Pashkova: Variants of SNPs – the Brain Natriuretic Peptide Gene Polymorphism and Appropriate Structural and Functional Parameters of the Myocardial State in Men with Essential Hypertension and Chronic Heart Failure Living in Podillia Region



Problem statement and analysis of the recent research

Chronic heart failure (CHF) ranks first in the incidence of cardiovascular diseases [4]. Framingham Heart Study researchers identify that the incidence of heart failure doubles every decade, and its presence increases the risk of deaths by 4 times - ranging from 15% to 50%. Patients with hypertension develop this complication 6 times more often [6]. In Ukraine, the prevalence of clinically significant CHF NYHA functional class (FC) II-IV among the population is about 2 million people [5].

Despite the advance in modern medicine, mortality and hospitalization rates due to CHF remain to be very high; therefore, it is important to search for specific markers which would help estimate cardiovascular risk and effectiveness of treatment in patients with essential hypertension (EH) and CHF [2].

According to the European Society of Cardiology (ESC, 2016) guidelines concerning the diagnosis and treatment of acute and chronic heart failure, there is evidence for determining the brain natriuretic peptide (BNP) to exclude or confirm the diagnosis of heart failure in hospitalized patients with complaints of dyspnea. The threshold level of BNP plasma is ≥ 100 pg/ml in patients with acute onset of heart failure. In patients with gradual onset and stable course of CHF the threshold level of BNP plasma is ≥ 35 pg/ml. Normal peptide levels in untreated patient with dyspnea and ankle swelling have a high negative predictive value and deny heart failure as a possible cause of symptoms, which is especially important in the first contact with the patient [18]. Thus, in case of suspected heart failure the first step is to determine the plasma BNP level as an early indicator of initial functional disorders of the cardiovascular system [16].

The researchers of the Department of Internal Medicine No 2 of the Vinnytsia National Pirogov Memorial Medical University calculated the threshold BNP levels during screening studies of structural and functional disorders of the left ventricle (LV) in men with EH at the age of 40-60 years. The threshold BNP level equaling to 50 pg/ml, with an accuracy of 81.73% allows detecting individuals with diastolic dysfunction and preserved systolic function; the threshold BNP level equaling to 150 pg/ml, with an accuracy of 86.00 % may be used as an additional diagnostic criterion for systolic dysfunction in patients with EH [3, 7].

All natriuretic peptides are known to be produced from a common precursor; however, their production is regulated by own gene. In contrast to the already well-known biomarker BNP and its role in the early diagnosis and treatment of cardiovascular diseases, information about the structural organization of the BNP gene and the possible influence of its polymorphic genotypes on the plasma peptide levels are not numerous.

The BNP gene is located on the first chromosome and consists of three exons and two introns. The most physiologically significant polymorphism of the BNP gene was determined and studied, namely, the replacement of thymine by cytosine at position 381 (T-381C) [1, 8, 12]. This polymorphism among the Ukrainian population, however, has not been studied. Considering modern achievements in the study of the pathogenesis of CHF, we can assume the influence of the BNP gene polymorphism on the development and progression of cardiac manifestations of the disease.

The objective of the research was to examine structural and functional indicatorsof the myocardial state in men with EH and co-existent CHF Class IIА being carriers of different variants of the BNP gene at the age of 40-60 years living in Podillia region.

Materials and methods

There were examined 141 men at the age of 40-60 years living in Podillia region. Among them, 62 men (Group I) were diagnosed with stage II EH, stage II-III LV hypertrophy and CHF NYHA FC 0-І (the average age was 49.19±0.66 years) and 50 men (Group II) were diagnosed with EH and co-existent CHF Class IIA according to the classification of M.D. Strazhesko – V.H. Vasylenko and NYHA FC II-III (the average age was 50.14±0.99 years). Patients of both groups were representative according to age. The diagnosis of EH and CHF weremade on the basis of the patients’ complaints, anamnesis, physical examination, laboratory and instrumental methods of examination according to the guidelines of the European Society of Hypertension (ESH) and the ESC in 2013 аnd 2016 ESC guidelines for the diagnosis and treatment of acute and chronic heart failure [9, 17]. All patients were observed since December 2013 to July 2014.

Exclusion criteria included symptomatic arterial hypertension (AH), impaired renal and liver functions, coronary heart disease prior to EH, endocrine, hematological, neoplastic and autoimmune disorders, patients with EH complications, namely, myocardial infarction, acute cerebrovascular accident. Genotyping of the BNP gene was performed using the polymerase chain reaction after isolation of genomic DNA from white blood cells of venous blood. This study was carried out in collaboration with the Research Institute of Genetic and Immunological Grounds of Pathology and Pharmacokinetics - Poltava Ukrainian Medical and Dental Academy. The frequency distribution of gene polymorphism among the population was tested according to the Hardy-Weinberg equilibrium equation.

Structural and functional indicators of the myocardial state were assessed by cardiac ultrasound with ultrasound device ULTIMA RA (Radmir, Ukraine). By means of linear dimensions we determined LV mass using the formula recommended by the American Society of Echocardiography (ASE) and subsequent calculation of the left ventricular myocardium mass index (LV MMI) [13]. In accordance with the Penn Convention and ASE formula the LV MMI ≥ 115 g/m2 was considered as a criterion for LV hypertrophy in men according to the guideline of the ESH and the ESC in 2013 [9]. LV diastolic function was evaluated according to current recommendations [15]. The state of LV systolic function was evaluated by the value of ejection fraction (EF). Systolic function was considered preserved when EF was >45%. The mathematical processing was performed on a personal computer using a standard statistical package STATISTICA 6.1.

Results and discussion

The frequency distribution of the BNP genotypes among men with EH of varying severity living in Podillia region corresponded to the Hardy-Weinberg equilibrium. In men with stage II EH the frequency of the T381T genotype of the BNP gene was 35.48% (n=22), the frequency of the T381C genotype was 48.39% (n=30) and the frequency of the C381C genotype was 16.13% (n=10) (рсс-тт>0.05; ртс-сс≤0.05; ртс-тт>0.05). Among patients with CHF Class IIA, the frequency of the T381T genotype of the BNP gene was 42.00% (n=21), the frequency of the T381C genotype was 46.00% (n=23) and the frequency of the C381C genotype was 12.00% (n=6) (рсс-тт>0.05; ртс-сс≤0.05; ртс-тт>0.05). In men with EH of varying severity, the Т381С genotype and C allele of the BNP gene prevailed (Fig. 1).

Fig. 1.

Frequency distribution of the BNP genotypes and alleles in men with stage II EH and patients with EH and co-existent CHF Class IIA (%) living in Podillia region.

gmj-23-gmj.2016.3.49-g1.jpg

the difference is significant (p<0.05) when compared to:

* - the T381C genotype/C allele within each group

The obtained results were consistent with the data obtained when studying the residents of other populations of the world. It was found that among patients with EH living in the USA [8], Novosibirsk region of Russian Federation [1] and Germany [11] carriers of the T381С genotype and C allele of the BNP gene predominated. The frequencies of polymorphic genotypes of the BNP gene did not differ significantly between patients from different populations.

During statistical analysis due to the small number of carriers of С381С genotype, T381С heterozygote and С381С homozygote were combined in one group - C allele carriers. The frequency of C allele carriers among men with stage II EH was 64.52% (n=40), among men with EH and co-existent CHF Class IIA – 58.00% (n=29).

The analysis of structural and functional indicators of the myocardial state in men with stage II EH being carriers of different variants of the BNP gene found that in patients with T381T genotype the indicators of the size and volume of the LV: end diastolic diameter (EDD), end systolic diameter (ESD), end-diastolic volume index (EDVI), end-systolic volume index (ESVI), cardiac index (CI), LV MMI, minute blood volume index (MBVI), interventricular septal thickness (IST) and posterior left ventricular wall (PLVW) thickness were significantly higher than in carriers of C allele of the BNP gene (р<0.05).

In patients with symptoms of CHF Class IIA being carriers of the Т381Т genotype the size and volume of the LV at the end of systole and diastole were significantly larger, the indicators of the LV MMI and LV wall thickness were higher than in carriers of C allele of the BNP gene (р<0.05). Among men with EH and co-existent CHF Class IIA, relative wall thickness (RWT) was significantly lower in T381T homozygotes than in carriers of C allele and can display a tendency to the formation of LV eccentric hypertrophy. When studying systolic function in patients with symptoms of CHF Class IIA it was found that EF was the lowest in carriers of the Т381Т genotype (р<0.05). The indicators of the size and volume of the LV, PLVW thickness and systemic hemodynamics (systolic blood pressure (SBP), diastolic blood pressure (DBP)) regardless of the BNP genotype were significantly higher in men with EH and co-existent CHF Class IIА compared to patients with stage II EH (p<0.05) (Table 1).

Table 1.

Structural and functional indicators of the myocardial state and systemic hemodynamics in men with stage II EH and patients with symptoms of CHF Class IIA being carriers of different variants of the BNP gene, (M±m)

Indicators Patients with stage II EH, T381T homozygotes (n=22) Patients with stage II EH, carriers of C allele (n=40) Patients with CHF Class IIA, T381T homozygotes (n=21) Patients with CHF Class IIA, carriers of C allele (n=29) р
EDD,cm 5.19±0.08 4.61±0.06 5.73±0.03 5.58±0.03 р2-1#;р3-1*; р4-2*;р4-3*
ESD, cm 3.51±0.09 3.19±0.05 4.69±0.03 4.16±0.08 р2-1#;р3-1*; р4-2*;р4-3*
PLVW thickness, cm 1.32±0.02 1.18±0.02 1.37±0.01 1.30±0.01 р2-1*;р3-1*; р4-2#;р4-3*
IST,cm 1.36±0.03 1.22±0.02 1.39±0.02 1.31±0.01 р2-1*; р4-2#;р4-3*
RWT, st.units 0.54±0.03 0.51±0.01 0.43±0.002 0.47±0.006 р3-1*; р4-2*;р4-3*
LV MMI, g/m2 176.24± 3.43 142.13± 4.65 204.61± 6.69 183.53± 3.19 р2-1#;р3-1*; р4-2#;р4-3*
EDVI, ml/m2 61.07±1.67 53.20±0.92 82.57±2.49 71.63±1.40 р2-1*;р3-1#; р4-2#;р4-3*
ESVI, ml/m2 26.28±1.26 21.14±0.80 49.78±2.95 37.46±0.87 р2-1*;р3-1#; р4-2#;р4-3*
EF,% 56.34±1.38 59.92±1.40 42.08±0.53 48.62±0.68 р3-1#;р4-2#; р4-3*
MBVI, ml/m2 3364.21± 162.77 2528.75± 121.14 2795.45± 122.04 2068.62± 73.88 р2-1*;р3-1*; р4-2*;р4-3*
СІ,l/m2 3.26±0.51 2.43±0.14 3.31±0.08 2.51±0.19 р2-1*;р4-3*
SBP,mm Hg 159.80± 1.11 141.60± 2.05 177.64± 4.29 166.52± 1.64 р2-1*;р3-1*; р4-2*;р4-3*
DBP,mm Hg 91.25± 1.39 84.60± 1.06 109.22± 4.09 101.13± 1.99 р2-1*;р3-1*; р4-2*;р4-3*

Notes:

* - the difference is significant at p<0.05;

# - the difference is significant at р<0.01.

The next step of the study was the analysis of frequency distribution of the BNP genotypes among men with EH and co-existent CHF Class IIA with different states of LV systolic function. For this purpose, patients with symptoms of CHF were distributed into two groups – Group I included 20 patients with preserved systolic function (EF>45%), Group II included 30 men with reduced systolic function (EF<45%). Among patients with the absence of LV systolic dysfunction carriers of C allele (73.70%) dominated significantly (p<0.05). However, in patients with the presence of LV systolic dysfunction T381T homozygotes of the BNP gene prevailed - 60% of patients (р≤0.05) (Fig. 2).

Fig. 2.

Frequency distribution of the BNP genotypes among patients with EH and co-existent CHF Class IIA with different states of LV systolic function (%) living in Podillia region.

gmj-23-gmj.2016.3.49-g2.jpg

Notes:

* - the difference is significant when compared within the group with preserved systolic function (p<0.05);

# - the difference is significant when compared within the group with the presence of LV systolic dysfunction (p<0.05);

& - the difference is significant when compared between the BNP genotypes (p<0.05).

The previous study found differences in frequency distribution of the BNP e genotypes among patients with CHF Class II A with different states of LV systolic function, therefore, it was decided to continue the study of structural and functional indicators of the myocardial state in these groups.

In patients with EH and co-existent CHF Class IIА (EF>45%) significantly higher indicators of EDD, ESD, IST, PLVW thickness, EDVI, ESVI, LV MMI, MBVI were observed. When studying the parameters of central hemodynamics, the levels of SBP and DBP in carriers of the Т381Т genotype were significantly higher than in carriers of C allele (р<0.05); however, there was no significant differences in heart rate (HR) between patients of this group (р>0.05). In patients with EH and co-existent CHF Class IIА with preserved systolic function, the levels of blood pressure (BP) were significantly higher in T381T homozygotes of the BNP gene and corresponded to stage II AH; among C allele carriers they corresponded to stage I AH.

Among patients with EH and co-existent CHF Class IIА (EF<45%) significantly higher indicators of the size of the LV, EDVI, ESVI, and LV MMI were observed in carriers of the Т381Т genotype. Among patients with EH and co-existent CHF Class IIА and decreased systolic function, the levels of BP were significantly higher in T381T homozygotes of the BNP gene and corresponded to stage III AH; among C allele carriers they corresponded to stage II AH. One of the possible explanation of the obtained results is the presence of lower concentrations of the BNP in T381T homozygotes [17], since the BNP is the most universal counter-regulating substance against the activity of the renin–angiotensin–aldosterone system. The latter one, according to most researchers, is partially associated with changes in the myocardium in chronic BP increase (Table 2).

Table 2.

Structural and functional indicators of the myocardial state and systemic hemodynamics in men with symptoms of CHF Class IIA and different states of LV systolic function being carriers of different variants of the BNP gene, (M±m)

Indicators T381T homozygotes, EF>45% (n=7) T381T homozygotes, EF<45% (n=14) Carriers of C allele, EF>45% (n=13) Carriers of C allele, EF<45% (n=16) р<0.05
EDD,cm 5.59±0.03 6.02±0.05 5.43±0.03 5.88±0.04 р2-13-1; р4-24-3
ESD, cm 4.51±0.07 4.87±0.06 4.11±0.05 4.29±0.08 р2-13-1; р4-24-3
PLVW thickness, cm 1.36±0.02 1.43±0.04 1.28±0.03 1.35±0.02 р2-13-1; р4-24-3
IST, cm 1.35±0.03 1.44±0.02 1.27±0.01 1.36±0.01 р2-13-1; р4-24-3
RWT, st.units 0.46±0.002 0.42±0.002 0.52±0.004 0.47±0.003 р2-13-1; р4-24-3
LV MMI, g/m2 193.42±5.45 215.61±4.68 171.53±3.19 195.19±4.23 р2-13-1; р4-24-3
EDVI, ml/m2 76.57±2.49 89.57±3.23 65.63±1.62 77.63±3.16 р2-13-1; р4-24-3
ESVI, ml/m2 40.34±2.17 51.78±3.35 31.46±1.92 42.46±2.68 р2-13-1; р4-24-3
EF,% 46.12±162 35.61±0.44 49.62±1.36 44.62±0.67 р2-14-2; р4-3
MBVI, ml/m2 2849.02± 81.13 2451.83± 136.95 2106.62± 73.88 2023.62± 73.88 р3-1
SBP,mm Hg 166.64± 2.67 187.64± 3.24 153.41± 1.64 176.52± 2.13 р2-13-1; р4-24-3
DBP,mm Hg 107.67± 3.32 116.14± 3.06 97.08± 1.65 104.52± 1.52 р2-13-1; р4-2
HR, bpm 84.89±2.03 95.89±2.03 73.95±1.50 82.38±1.50 р4-3

The results of hemodynamic changes in patients with EH and CHF were consistent with the data obtained by other researchers. Thus, E.N. Berezikova (2013) found that the inheritance of the T381T genotype with the presence of T allele is associated with high risk of the development of CHF, its severe and unfavorable course in patients with coronary heart disease [1]. The study of I. Gorący et al. (2015) conducted in Poland among healthy newborns of both sexes found that in boys the inheritance of the Т381Т genotype and T allele is associated with larger LV mass and higher indicators of DBP at birth than in T381C heterozygotes and С381С homozygotes and carriers of С allele of the BNP gene [10]. K.L. Ellis et al. (2011) showed that patients with coronary heart disease of both sexes being carriers of the Т381Т genotype and T allele of the BNP gene have significantly higher indicators of the EDVI and ESVI than carriers of C allele of the BNP gene [14].

Conclusions

  1. Among men with EH of varying severity living in Podillia region the Т381С genotype and C allele of the BNP gene dominate (р<0.05).

  2. In patients with symptoms of CHF Class IIA being carriers of the Т381Т genotype the size and volume of the LV at the end of systole and diastole were significantly larger, the indicators of the LV MMI and LV wall thickness were higher than in carriers of C allele of the BNP gene (р<0.05).

  3. In patients with EH and co-existent CHF Class IIА (EF<45%) being carriers of the Т381Т genotype, the highest indicators of the size of the LV, ESVI, EDVI, LM MMI were observed.

  4. The inheritance of the T381T genotype by patients with symptoms of CHF Class IIA was found to be associated with higher levels of SBP and DBP irrespective of the state of LV systolic function.

Prospects for further research

We plan to investigate plasma levels of the BNP in men with EH and CHF being carriers of various BNP genotypes at different structural and functional indicators of the myocardial state.

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Copyright (c) 2017 Іuliia P Pashkova

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