RU2793062C1 - Method for establishing a predisposition in the formation of polar stress syndrome in students aged 7-13 years living in the polar regions - Google Patents

Abstract

FIELD: medicine.

SUBSTANCE: medical genetics and pediatrics, used to determine the predisposition to the formation of polar stress syndrome in students 7-13 years old living in the polar regions. A blood sample and a sample of buccal epithelium are taken from a student. The relative content of CD3+CD95+ lymphocytes and the level of dopamine are determined in the blood. DNA is isolated from the buccal epithelium. Genotyping polymorphism of genes TNF and ANKK1/DRD2 is carried out. With the simultaneous fulfillment of the following conditions: the presence of a combination of the GG genotype of the TNF G308A gene (rs1800629), as well as the TT genotype of the ANKK1/DRD2 gene C2137T (rs1800497), an increase in the content of relative CD3+CD95+ lymphocytes in the blood compared to the upper limit of the physiological standard, equal to 15-25%, by 1.2 or more times, and an increase in the level of dopamine compared to the upper limit of the physiological standard, equal to 5.6-44 pg/cm³, by 1.7 or more times, conclude that there is a predisposition to the formation of polar tension syndrome among students living in the polar regions.

EFFECT: reliable detection of predisposition to the formation of polar stress syndrome in schoolchildren aged 7-13 years old living in the polar regions due to the circuit of the proposed method and modes of its implementation.

1 cl, 1 tbl, 2 ex

Description

The invention relates to the field of medicine, namely to medical genetics, pediatrics, and is associated with the development of a method for predicting the danger of the formation of "polar stress syndrome" in schoolchildren 7-13 years old living in the Arctic, i.e. in particular, in the territory of the North of Siberia.

The invention can be used for making a preliminary diagnosis both in specialized clinics when examining children, and in traditional health care institutions.

Human health, including schoolchildren, in the Arctic is subject to the influence of a complex set of factors, both social and geophysical. The study of rhythm disturbances in various body functions opens up the prospect of using genetic information as a criterion for assessing the state of adaptive stress in schoolchildren.

Currently, a large contingent of people, including children, live in the Arctic for a long time in extreme climatic and geographical conditions of the Arctic zone of the Russian Federation. In connection with the foregoing, the development of effective means of monitoring the condition and health of children associated with living in such difficult conditions seems to be an important area of scientific research, as it is a necessary condition for the effective preservation of health. The invention is based on the task of creating an effective method for predicting the risk of the formation of "northern stress", i.e. "Polar voltage syndrome" among schoolchildren aged 7-13 living in the Arctic zone of the Russian Federation (in the Arctic).

The factors influencing the occurrence of "northern stress" are:

- weather conditions (a sharp change in temperature);

- oxygen starvation, causing difficulty breathing and shortness of breath;

- the presence of strong winds, which causes excitation of the central nervous system, headaches, anxiety, and sometimes a feeling of melancholy;

- a sharp change in illumination (polar day and polar night), which causes drowsiness, constant loss of strength, fatigue, significant difficulties in mastering the curriculum, reduced immunity and exacerbated diseases.

- frequent magnetic storms, which causes headaches, emotional instability, nervousness, hypersensitivity, exacerbation of chronic diseases, loss of strength.

- geochemical conditions of the area (lack of fluorine, calcium, magnesium and iodine), which leads to a violation of the metabolic processes of the body, the activity of the nervous and muscular systems.

Problems arising from the combination of the above factors can lead to "polar voltage syndrome". “Polar stress syndrome” is a specific form of chronic psycho-emotional stress, which is accompanied by sleep disturbance, shortness of breath, increased fatigue and asthenia (Kaznacheev V.P. Mechanisms of human adaptation in high latitudes, journal Medicine, 1980. 200 s. [in Russia] ).

This term was first proposed by V.P. Kaznacheev in 1974. Behind it is not just a set of external symptoms, but a certain internal mechanism for the formation of a special state of the human body. "Polar tension syndrome" reflects the mechanism of development of adaptation resistance reactions, subsequently leading to the development of diseases (https://congress.oh-events.ru/doc/arch/2019_RTStress-ShuraevaEV.pdf).

The generalized data of scientists who discovered the “polar stress syndrome” make it possible to determine its main components (Khasnulin V.I. “Modern ideas about the mechanisms of formation of northern stress in humans at high latitudes” https://cyberleninka.ru/article/n/sovremennye -predstavleniya-o-mehanizmah-formirovaniya-severnogo-stressa-u-cheloveka-v-vysokih-shirotah): - lipid hyperperoxidation (oxidative stress); - insufficiency of detoxification processes and excretory processes: - disorders of the northern type of metabolism; - northern tissue hypoxia; - immune deficiency; - polyendocrine disorders; - regenerative-plastic insufficiency; - violations of electromagnetic homeostasis; - functional dissymmetry of interhemispheric relationships; - desynchronosis; - psycho-emotional stress; - meteopathy.

From the prior art, no known solutions have been identified that allow predicting the risk of the occurrence of precisely “northern stress”. However, a number of publications reflect clinical and laboratory methods for assessing predisposition to the development of stress as such using laboratory prognostic criteria.

Clinical and laboratory methods for assessing predisposition to the development of stress are as follows:

- RF patent No. 2508545 "Method for determining the individual sensitivity of a person to oxidative stress." According to this method, in a suspension of peripheral blood neutrophils of a specific individual, the intensity of the initial chemiluminescence (IC) and chemiluminescence (NC) is recorded after they are heated in test tubes in a water bath for 30-60 seconds at a temperature of 42°C by counting the number of pulses per minute. If the value of IC / NC is less than 3, the studied are classified as stress-sensitive.

- "Method for assessing the levels of psycho-emotional stress" (K.V. Sudakov "Social and biological aspects of psycho-emotional stress: ways to protect against its undesirable consequences", Bulletin of the International Academy of Sciences, 2006, No. 1, pp. 8-14). It includes instrumental registration of the most informative physiological indicators (heart rate, respiratory rate, systolic and diastolic blood pressure, electromyogram, galvanic skin response, etc.).

- RF patent No. 2761743 "Method for assessing the chronoarchitectonics of vascular wall tension" by means of outpatient monitoring of systolic and diastolic blood pressure in a group of individuals or individually in an individual patient. The specified technical solution can be used for residents of the Far North to assess the risk of vascular accidents in the winter or summer season at the time of the day under study and their further prevention.

The disadvantages of the known methods is that these methods do not take into account such an objective criterion as a genetic predisposition to the development of northern stress and do not include the identification of candidate genes, the polymorphism of which underlies the formation of such a polar stress.

From the patent of the Russian Federation No. 2534821, a method for diagnosing oxidative stress is known by measuring the expression of oxygen-sensitive genes (options). The method involves measuring the expression of the following oxygen-sensitive genes: Pdk4, Nid2, Golph2, Actr1a, Api5, Mrp13, Scn7a, Acox3, Lamr1, Sv2b, Stmn2, Trpc3, Crebzf, Herd, Ssc1, Snrpb, Zfhx1b, sc125b, Rn. 48050, Rn. 42485, MPP3, AI 101224, Abca1, Appbp1, Rn. 96234, Nptxr, Rn. 16755, Vamp2, Ghr, Calm1, Cyp51, and ID1, and no distinct difference in the level of expression of these genes between oxidatively stressed and unexposed animals, suggest the presence of oxidative stress in the mammal. The method allows diagnosing the state of oxidative stress in the body both during and after exposure to oxidative stress.

However, this well-known method, firstly, establishes a state of stress that has already taken place, i.e. not in the early stages of its formation; and secondly, it establishes only one of the components of stress - oxidative stress, while the formation of the "polar stress syndrome" as a whole is affected by a number of factors, as mentioned above; thirdly, there is no indication in the publication that this method is applicable to schoolchildren living in the conditions of the Arctic.

At the same time, known methods for establishing a predisposition to the formation of a "polar stress syndrome" in schoolchildren aged 7-13 years old living in the conditions of the Arctic have not been identified from the prior art, so it is not possible to make a choice of the closest analogue to the claimed object.

The technical result achieved by the invention is to ensure the reliability of identifying a predisposition to the formation of "polar stress syndrome" in schoolchildren 7-13 years old living in the Arctic, with the possibility of subsequently judging the development of such conditions in children already at the early stages of its formation .

The specified technical result is achieved by the proposed method of establishing a predisposition to the formation of "polar tension syndrome" in schoolchildren 7-13 years old living in the conditions of the Arctic, which consists in taking a blood sample and a sample of the buccal epithelium from the student, determining the relative content of lymphocytes in the blood sample CD3+CD95+ and dopamine levels, DNA is isolated from a sample of buccal epithelium of deoxyribonucleic acid, then genotyping of the TNF and ANKK1/DRD2 gene polymorphisms is carried out on a detecting amplifier using a PCR polymerase chain reaction, using a DNA region as a primer by examining the genotypes of the TNF G308A gene ( rs1800629), while establishing the GG genotype for the specified gene, and the ANKK1 / DRD2 C2137T gene (rs1800497), while establishing the TT genotype for the specified gene, and while simultaneously fulfilling the following conditions: the presence of a combination of the GG genotype of the TNF G308A gene (rs1800629), and as well as the TT genotype of the ANKK1/DRD2 gene С2137Т (rs1800497), an increase in the content of relative CD3+CD95+ lymphocytes in the blood compared to the upper limit of the physiological norm equal to 15-25%, by 1.2 or more times, and an increase in the level of dopamine compared to the upper limit of the physiological norm, equal to 5.6-44 pg / cm ³ , by 1.7 or more times, they conclude that there is a predisposition to the formation of the “polar stress syndrome” in schoolchildren living in the conditions of the Arctic.

The proposed technical result is achieved due to the following.

As criteria for the implementation of the predisposition to establish the formation of the "polar tension syndrome" in schoolchildren aged 7-13 years old living in the Arctic, it is recommended to use the genotypes of candidate genes: the GG genotype of the tumor necrosis factor gene TNF G308A (rs1800629) and the TT genotype of the dopamine receptor gene ANKK1 /DRD2 C2137T (rs1800497).

The TNF gene, tumor necrosis factor, is an adipokine and a cytokine. TNF is a member of the TNF superfamily, which consists of various transmembrane proteins with a TNF homologous domain. As an adipokine, TNF contributes to insulin resistance and is associated with obesity-induced type 2 diabetes. As a cytokine, TNF is used by the immune system to signal cells. If macrophages (certain white blood cells) detect an infection, they release TNF to alert other cells of the immune system as part of the inflammatory response.

TNF signaling occurs through two receptors: TNFR1 and TNFR2. TNFR1 is constitutively expressed on most cell types, while TNFR2 is limited mainly to endothelial, epithelial, and immune cell subsets. TNFR1 signaling tends to be pro-inflammatory and apoptotic, while TNFR2 signaling is anti-inflammatory and promotes cell proliferation. Suppression of TNFR1 signaling is important for the treatment of autoimmune diseases, while TNFR2 signaling promotes wound healing.

The allelic state of the TNF gene provides for the following states: homozygous - GG or AA, or heterozygous - GA.

During the study, it was found that against the background of the presence of a typical homozygous GG polymorphism of the TNF G308A (rs1800629) gene, there are high levels of the cell death receptor - CD3+CD95+ lymphocytes encoding the synthesis of tumor necrosis factor, part of the receptor of which is CD95+ reception. The physiological meaning of this immunoregulatory scenario in the conditions of the Far North is associated with the acceleration of metabolic and immune processes of rapid death and a change in cell phenotypes, reflecting the need to compensate for energy losses and maintain an elevated level of cytokine anti-inflammatory profiles, which is accompanied by constant inflammation, immunoregulatory stress and depression.

Also, as a diagnostic criterion in the proposed method, it is proposed to use the presence of the ANKK1/DRD2 gene. The regulation of the expression of the ANKK1 gene is influenced by the DRD2 gene located next to it. One of the polymorphic loci of the ANKK1 gene is located at position 2137 in the 8th exon; cytosine (C) can be replaced by thymine (T) in it. This region is called the genetic marker C2137T, or Taq1A (rs1800497 - dbSNP number).

The DRD2 gene encodes the dopamine receptor, a protein located on the surface of neurons, coupled to G-proteins and triggering a cascade of reactions inside the cell under the influence of dopamine, the “pleasure hormone”. Violation of the dopaminergic system is associated with neurological and mental diseases.

At present, the association of rs1800497 polymorphism with deviant behavior, predisposition to alcoholism, drug addiction, tobacco smoking, the development of psychosis, schizophrenia and manic-depressive disorders, Parkinson's disease, movement disorders, migraine, etc. has been noted. This is due to the fact that in the presence of rs1800497 polyformism (T allele), there is a decrease in the density of D2 dopamine receptors in all areas of the striatum of the brain. Also, the dependence of the rs1800497 polymorphism with variants of the FTO gene ("Fat mass and obesity-associated protein") was found, while the risk of obesity and diabetes may be increased.

Dopamine is a mediator of the sympathetic-adrenal system. It is released in response to stress, including cold, to adapt and mobilize the body's energy reserves. It was found that residents of the Yamalo-Nenets Autonomous Okrug have an increased activity of the thyroid gland and a higher level of dopamine.

The variant homozygous TT genotype of the ANKK1/DRD2 gene С2137Т (rs1800497), which is predominant in the adolescent population of the Arctic, was established predominantly in the adolescent population of the Arctic, associated with low expression of the dopamine receptor, a decrease in the density of D2 receptors, which leads to an excess content of the ligand in the serum due to a discharged reception and compensatory increases the effects associated with stimulated food intake and additional energy compensation, which, however, is also a factor in the constant maintenance of high stress readiness and the formation of depression (“polar tension syndrome”).

Due to the use of buccal epithelium (biological material samples from the buccal mucosa) as the test material, simplicity and reliability of studies are ensured, as well as obtaining the necessary information in terms of isolating deoxyribonucleic acid (DNA) from the specified sample and using polymerase chain reaction (PCR) to perform genotyping of polymorphism these TNF genes and the ANKK1/DRD2 gene. In this case, it is proposed to use a DNA segment of the TNF G308A gene (rs1800629) and the ANKK1/DRD2 C2137T gene (rs1800497) as a primer, while establishing the GG genotype for the first gene, and the TT genotype for the second gene.

Moreover, from the prior art, methods for detecting a predisposition to the development of "polar stress syndrome" in schoolchildren aged 7-13 living in the conditions of the Arctic are unknown, using at the same time precisely the polymorphisms of these genes.

When assessing a high degree of predisposition to the development of the "polar tension syndrome" in schoolchildren aged 7-13 years old living in the conditions of the Arctic, such laboratory indicators as the content of CD3 + CD95 + lymphocytes were also determined in the blood sample (connection with the TNF G308A gene (rs1800629), and the level of dopamine in a blood sample of a schoolchild (relationship with the ANKK1/DRD2 gene С2137Т (rs1800497).

It is thanks to the expansion of information indicators associated with polymorphic variants of these genes that the accuracy of the potential appearance of the occurrence of the “polar tension syndrome” in a child will be ensured.

Based on the foregoing, it can be concluded that the set technical result is achieved through a set of operations of the proposed method, their sequence and modes of its implementation.

The proposed method is implemented as follows.

1. A group of schoolchildren aged 7-13 from the same ethnic population is selected. Then, a blood sample is taken from the child being examined to determine the content of relative CD3+CD95+ lymphocytes in the blood and the level of dopamine. These indicators determine:

- CD3+CD95+ lymphocytes by flow cytometry on a FACSCalibur flow cytometer (Becton Dickinson, USA), using panels of labeled monoclonal antibodies according to the Method set out in E.R. Polosukhina, S.V. Kusnetsov, T.N. Zabotina, A.D. Shirin, G.I. Kaletin, D. Yu. Blokhin, A.G. Turkina, N.V. Tsvetaeva, N.P. Logatcheva, M.R. Tenuta, Yu. V. Shishkin, Z.G. Kadagidze, N.V. Khoroshko, M.A. Volkova & A. Yu. Baryshnikov. CD95 (Fas/APO-1) Antigen Expression on the Cells of Myelodysplastic Syndromes, Acute Myelogenous Leukemia, and Chronic Myelogenous Leukemia Patients // Acute Leukemias VII Experimental Approaches and Novel Therapies Hiddemann et al. (eds) Springer-Verlag Berlin Heidelberg. - 1998. - p.96 https://link-springer-com.translate.goog/chapter/10.1007/978-3-642-71960-814?error=cookies_not_supported&error:=cookies_not_supported&code=05011daf-110d-49d4-aa6f- 5a587c69d41f&code=35f8c998-24a7-449e-8698-3b8ffle7568dc&_x_tr_sl=auto&_x_tr_tl=en&_x_tr_hl=en

- dopamine level by enzyme immunoassay according to the instructions for the Dopamine ELISA Fast Track commercial kit (Labor Diagnostika Nord GmbH, Germany).

It is established whether there is an excess of the concentration of CD3+CD95+ lymphocytes in the blood and the level of dopamine in the blood sample over their physiological norm (the physiological norm of CD3+CD95+ lymphocytes is 15-25%; the physiological norm of dopamine is 5.6 - 44 pg/cm ³ ).

2. Also, a sample of the buccal epithelium is taken from the specified child (in the form of a smear from the buccal mucosa). After taking the material, the swab (the working part of the probe with a cotton swab) is placed in a sterile Eppendorf tube with 500 μl of the transport medium (sterile 0.9% NaCl solution). The test tube with the solution and the working part of the probe is closed.

Next, DNA is isolated from the sample. For this, samples in an amount of 100 μl are lysed with 300 μl of a lysing solution, which is a 0.5% solution of sarcosyl and proteinase K (20 mg/ml) in acetate buffer (pH 7.5). Then a sorbent (kaolin) is added and the samples are washed with phosphate-buffered saline (pH 7.2) from proteins and a mixture of isopropyl alcohol: acetone from lipids by successive washing procedures. Nucleic acids remain on the sorbent. Next, the DNA adsorbed on the sorbent from the sample is extracted with a TE buffer, which is a mixture of 10 mM Tris-HCl and 1 mM EDTA (pH 8.0). The extract is subjected to centrifugation. After centrifuging the tube, the supernatant contains purified DNA.

The resulting material is ready for the polymerase chain reaction (PCR). PCR is carried out on a detecting amplifier with real-time hybridization-fluorescence detection using ready-made sets of primers and probes manufactured by Sintol CJSC, Russia, in which DNA regions of the TNF G308A gene (rs1800629) and the ANKK1/DRD2 gene were used as primers C2137T (rs1800497).

An amplification reaction is carried out, this is achieved by preparing their own reaction mixture to study the allelic state of each gene in an individual child. 0.1 µl of the prepared mixture of primers (a term accepted in genetics for end labeled nucleotides that limit (cut off) the amplifiable chain of nucleotides of the gene) and probes for the selected TNF G308A gene (rs1800629) and the ANKK1/DRD2 C2137T gene (rs1800497) are added to each tube. ) (we used Reagent kits for the determination of polymorphism TNF G308A (rs1800629 and gene ANKK1/DRD2 С2137Т (rs1800497) ZAO Syntol, Russia). The remaining components necessary for PCR are added to each tube: nucleotides (deoxynucleoside triphosphates: 10 mM dATP, dTTP , dGTP, dCTP), buffer (100 mM Tris-HCl buffer, 500 mM KCl, 40 mM MgCl ₂ ) and Tag F-polymerase. A 10 µl sample is added. Thus, the total volume of the reaction mixture is 25 µl. Each the test tube is tightly closed with a stopper and installed in the cycler.

When conducting PCR, amplification and detection are carried out on a CFX96 detection cycler from Bio-Rad.

A universal amplification program selected by the reagent manufacturer is used. It includes several stages: stage 1 - activation of TaqF polymerase ("hot start" mode) lasts 15 minutes at 95°C; stage 2 - amplification setup cycles without fluorescence measurement (5 cycles); Stage 3 - working cycles of amplification with fluorescence measurement (40 cycles).

Each amplification cycle includes DNA denaturation (5 s at 95°C), primer annealing (20 s at 60°C), and the DNA polymerization reaction itself (15 s at 72°C).

The registration of the fluorescence signal arising from the accumulation of amplification products of DNA sections is carried out in the "real time" mode after the stage of primer annealing for selected genes via the VIC channel - for the detection of one of the allelic variants of the genes, and via the FAM channel - for an alternative variant.

The results are interpreted based on the presence (or absence) of the intersection of the fluorescence curve with the threshold set at a given level, which corresponds to the presence (or absence) of the cycle threshold (N) value in the corresponding column in the results table displayed in the CFX96 cycler software.

The state of the TNF gene in the studied DNA region G308A (rs1800629) and the state of the ANKK1/DRD2 gene in the studied DNA region C2137T (rs1800497) are determined by the ratio of threshold cycles obtained from two detection channels (allelic discrimination method). There were two possible variants of the state of the gene: homozygous, in the case when one of the values of the threshold cycle is not determined (below the threshold line) and heterozygous, in the case when two values of the threshold cycles were obtained and parabolic fluorescence curves were obtained through these channels. Depending on the accumulation of which amplification product occurs in the reaction, a typical homozygous (GG), or variant homozygous (AA) state, or a heterozygous (GA) state of the TNF G308A (rs1800629) gene is established, and a typical homozygous (CC) is also established. or variant homozygous (TT) state, or heterozygous (CT) state of the ANKK1/DRD2 C2137T (rs1800497) gene.

3. And under the following conditions: with the simultaneous presence of a combination of the GG genotype of the TNF G308A gene (rs1800629), as well as the TT genotype of the ANKK1 / DRD2 gene С2137Т (rs1800497), with an increase in the content of relative CD3 + CD95 + lymphocytes in the blood compared to the upper limit of the physiological norm equal to 15-25%, 1.2 or more times, and with an increase in the level of dopamine compared with the upper limit of the physiological norm, equal to 5.6-44 pg / cm ³ , 1.7 or more times, they conclude about a high predisposition to the formation of the “polar stress syndrome” among schoolchildren living in the conditions of the Arctic.

When conducting tests on the implementation of the proposed method, two groups of children were formed, the first - the observation group - permanently residing in the Arctic (the city of Norilsk): schoolchildren aged 7-13 years, who, according to the results of the neuropsychological testing (STROOP test), found a significant decrease in indicators figurative memory, numerical memory, level of attention, the presence of irritability, tearfulness, sleep disturbances, frequent headaches, emotional instability. The comparison group also included children - schoolchildren aged 7-13 years old, living outside the conditions of the formation of the "polar tension syndrome" (Bratsk), who, according to the results of the neuropsychological testing (STROOP test), did not show a decrease in figurative memory, numerical memory, and also the level of attention, in the anamnesis there were no complaints about the presence of irritability, tearfulness, sleep disturbances, frequent headaches, emotional instability.

The observation group included 154 children. The comparison group included 103 children. The study groups were comparable in terms of age, gender, ethnic composition, comorbidity, socioeconomic level of the family, quality and composition of nutrition.

A sample of venous blood was taken from all the examined children, the study of which was carried out according to the above scheme. A sample of the buccal epithelium was also taken to determine the polymorphism of the studied genes by the PCR reaction.

The data obtained as a result of studies for children from the observation group and the comparison group are shown in Table 1.

The data shown in Table 1 shows the following. In schoolchildren - patients 1-4 (children from the observation group) of Table 1, it was found that there were complaints in the form of tearfulness, irritability, anxiety and mood swings, features of social and behavioral reactions indicating the formation of a "polar tension syndrome" in them, a negative deviation of indicators neuropsychological testing (STROOP test) from similar values of the contingent of the comparison group living outside the conditions of northern stress - a decrease in figurative memory, numerical memory, as well as the level of attention. And in these patients 1-4, there was a combination of the GG genotype of the TNF G308A gene (rs1800629), as well as the TT genotype of the ANKK1/DRD2 gene C2137T (rs1800497), a high content of relative CD3+CD95+ lymphocytes in the blood compared to the upper limit of the physiological norm of 1, 2 - 1.56 times, a high level of dopamine compared to the upper limit of the physiological norm by 1.7 - 1.84 times. Those. this confirms the reliable connection of the prognostic criteria proposed in the claimed method with the risk of the onset of the "polar stress syndrome" in the examined schoolchildren.

Schoolchildren 5-14 of Table 1 did not have manifestations of the “polar tension syndrome”. And studies have shown that the presence of these prognostic criteria is uncharacteristic for them, i.e. they did not have a simultaneous combination of the GG genotype of the TNF G308A gene (rs1800629), as well as the TT genotype of the ANKK1/DRD2 gene С2137Т (rs1800497), and also there was no increased content of lymphocytes (more than 1.2 times higher than the physiological norm) and dopamine levels ( above the limits of the physiological norm by more than 1.7 times).

Thus, high levels of the cell death receptor CD95+ have been established against the background of the presence of a typical homozygous GG polymorphism of the TNF G308A gene (rs1800629), which encodes the synthesis of tumor necrosis factor, part of the receptor of which is the reception of CD95+, the physiological meaning of this immunoregulatory scenario in the conditions of the Far North is associated with an acceleration metabolic and immune processes of rapid death and a change in cell phenotypes, reflecting the need to compensate for energy losses and maintain an elevated level of cytokine pro-inflammatory profiles, which is accompanied by constant inflammation, immunoregulatory stress and depression (“polar tension syndrome”).

At the same time, there is an increased expression of the neurotransmitter dopamine in the blood serum, which belongs to the catecholamine family, which regulates blood pressure, thermoregulation, behavioral and emotional processes, the need for which increases in the conditions of the Far North.

A variant homozygous TT genotype of the ANKK1/DRD2 C2137T (rs1800497) gene, predominant in the adolescent population of the Arctic (7-13 years), was established, associated with low expression of the dopamine receptor, a decrease in the density of D2 receptors, which leads to an excessive content of the ligand in the serum due to discharged reception and compensatory increases the effects associated with stimulated food intake and additional energy compensation, which at the same time is a factor in maintaining a constant high stress readiness and the formation of depression (“polar tension syndrome”).

To illustrate the implementation of the proposed method, two examples are given for specific patients of the same age and ethnicity.

Example 1. Patient, 10 years old, Russian. Born and permanently resides in the Arctic (Norilsk) 69 ⁰ North latitude. In the anamnesis, there were no signs indicating manifestations of the “polar tension syndrome”. However, a typical homozygous GG polymorphism of the TNF G308A gene (rs1800629), as well as a variant homozygous TT genotype of the ANKK1/DRD2 gene С2137Т (rs1800497) was found in the patient's buccal epithelium sample. In a blood sample, it was found that the content of the CD3 + CD95 + cell death receptor - an immune marker of emerging stress - was 35%, i.e. above the normal range (15-25%) more than 1.2 times. Increased expression of the neurotransmitter dopamine in the blood serum - a marker of behavioral and emotional processes and the formation of depression - 80 pg / ml, i.e. above the normal range (5.6-44 pg / ml) by more than 1.7 times. This made it possible to predict the risk of developing the “polar tension syndrome” in this child in the future. Observation of this child for 12 months showed that during this time, stress reactions gradually formed against the background of immune inflammation and accelerated cell death and excessive sympathetic-catecholamine activity of the nervous system, manifested by complaints in the form of tearfulness, irritability, anxiety and mood swings, changes the results of neuropsychological testing (STROOP test) in the form of a decrease in figurative memory, numerical memory, attention level, as well as hypersympathicotonia according to the results of variational pulsometry, which ultimately led to depression in the child in the form of "polar tension syndrome".

Analyzing the above genetic criteria and laboratory parameters that characterize this student, we can conclude that the patient has changes in the immune profile in the form of hyperreactivity of the inflammatory process according to the criterion of the activation marker CD3 + CD95 +, which characterizes the presence of excessive cell death, hypercytokinemia and asthenia, which lies in based on the immune mechanism of depressive disorders, and there is also hyperproduction of serum dopamine, which, against the background of an SNP defect in the dopamine D2 receptor, led to the accumulation of catecholamine (dopamine), disruption of behavioral and emotional processes, hypersympathicotonia with the formation of depression. This proves that, according to the proposed method, this patient was correctly predicted the risk of forming the "polar tension syndrome".

Example 2. Patient, 12 years old, Russian, student, healthy. Permanently resides in a territory that does not belong to the Arctic (Bratsk) 56 ⁰ North latitude. The presence of a variant homozygous AA polymorphism of the TNF G308Ars1800629 gene, as well as a typical homozygous CC genotype of the ANKK1/DRD2 C2137T rs 18004976 gene, was found. in the normal range (15-25%). Expression of the neurotransmitter dopamine in blood serum - a marker of behavioral and emotional processes and the formation of depression - 19 pg / ml, i.e. in the normal range (5.6-44 pg / ml). Thus, normal values of the receptor that triggers cell death were established against the background of the variant AA genotype of the TNF G308A gene (rs1800629), as well as the production of serum dopamine corresponding to the norm under conditions of the polymorphically unchanged dopamine receptor gene ANKK1/DRD2 C2137T (rs1800497) - the CC genotype, which together, it does not create conditions for the formation of stress reactions of immune genesis and excessive sympathetic-catecholamine activity of the nervous system, there are no complaints in the form of tearfulness, irritability, anxiety and mood swings, and according to the results of variational pulsometry, eutonia is observed, which ultimately does not lead to the development of depression in the form "syndrome of polar tension".

Observation of this student for 12 months showed that he did not experience "northern stress", the results of neuropsychological testing (STROOP test) showed the absence of violations of figurative memory, numerical memory, attention level. That is, the presence of the genetic program of one of the pathogenetically significant polymorphisms of the TNF G308Ars1800629 gene in the form of the AA variant genotype, as well as the presence of the typical genotype of the ANKK1/DRD2 dopamine receptor gene С2137Т rs 18004976 - СС, does not lead to the development of stress and "polar tension syndrome" in this patient.

Thus, the above data show that when implementing the proposed method using the proposed criteria, its purpose is ensured.

The claimed method allows to establish with sufficient certainty the precursors of the formation of depression in the form of "polar stress syndrome" in adolescent schoolchildren aged 7-13 years old living in the conditions of the Arctic, according to the claimed genetic criteria - a combination of two genotypes of candidate genes - the GG genotype of the TNF G308Ar gene (s1800629) and TT genotype of the ANKK1/DRD2 gene С2137Т (rs1800497), as well as an increase in the content of relative CD3+CD95+ lymphocytes in the blood compared to the upper limit of the physiological norm equal to 15-25%, by 1.2 or more times, and an increase in dopamine levels by compared with the upper limit of the physiological norm, equal to 5.6-44 pg / cm ³ , by 1.7 or more times, and plan preventive measures.

Claims (1)

A method for establishing a predisposition to the formation of a "polar tension syndrome" in schoolchildren aged 7-13 years old living in the conditions of the Arctic, characterized in that a blood sample and a sample of buccal epithelium are taken from the student, the relative content of CD3 + CD95 + lymphocytes and the level of dopamine are determined in the blood sample , extracting deoxyribonucleic acid DNA from a sample of the buccal epithelium, then, using a PCR polymerase chain reaction, genotyping the polymorphism of the TNF and ANKK1/DRD2 genes is carried out on a detecting amplifier using a DNA region as a primer by examining the genotypes of the TNF G308A (rs1800629) gene, while establishing for the specified gene, the GG genotype, and the ANKK1/DRD2 gene С2137Т (rs1800497), while establishing the TT genotype for the specified gene, and while the following conditions are met: the presence of a combination of the GG genotype of the TNF G308A gene (rs1800629), as well as the TT genotype of the ANKK1/ gene DRD2 С2137Т (rs1800497), an increase in the content of relative CD3+CD95+ lymphocytes in the blood compared to the upper limit of the physiological norm equal to 15-25% by 1.2 or more times, and an increase in the level of dopamine compared to the upper limit of the physiological norm equal to 5.6-44 pg/cm ³ , by 1.7 or more times, they conclude that there is a predisposition to the formation of the "polar stress syndrome" in schoolchildren living in the conditions of the Arctic.