinations. Clinical, biochemical, and enzyme-linked immunosorbent assay measurements were performed at the National Institute for Public Health and the Environment (Bilthoven, the Netherlands) within the framework of scientific cooperation. Insulin resistance was characterized by the HOMA-IR index, HOMA-IR/adiponectin, adiponectin/leptin, HOMA-IR/leptin, Adipo-IR and a number of other surrogate indices. The pathological impact of dyslipidemia was assessed by Castelli risk indices I and II and triglyceride-glucose index. Determination of the single-nucleotide substitution of cytosine for thymine in exon 5 of the SIRT1 gene rs2273773 (C > T) was carried out by polymerase chain reaction with two pairs of opposite primers. Electrophoretic fractionation determined the following genotypes for the rs2273773 (C > T) SIRT1 gene: CC — 314/228 bp; CT — 314/228/135 bp; TT — 314/135 bp. Allele frequencies for the studied genotype were calculated. The normality of the distribution of variables was determined using the Kolmogorov-Smirnov criterion. To compare the indicators characterized by normal distribution, the unpaired two-tailed Student's t-test was used, to compare the parameters with non-normal distribution, the Mann-Whitney test. To statistically assess the differences observed between the empirical and theoretical frequencies of the variation series, the χ2 (chi-square) test was used. Results. Genotyping according to the above-mentioned SNP indicates that there is no accumulation of certain genotypes under the conditions of the studied pathology. A preliminary analysis of the polymorphism functional significance was performed using a recessive inheritance model, i.e. CC-carriers (n = 24) versus CT + TT (n = 37). Analysis of clinical parameters revealed that this SNP did not have a statistically significant effect on the age of manifestation of type 2 DM, the degree of diabetes compensation, HOMA-IR, atherogenicity indices, blood pressure, and degree of obesity. However, there was a trend (0.05 < P < 0.1) towards worse indicators characterizing insulin resistance of adipose tissue (HOMA-IR/adiponectin, HOMA-IR/leptin, adiponectin/leptin) in carriers of the mutant T-allele. The above was accompanied by a shift in the triglyceride-glucose index (CC-carriers: 8.15 (4.29; 14.81) units vs 10.91 (6.88; 16.39) units in the CC + CT group, 0.05 < P < 0.1) and dyslipidemia parameters (triglycerides, high-density lipoprotein cholesterol). The mutant T-allele has been found to have a negative effect on indicators related to the hormonal function of adipose tissue. Thus, there is a statistically significant decrease in the levels of the most active form of anti-inflammatory adipocytokine — high molecular weight adiponectin — in T-allele carriers relative to the indicator for the CC genotype (1.96 ± 0.19 vs 3.09 ± 0.50 mg/L, respectively, P < 0.05). This was accompanied by a decrease in the levels of vaspin, an adipokine that increases insulin sensitivity, inhibits the synthesis of pro-inflammatory cytokines, and is involved in the regulation of food intake (CC-carriers: 0.33 (0.20; 0.64) vs 0.19 (0.08; 0.49) μg/L in the CC + CT group, P < 0.05). There are trends toward higher levels of chronic inflammation markers (TNF-α, IL-6, and IL-1b), which, however, do not reach statistical significance. In addition, carriers of the T-allele had significantly higher superoxide dismutase activity (24.77 ± 0.92 vs 20.46 ± 0.68 kU/mmol Hb in CC-carriers, P < 0.01) and levels of total (44.24 ± 3.45 vs 31.39 ± 2.96 μmol/mmol Hb in CC-carriers, P < 0.01) and reduced glutathione (35.10 ± 2.96 vs 21.86 ± 2.24 μmol/mmol Hb in CC-carriers, P < 0.01) in erythrocytes. The above is most likely compensatory in nature and indicates active lipoperoxidation processes. Conclusions. Analysis of the hormonal activity of adipose tissue suggests a higher cardiovascular risk in carriers of the minor allele of the C > T rs2273773 polymorphism of the sirtuin-1 gene among overweight patients with type 2 diabetes in the East Ukrainian population. The results obtained can be used to optimize prevention and treatment regimens for overweight individuals and patients with type 2 diabetes, which will help maintain working capacity and increase the duration and quality of life.