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Ikramov Otabek Azizbekovich, doctoral, student of the Andijan State Medical Institute
Karimova Muyassar Khamitovna, Deputy Director for Science of the Republican Specialized Scientific and Practical Medical Center for Eye Microsurgery
Boboev Kodirjon Tukhtaboevich, Head of the Department of the Research Institute of Hematology and Blood Transfusion, Ikramov Otabek Azizbekovich, Head of the Department of the Andijan State Medical Institute
Ibragimov Zafar, Research Institute of Hematology and Blood Transfusion, Republic of Uzbekistan E-mail: [email protected]
ANALYSIS OF POLYMORPHISM ASSOCIATION RS1107946 COL1A1 GENE WITH RISK OF MYOPIA DEVELOPMENT
Abstract: The study of candidate genes confirms that short-sightedness is, in fact, so poly-physiological and complex pathology that studies of individual candidate genes are unlikely to demonstrate the fullness of the interactions and relationships necessary to take into account the significance of most susceptibility genes for the development of this disease.
Keywords: myopia, polymorphism rs1107946, gene COL1A1, clinical refraction, allele, genotype.
As you know, myopia (H52.1) is widespread throughout the world the anomaly of refraction, representing a variety of refractive errors, resulting in significant social and socio-economic problems [1,2]. Over the past decade a much-expanded understanding of the molecular biology of predetermining the error of refraction, which further confirms the assumption that shortsightedness is the result of a complex interaction between genetic predisposition and environmental exposure [4]. For many years in the world literature there are different opinions about whether the myopia predominantly genetic or caused by external factors [5, 6].
Double studies with identical (monozygotic) twins demonstrated greater consistency with the presence and severity of refractive error than in nonidentical (dysygotic) twins, which can be considered as confirmation of the importance of genetic factors in the development of myopia [7, 8]. Similar studies of myopia have confirmed that heri-tability at myopia reached 91%. Another confirmation of the importance of genetic factors in the origin and development of myopia can be observed associations between the power of refraction in parents and their children [9, 10]. Studies of patients with myopia, in which the traditional genetic analysis of the connections were carried out, revealed a number of potential loci for genes associated with myopia, but
the etiological role of the so-called "causal" genes was not proven [11, 12]. In this regard, as genes associated with the development of myopia, first of all, the genes of collagens and genes of enzymes involved in the metabolism of collagens are considered. Collagen is a protein that strengthens and supports many body tissues, including cartilage, bone, tendon, skin and sclera of the eye.
Purpose of the study. Determination of the associative relationship of polymorphism rs1107946 of the COL1A1 gene in patients with myopia of the Uzbek population.
Material and methods of investigation. The survey group consisted of 237 people residing in the territory of Uzbekistan. In the group of conditionally healthy individuals who did not have a history of myopia, 137 people were included. The main group included 100 patients. The diagnosis of myopia was established on the basis of data obtained by clinical-ophthalmological (biomicroscopy, perimetry, au-torefractometry) and instrumental research in the clinic of the Andijan State Medical Institute of the Ministry of Health of the Republic of Uzbekistan. The molecular-genetic part of this work was performed on the basis of the Department of Molecular Medicine and Cell Technologies of the Scientific Research Institute of Hematology and Blood Transfusion of the Ministry of Health of the Republic of Uzbekistan.
The material for molecular genetic analysis was DNA Australia) using the "Sintol" test system (Russia) according
samples isolated from peripheral venous blood leukocytes. DNA extraction was performed using the Ampli Prime Ribo-prep DNA extraction kit (Nextbio LLC, Russia) in accordance with the instructions of the manufacturer. Determination of the concentration of the obtained nucleic acid preparation in the samples was carried out spectrophotometrically on a NanoDrop-2000 instrument (NanoDrop Technologies, USA). The polymorphism rs1107946 of the COL1A1 gene was tested on a Rotor Gene 6000 PCR amplifier (Corbett,
to the manufacturer's instructions.
The statistical processing of the results was carried out using a computer package of statistical programs "WINPEPI 2016, Version 11.65".
Results and discussion. Molecular genetic study in the control group showed that the detection rate of the minor allele A polymorphism rs1107946 of the COL1A1 gene was 18.2%. The distribution of genotypes in the control and main groups corresponded to the Hardy-Weinberg equilibrium (PXB) (Table 2, 3).
Table 1.- The heterozygosity of the polymorphic variant rs1107946 of the COL1A1 gene in the study groups
Study groups n Level Coefficient of deviation D*
heterozygosity
Hobs Hexp
Control group 137 0.277 0.298 -0.07
Main group 100 0.49 0.416 0.17
Notes: D* is the coefficient of relative deviation, Hobs is the observed heterozygosity. Hexp - expected heterozygosity
whose level corresponds to the optimal values (D * = -0.070). Among patients with myopia, in the presence of moderate heterozygosity Hexp = 0.41 (0.4 < Hexp < 0.5), a moderate excess of heterozygotes D* = 0.17 is shown.
In the control group, relatively low gene diversity was revealed: the level of expected heterozygosity, Hexp, corresponds to 0.298, where 0.2 < Hexp < 0.3. The coefficient D* takes negative values, which indicates a lack of heterozygotes,
Table 2.- Distribution of alleles and genotypes of polymorphism rs1107946 of the gene COL1A1 in the study groups
Study groups Frequencies of alleles HWE Frequency of genotypes
С А А/А
Control group (n = 137) 0.82 0.18 The observed 0.679 0.277 0.044
Expected 0.668 0.298 0.033
x2 0.276 0.202 0.453
I p u I x2 p = 0.410; x2 = 0.678
Main group (n = 100) 0.71 0.30 The observed 0.46 0.49 0.05
Expected 0.497 0.416 0.087
x2 0.276 1.32 1.57
1 p u 1 x2 p = 0.205; x2 = 3.17
In this case, theoretically, the expected frequency of distribution of genotypes in the control group was - C/C = 0.668, C/A = 0.298, a/a = 0.033, and actually observed: C/C = =0.679; C/A = 0.277; A / A = 0.044.
The data obtained in the main group of patients are also consistent with RXB, where x2 = 0.41 and p = 0.68. The expected frequency ofgenotypes in this group was: C/C = 0.497; C/A = 0.416; A/A = 0.087, and the observed: C/C = 0.46; C/A = 0.49; AA = 0.05.
Analysis of the prevalence of genotypes revealed that in the group of patients with myopia the prevalence of the C/C genotype is 46.0% (n = 46), the C/A genotype 49.0%
(n = 49) and the A/A-5 genotype, 0% (n = 5), and in the population control group, the prevalence of the genotype is, respectively, 67.9% (n = 93), 27.7% (n = 38) and 4.4% (n = 6) (Table 3).
In the population sample of conditionally healthy individuals, the frequency of the major C allele is statistically significantly higher than in the main group and is associated with a low risk of developing the disease (x2 = 7.64; p = 0.0057; RR = 0.71; 95% CI: 0,58-0.89) (table 4).
For the T allele, the presence of a direct association with the carriage of this allele and the likelihood of the disease was shown (RR = 1.40, 95% CI: 1.13-1.74). In the main group for
the homozygous C/C genotype, a reduction in the risk of the increases by 1.7 times (x2 = 10.9, p = 0.001, RR = 1.70, 95% disease is shown: RR = 0.60 (95% CI: 0.45-0.81), x2 = 10.53;CI: 1.26-2.30), for homozygotes A / A - 1.37 times (x2 = 0.25, p = 0.0012, whereas genotypes with an unfavorable allele lead p = 0.62, RR = 1.37, 95% CI: 0.69-2.74). to a significantly increased risk: for heterozygotes of C/A it
Table 3.- Frequency distribution of alleles and genotypes of polymorphisms of the gene COL1A1 in patient groups and controls
Group n Frequencies of alleles Frequency of distribution o: : genotypes
C A C/C C/A A/A
n % n % n % n % n %
Myopia 100 141 70.5 59 29.5 46 46.0 49 49.0 5 5.0
Control group 137 224 81.8 50 18.2 93 67.9 38 27.7 6 4.4
Table 4.- Associative relationship between polymorphism rs1107946 of the gene COL1A1 and the risk of developing myopia (n = 100)
Alleles and genotypes Statistical difference
Relative risk Odds ratio X2 Yat es corrected Chi-square p-value
RR 95% CI: OR 95% CI:
C 0.71 0.58-0.89 0.53 0.35-0.82 7.64 0.0057
A 1.40 1.13-1.74 1.87 1.22-2.89
C/C 0.60 0.45-0.81 0.40 0.24-0.69 10.53 0.0012
C/A 1.70 1.26-2.30 2.61 1.50-4.53 10.91 0.001
A/A 1.37 0.69-2.74 1.68 0.49-5.81 0.25 0.62
In the main group, the chance of finding the C allele was statistically significantly lower in the group of patients: OR = 0.53; 95% CI: 0.35-0.82; x2 = 7.64; p = 0.0057, whereas the chance of finding the A allele increases more than 1.87 times (OR = 1.87, 95% CI: 1.22-2.89). The protective function of the C/C genotype has been revealed in relation to the risk of developing the disease: the frequency of this genotype is statistically significantly lower in the sample of patients (OR = =0.40, 95% CI: 0.24-0.69, x2 = 10.53; p = 0,0012), whereas the unfavorable genotypes of C/T and T/T occur 2.6 and 1.7 times more often in the patients of the main group than in the control group (OR = 2.61; 95% CI: 1.50-4, 53, and OR = 1.68, 95% CI: 0.49-5.81, respectively).
Distribution of genotypes rs1107946 in groups of people with medium degree myopia and high-grade myopia did not have statistically significant differences from those in the control group (p = 0.400 and p = 0.338).
The study of candidate genes confirms that short-sightedness is, in fact, so poly-physiological and complex pathology that studies of individual candidate genes are unlikely to demonstrate the fullness of the interactions and relationships necessary to take into account the significance of most susceptibility genes for the development of this disease.
Thus, the analysis of rs1107946 polymorphism of the COL1A1 gene allows us to draw the following conclusions:
1. The incidence of polymorphism rs1107946 of the COL1A1 gene of the minor A allele in patients with myopia in Uzbekistan was 30%.
2. A statistically significant association of the A allele and the heterozygous genotype C / A of polymorphism rs1107946 of the COL1A1 gene with the risk of myopia development was revealed.
3. The presence of the C allele and the C / S genotype of the polymorphism rs1107946 of the COL1A1 gene lowers the risk of myopia development.
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