Relationship between N-terminal pro-B type natriuretic peptide and extensive echocardiographic parameters in mild to moderate aortic stenosisM Cemri, U Arslan, SA Kocaman, A Cengel
Department of Cardiology, Gazi University Medical School, Ankara, Turkey
Context: The N-terminal pro-B type natriuretic peptide levels (NT pro-BNP) are increased in cases of volume or pressure overload. Aims: To examine NT pro-BNP levels and enclose whether any relationship is present between the levels of NT pro-BNP and extensive echocardiographic parameters in asymptomatic patients with mild to moderate aortic stenosis (AS). Settings and Design: A cross-sectional study about the NT pro-BNP levels was conducted in 37 asymptomatic AS patients and compared with 40 controls. Methods: Patients <70 years old with mild to moderate AS with a peak transaortic gradient >20 mm Hg in transthoracic echocardiogram were included in our study. Extensive echocardiographic parameters and NT pro-BNP levels were obtained from these patients and these indices were compared with the control population selected from the patients who had similar clinical characteristics with the AS patients. Statistical Analysis: NT-proBNP values were found to be distribution free. Spearman correlation coefficient was used for correlation analysis. Mean values were compared by the Kruskal-Wallis test. Results: The NT pro-BNP levels were increased in patients with AS (median; interquartiles range: 686 [449-855] pg/mL vs. 140 [116-150] pg/mL, P < 0.001). Among patients with AS, when correlation analysis was performed mean transaortic gradient, aortic valve area index, myocardial performance index, E m /A m ratio, left-ventricular mass index (LVMI) and E/E m ratio had correlations (r=0.38, P = 0.026; r=-0.46, P =0.008; r=0.19, P =0,049; r=-0.22, P =0.04, r=0.49, P =0.003 and r=0.53, P <0.001 respectively) with plasma NT pro-BNP levels. The LVMI (r = 0.49, P = 0.003) and E/E m ratio (r = 0.53 P < 0.001) have the strongest correlations when compared to other parameters. Conclusion: Plasma NT pro-BNP levels are increased in even asymptomatic patients with AS and correlated with several echocardiographic parameters related to severity of AS and degree of diastolic dysfunction. As a result, NT pro-BNP levels may be used in the follow-up of asymptomatic patients having mild to moderate AS.
Keywords: Aortic stenosis, echocardiography, N-Terminal pro-BNP
Brain natriuretic peptide and N-terminal pro-B type natriuretic peptide (NT pro-BNP) have been shown to be increased in plasma in states of pressure or volume overload. ,, The NT pro-BNP has been demonstrated to be increased in patients with severe aortic stenosis (AS) and it correlates with functional status,  mean pressure gradient  and aortic valve area  and predicts symptom development in asymptomatic AS and postoperative outcome after valve replacement.  The NT-proBNP is elevated in patients with aortic valve disease linked to disease severity and decreases after successful surgical therapy but increases in conservatively treated patients. , Brain natriuretic peptide has been also found to be a predictor of outcome and helpful for risk stratification in patients with AS. 
Therefore, NT pro-BNP levels can be used as a simple biomarker for the follow-up of AS in outpatient clinics. With this hypothesis, our aim was to examine the levels of NT pro-BNP levels and study the relationship between the levels of NT pro-BNP and extensive echocardiographic parameters in asymptomatic patients having mild to moderate AS.
Asymptomatic patients <70 years old with mild to moderate AS with a peak transaortic gradient >20 mm Hg in transthoracic echocardiogram were included in our study. Mild and moderate AS was defined as previously published.  Mean transaortic gradients were used to differentiate mild and moderate AS. Mean gradients between 25 and 40 mm Hg were accepted as moderate AS and mean gradients <25 mm Hg were accepted as mild AS. Echocardiographic parameters and NT pro-BNP levels were obtained from these patients and these indices were compared to the control population selected from the patients who had similar clinical characteristics with the AS patients. The patients with no cardiovascular disease constituted the control group and they were using no drugs primarily effecting the cardiovascular system. Informed consent was obtained from all subjects and the study protocol was approved by the local ethical committee. All the patients were in class I according to New York Heart Association (NYHA) classification. In patients with AS, the echocardiographic parameters were inserted in a correlation analysis with plasma NT pro-BNP levels.
Patients with hypertension, diabetes mellitus, atrial fibrillation, coronary artery disease defined as >50% narrowing in at least one coronary artery in a previous angiogram, history of myocardial infarction, acute coronary syndrome or typical angina pectoris, renal failure (serum creatinine level >1.5 mg/dL) and concomitant valvular disease such as mitral stenosis (mitral valvular area <2.5 cm 2 ) and moderate to severe aortic or mitral regurgitation were excluded from the study.
NT pro-BNP measurement
Venous blood from all patients was taken from an antecubital vein and placed in tubes filled with ethylenediaminetetraacetic acid. The specimens were centrifuged for 1 h and plasma was frozen at -80°C until analysis. The NT pro-BNP was measured by an electrochemiluminescence immunoassay (Elecsys proBNP, Roche Diagnostics, Mannheim, Germany).
All patients underwent complete transthoracic studies including two dimensional, color flow and pulsed Doppler as well as tissue Doppler imaging with a GE-Vingmed Vivid 7 system (GE-Vingmed Ultrasound AS, Horten, Norway) using a 2.5-3.5 MHz transducer. Standard transthoracic views were used to obtain left ventricular ejection fraction (EF), left ventricular end-diastolic diameter (LVEDD), interventricular septal diameter (IVSD), and left ventricular posterior wall diameter (PWD) in diastole. The LV diameters, EF, and the left-ventricular mass index (LVMI) were determined from M-mode traces recorded from the parasternal long-axis view according to the established standard.  Left ventricular mass (LV mass, in grams) was calculated according to the following formula: 1.04 × [(LVEDD + IVSD + PWD) 3 - LVEDD 3 ] - 13.6 (diameter in centimeters).  All dimensions were indexed to body surface area (BSA). The LVMI was calculated by division of LV mass by BSA.
With continuous Doppler, mean, and maximum gradients between aorta and left ventricle were assessed and aortic valve area (AVA) and AVA index (AVAi) were calculated. These were taken as indices of AS severity.
Transmitral pulsed Doppler was recorded in the apical 4-chamber view: early (E) and atrial (A) peak velocities, E/A ratio, isovolumic relaxation time (IVRT) (ms), isovolumic contraction time (IVCT) (ms), and ejection time (ET) were calculated. Myocardial performance index (MPI) was measured as (IVRT + IVCT)/ET.
With tissue Doppler imaging, in apical 4 chamber view, a 5-mm pulsed Doppler sample volume was placed at the level of septal mitral annulus. Peak systolic (Sm), peak early (Em) and late (Am) diastolic mitral annular velocities, and Em/Am ratio were calculated.
The intraobserver variability of echocardiographic measurements was <6% and all examinations were performed by an experienced cardiologist who was blinded to the patient's clinical information.
Continuous variables were expressed as mean ± SD; categorical variables were defined as percentage. Data were tested for normal distribution using the Kolmogorov-Smirnov test. Continuous variables of normally and nonnormally distributed variables were compared with an independent t -test and Mann-Whitney U -test and the χ2 test was used for the categorical variables. The NT-proBNP values were not normally distributed. Spearman correlation coefficient was used for correlation analysis. Mean values were compared by Kruskal-Wallis test among different groups. All tests of significance were two-tailed. Statistical significance was defined as P < 0.05. The SPSS statistical software (SPSS for windows 11.5, Inc, Chicago, IL, USA) was used for all statistical calculations.
Thirty-seven patients with AS (17 mild, 20 moderate) were enrolled in the study and their NT pro-BNP levels were compared to 40 control patients. The baseline characteristics of the patients were highlighted in [Table - 1]. NT pro-BNP levels were significantly increased in patients with mild to moderate AS [Table - 1]. [Figure - 1] demonstrates the NT pro-BNP levels in control group, mild AS, and moderate AS. There was a step-by-step increase with the increase in the severity of AS [median; interquartiles range: 140 (116-150), 490 (267-679), and 890 (718-1140), respectively].
Among patients with AS, when correlation analysis was performed mean transaortic gradient, AVA index, MPI, E m /A m ratio, LVMI, and E/E m ratio had correlations with plasma NT pro-BNP levels [Table - 2]. The LVMI and E/E m are the two parameters which have the strongest correlations [Figure - 2],[Figure - 3].
The diagnostic value of BNP and NT-proBNP has been mainly investigated in patients with heart failure. In a large number of studies, it has been consistently found that BNP and NT pro-BNP are elevated in patients with heart failure and may be used for prognostic assessment and follow-up of heart failure treatment.  The NT pro-BNP levels are used mainly to rule out acute heart failure in an emergency setting if NT-proBNP values are <300 pg/mL.  Besides the levels of NT pro-BNP have been shown to be elevated in stable and unstable coronary artery disease and several valvular diseases. 
In the present study, we have found increased NT pro-BNP levels even in asymptomatic patients with mild to moderate AS. The increase in plasma NT pro-BNP levels was also correlated with several echocardiographic parameters especially LVMI and E/E m ratio. Our results are in concordance with several studies in the literature in which the NT pro-BNP levels have been studied in AS and confirm previously reported findings. ,,,,
In AS, the elevated left ventricular systolic wall stress due to increased afterload acts as a stimulus for myocardial cell hypertrophy characterized by the increased expression of natriuretic peptides in the ventricle , which is a cause of increased plasma BNP levels in patients with AS. Reverse LV remodeling after aortic valve replacement with regression of myocardial hypertrophy results in improvement of LV longitudinal myocardial strain and decrease of NT pro-BNP plasma levels.  The relation observed between the concentrations of natriuretic peptides and LVMI in the study by Qi et al. ,  was closer for BNP and NT-proBNP than for ANP and NTproANP; which supports other studies by Kohno et al.  and Yamamoto et al. ,  who found that BNP and ANP plasma concentration was higher in hypertensive patients with left ventricular hypertrophy than in those without. In our study, we also observed a significant correlation between LVMI and NT-pro BNP levels which is supported by the above-mentioned studies.
Surprisingly, the NT pro-BNP levels were found to be high for asymptomatic patients in our study. Bergler-Klein et al. found a cut-off point at 80 pmol/L (approximately 670 pg/mL) in their study for prediction of symptom development in AS.  Weber et al. determined a cut off point of 900 pg/mL in patients >50 years and 450 pg/mL in patients <50 years to rule in heart failure.  We found a median NT pro-BNP level, 686 pg/mL, which is similar to these cut-off values in asymptomatic patients with mild to moderate AS. However, our results are consistent with another study by Weber et al. , in which the NT pro-BNP levels have a mean of 601 pg/mL in asymptomatic patients with AS.  In this study, the plasma levels of NT-pro BNP had a mean of 1467 pg/mL in severe asymptomatic AS, in New York Heart Association class IV patients with severe AS, these levels had been found to be as high as a mean of 5273 pg/mL. These different values for NT-proBNP may be caused by the patient selection, mean age of the population, different selection of transaortic gradients to determine AS severity, and different methods for NT pro-BNP measurements in different studies.
The elevated plasma levels of NT pro-BNP in severe AS were observed in several studies. ,, In these studies, this increase was related to functional capacity, mean transaortic gradient, and AVA. We have also found correlations between mean gradient and AVA indices and plasma NT pro-BNP levels in patients with mild to moderate AS. Besides Weber et al.  found a correlation between the severity of AS and plasma NT pro-BNP levels and in this study, patients having a mean transaortic gradient <30 mm Hg had even increased NT pro-BNP levels. Our results are in concordance with this finding, because plasma NT pro-BNP levels are higher in patients having moderate AS than patients having mild AS and in patients with mild AS than the control group.
An important aspect of our study is the relationship between the extensive echocardiographic parameters including tissue Doppler parameters and plasma NT pro-BNP levels. This relationship is disclosed in even asymptomatic patients with mild to moderate AS in our study. Especially E/Em ratio and LVMI are correlated with the plasma NT pro-BNP levels. These findings are supported by several studies in the literature. The E/Em ratio has been shown to be an important predictor for left ventricular filling pressures.  Dong et al.  have recently found a correlation between NT pro-BNP levels with echocardiographically determined left ventricular diastolic functions including E/Em ratio in ambulatory patients. The plasma NT pro-BNP levels are also increased in left ventricular hypertrophy. Yamamoto et al.  showed increased BNP levels in hypertensive patients with left ventricular hypertrophy. In another study,  it has been found that amount of left ventricular hypertrophy determines the plasma NT pro-BNP level in patients with hypertrophic cardiomyopathy and normal left ventricular EF. Our results implicating a relationship between LVMI and plasma NT pro-BNP levels are also consistent with these findings.
According to our findings, plasma NT pro-BNP levels may be useful in the follow-up of patients with mild to moderate AS because they can give information about the severity of disease, the degree of diastolic dysfunction and the onset of symptoms as these levels are correlated with E/Em ratio which is a useful measure concerning left ventricular end diastolic pressures. In one study, there has been a direct correlation between E/Em ratio and pulmonary capillary wedge pressure  which is one of the best measures for deterioration of left ventricular functions. Therefore, this ratio may be used for determination of the onset of heart failure symptoms in AS.
Our study has several limitations. First of all, the coronary angiography data of the patients were not available as these patients had no symptoms to warrant coronary angiography. The study was a cross-sectional one and no long-term follow-up of these patients were available. As a result, the outcomes of the patients were unknown.
In conclusion, plasma NT pro-BNP levels are increased in even asymptomatic patients with AS and correlated with several echocardiographic parameters related to severity of AS and degree of diastolic dysfunction. As a result, NT pro-BNP may be used in the follow-up of asymptomatic patients having mild to moderate AS.
[Figure - 1], [Figure - 2], [Figure - 3]
[Table - 1], [Table - 2]