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IV. СЛУЧАИ ИЗ ПРАКТИКИ
W 616.01:612.115
GENETIC CONTRIBUTION OF GENE POLYMORPHISM TO THROMBOTIC COMPLICATIONS IN SURGICAL PATIENTS
About the authors:
Yesenai B. Yespenbetov -cardioanesthesiologist of the first category
Key words:
folate metabolism genes, the genes of blood coagulation, venous thromboembolism, pulmonary embolism.
ABTopnap Typanbi:
EcneHôeroB EceHaé BaKûTxaHyëbi - KapMuoaHecre-3èonorMdpirep, 6ipiHrni caHarrb Miapirep.
Тушн сездер:
фолатты метаболизм гендер1, кан ую жуйесшщ гендер1, тамыр, тромбоэмболиялык аскынулар, тромбоэмболиялык екпе артериясы.
Miyerbekov E.M.2, Yespenbetov Ye.B. 1, Svyatova G.S., Berezina G.M.1
National Scientific Center of Surgery named after A.N. Syzganov, Almaty, Kazakhstan,1 Department of cardioanesthesiology and intensive care
Republican medical-genetic consultation of the Scientific Center for Obstetrics, Gynecology and Perinatology of the Ministry of Health of the Republic of Kazakhstan, Almaty, Kazakhstan 2
Abstract
Despite the advances of modern medicine in the study of the processes of hemostasis, thrombophilia is still the leading cause of death and disability in many developed countries. The frequency of venous thrombosis in the general population, according to international data, is 1-2 cases per 1000 population per year. Therefore, great interest is the study of genetic and biochemical factors involved in the change process of blood clotting.
Хирургиялык аурулардьщ тромботикалык аскынулардыц дамуына гендердщ кептурлЫпне генетикалык улес
Миербеков Е.М.2, Еспенбетов Е.Б.1, Святова Г.С., Березина Г.М.1
А.Н.Сь№анов атындаш Улттык, шлыми хирургия орталыш, Алматы к,., К,азак,стан,1 Кардиореанимация жэне реаниматология ôeëiMrneci
КР ДСжЭДМ Республикалык, акушерия, гинекология жэне перинатология Рылыми орталышныщ медико-генетикалык, консультациясы, Алматы к., Казакстан 2 Ацдатпа
Гемостаз, тромбофиль процесан зерделеуде замануи медицинанын, жeтicтiктeрiнe карамастан, квптеген дамыган елдерде влiм-жiтiм мен мугедеклктк непзп себеб'1 болып табылады. Тамыр трмбоздарынын, жилю жалпы популяцияда элемдк деректерге сэйкес, жыл сайын тургындардын, 1000-на 1-2 окиганы курайды. Сондыктан айтарлыктай кызыгушылыкты кан уюы процесамц взгерунде катысатын генетикалык пен биохимиялык факторларын зерделеу болып табылады.
Об авторах:
Еспенбетов Есенай Бакытжано-вич - врач кардиоанестезиолог, первой категории, e-mail: esen.esp@mail.ru
Ключевые слова:
гены фолатного метаболизма, гены свертывающей системы крови, венозные тромбоэмбо-лические осложнения, тромбоэмболия легочной артерии.
Генетический вклад полиморфизмов генов в развитие тромботических осложнений у хирургических больных
Миербеков Е.М.2, Еспенбетов Е.Б. 1, Святова Г.С., Березина Г.М.1
Национальный научный центр хирургии им. А.Н. Сызганова, г. Алматы, Казахстан,1 Отделение кардиоанестезиологии и реаниматологии
Республиканская медико-генетическая консультация Научного центра акушерства, гинекологии и перинатологии МЗ РК, г. Алматы, Казахстан 2
Аннотация
Несмотря на успехи современной медицины в изучении процессов гемостаза, тромбофилия по-прежнему является основной причиной смертности и инвалидизации во многих развитых странах мира. Частота венозных тромбозов в общей популяции, согласно мировым данным, составляет 1-2 случая на 1000 населения ежегодно. Поэтому большой интерес представляет изучение генетических и биохимических факторов, участвующих в изменении процессов свертывания крови.
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ВЕСТНИК ХИРУРГИИ КАЗАХСТАНА № 2(43)*2015
Introduction.
Despite the advances of modern medicine deep vein thrombosis (DVT) of the lower limbs and associated pulmonary embolism (PE) is a serious problem of modern health care. DVT and PE are a major cause of postoperative mortality (5% after general surgery and 24% after orthopedic surgery), lead to disability of patients, significantly increase the cost of treatment, require additional costs for rehabilitation and care [1-4]. In spite of the numerous scientific studies on the genetic basis of thrombophilia single answer about the genetic contribution of possible candidate genes to date does not exist that require additional study the population in the different ethnic groups with a detailed study of the role of some genetic variants that can influence the risk of thrombotic complications in surgical patients.
The aim of the study was to establish the role of genetic disorders of hemostasis in the pathogenesis of venous thromboembolic complications.
Materials and methods.
To study the clinical and diagnostic value of gene polymorphisms of folate metabolism (MTHFR, MTRR, MTR), coagulation (FV, FII, FGB, ITGB3), fibrinolytic hemostasis (PLANH-1) the molecular genetic testing of 112 patients with venous thrombosis (main group), 102 patients surgical patients without thrombotic complications (comparison group) and 100 healthy people (control group) was carried out. Given the existence of ethnic differences of the above polymorphisms study was conducted in patients of Kazakh population.
Isolation of DNA from peripheral blood of the subjects was conducted by saline followed by phenol-chloroform purification and testing in 3% agarose gel with different concentrations of phage X. Genotyping was performed by amplifying DNA in a polymerase chain reaction (PCR) using specific oligonucleotide primers was performed more restriction method to determine restriction fragment length polymorphism (RFLP). (RFLP).
Statistical analysis was carried out in the IBM SPSS Statistics program . Tables of contingency were composed. We calculated the absolute and relative indicators. To examine the relationship between gene and disease composition of respondents «Thrombosis» it was used a statistical method - %2 Pearson.
Results and discussion.
A molecular genetic study of genotype frequencies of polymorphisms C677T and A1298C
of MTHFR gene, A66G of MTRR gene, A2756G of MTR gene, 4G / 5G of PLANH-1 gene, G (-455) A of FGB gene, Leu33Pro of ITGB3 gene, G1691A of FV gene, G20210A of FII gene in the three groups (Table 1).
An important role in the occurrence of thrombosis belongs to carriage gene polymorphisms of folate metabolism, which is a complex cascade mechanism with the participation of a large number of enzymes. In folate metabolism genes it was revealed significantly higher values of carriage unfavorable genotypes CT and CC (13.39% and 21.43%) in the MTHFR gene in the main group of patients with thrombosis compared with a compared group (7.84% and 18.63) and control (5.0% and 10.0%, respectively). Also, statistically significant increases in the frequency of heterozygous carriers of C677T and A1298C of MTHFR genotypes were found in the main group (54.46% and 62.5%) compared with the comparison and control groups (from 43.0% to 56,86%) (x2 Pearson = 9,3; p-value = 0,055 and Pearson %2 = 26,8; p-value = 0,0001) (table 1).
The MTR and MTRR gene frequency of homozygous GG (8,04% and 8,93%) have significant differences (x2 Pearson = 17,9; p-value = 0,001 and Pearson %2 = 19,0; p-value = 0,001, respectively) in comparison with comparison groups (6.86% in both genes) and control (6.0% and 5.0%, respectively). Carriage of heterozygote GA of MTR and MTRR genes in the main group (56.25% and 45.54%) exceeds the rate in the comparison group (49.02% and 20.59%) and in the control group (30.0% and 29 0%, respectively).
The genes of blood coagulation (FV, FII, FGB, ITGB3) are represented by four polymorphisms. The frequency of occurrence of adverse homozygotes Pro / Pro of ITGB3 gene totaled almost 1% in all groups. Therefore, the importance of the development of thrombosis in surgical patients has heterozygous carrier Leu / Pro from 20.0% in the control group to 52.68% in the main group (x2 Pearson = 37,2; p-value = 0,0001).
Fibrinogen gene FGB G (-455) A encodes one of the chains of the protein fibrinogen, produced in the liver and is converted to insoluble fibrin - the basis of a blood clot in blood coagulation. The incidence of adverse AA homozygotes, GA heterozygotes in the main group (7.14% and 55.36%) was significantly above the rate in the control group (2% and 10.0%, respectively) (x2 Pearson = 71,4; p-value = 0,0001).
Prothrombin gene (Factor II) G20210A encodes a protein (prothrombin), which is one of the main factors of the coagulation system. Heterozygous genotype of AG showed a significantly higher rate (16.07%) as compared to the primary comparison group (4.90%) and the control (3.00%, respectively) (x2 Pearson = 14,2; p-value = 0.001) (Table 1).
The role of the factor 5 gene (Leiden mutation) G1691A is big in the development of thrombosis in surgical patients. Heterozygous genotype of AG showed a significantly higher rate (18.75%) as compared to the primary comparison group (6.86%) and control (1,00%) (x2 Pearson = 20,9; p-value = 0 , 0001).
Responsibility for fibrinolytic links of hemostasis is on the gene of plasminogen activator inhibitor (PLANH-1) on 4G / 5G polymorphism. This gene is the major antagonist of tissue plasminogen activator and urokinase, which in turn are plasminogen
activators that promote fibrinolysis (the dissolution of a blood clot). 4G allele had higher activity than the 5G, so the 4G allele carriers plasminogen concentration is higher than in carriers of allele 5G, which leads to an increased risk of thrombosis and thromboembolic complications in surgical patients. The frequencies of occurrence of homozygous 4G / 4G and heterozygotes 4G / 5G are significantly different in value in the main group (38.39% and 47.32%) compared with comparison groups (22.55 and 50.98%) and control (32,00 % and 43.00%, respectively) (%2 Pearson = 9,4; p-value = 0,05).
Conclusion. Molecular genetic studies of gene folate metabolism (MTHFR, MTRR, MTR), coagulation (FV, FII, FGB, ITGB3) and fibrinolytic hemostasis (PLANH-1) in the three study groups showed a significant adverse of carriage polymorphisms in the development of thromboembolic complications in surgical patients.
Table 1
Frequency of genotypes of polymorphisms of MTHFR, MTRR, MTR, PLANH-1, ITGB3, FGB, FII, FV genes, in %.
Genotype, allele Main group (n=112) Comparison group (n=102) Control group (n=100)
C677 T allele of MTHFR gene
n % ± m n % +m n % + m
CC 36 32,14+4,4 49 48,04+4,9 40 40,0+4,9
CT 61 54,46±4,7 45 44,12+4,9 55 55,0+5,0
TT 15 13,39+3,2 8 7,84+2,7 5 5,0+2,2
The differences in the three groups are valid: x2 Pearson = 9.3; p-value = 0,055
A1298C allele of MTHFR gene
AA 18 16,07+3,5 25 24,51+4,3 47 47,0+5,0
AC 70 62,50+4,6 58 56,86+4,9 43 43,0+5,0
CC 24 21,43+3,9 19 18,63+3,9 10 10,0+3,0
The differences in the three groups are valid: x2 Pearson = 26,8; p-value=0,0001
A66G allele of MTRR gene
AA 40 35,71+4,5 45 44,12+4,9 64 64,0+4,8
AG 63 56,25+4,7 50 49,02+4,9 30 30,0+4,6
GG 9 8,04+2,6 7 6,86+2,5 6 6,0+2,4
The differences in the three groups are valid: x2 Pearson = 17,9; p-value=0,001
A2756G allele of MTR gene
AA 51 45,54+4,7 74 72,55+4,4 66 66,0+4,7
AG 51 45,54+4,7 21 20,59+4,0 29 29,0+4,5
GG 10 8,93+2,7 7 6,86+2,5 5 5,0+2,2
The differences in the three groups are valid: x2 Pearson = 19,0; p-value=0,001
4G/5G allele of PLANH-1 gene
4G/4G 43 38,39+4,6 23 22,55+4,1 32 32,0+4,7
4G/5G 53 47,32+4,7 52 50,98+4,9 43 43,0+5,0
5G/5G 16 14,29+3,3 27 26,47+4,4 25 25,0+4,3
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ВЕСТНИК ХИРУРГИИ КАЗАХСТАНА № 2(43)-2015
The differences in the three groups are valid: x2 Pearson = 9,4; p-value=0,05
Leu33Pro allele of ITGB3 gene
Leu/ Leu 52 46,43±4,7 38 37,25±4,8 79 79±4,1
Leu/ Pro 59 52,68±4,7 63 61,76±4,8 20 20,0±4,0
Pro/ Pro 1 0,89±0,9 1 0,98±0,99 1 1,0±0,99
The differences in the three groups are valid: x2 Pearson = 37,2; p-value=0,0001
G (-455) allele of FGB gene
AA 8 7,14±2,4 11 10,78±3,1 2 2±1,4
AG 62 55,36±7,4 53 51,96±4,9 10 10±3,0
GG 42 37,50±4,6 38 37,25±4,9 88 88±3,2
The differences in the three groups are valid: x2 Pearson = 71,4; p-value=0,0001
G20210A allele of FII gene
AG 18 16,07 ± 3,5 5 4,90±2,1 3 3±1,71
GG 94 83,93±3,5 97 95,10±2,1 97 97±1,71
The differences in the three groups are valid: x2 Pearson = 14,2; p-value=0,001
G1691A allele of FV gene
AG 21 18,75±3,7 7 6,86±2,5 1 1 ±0,99
GG 91 81,25±3,7 95 93,14±2,5 99 99±0,99
The differences in the three groups are valid: x2 Pearson = 20,9; p-value=0,0001
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