Plasma amino acid concentrations (aminogram) show distinct patterns under various pathologic conditions. However, the plasma aminogram pattern in patients with pulmonary hypertension (PH) has not been elucidated. We sought to examine whether an aminogram could be predictive of clinical severity in patients with PH. We attained fasting plasma aminograms for 140 patients with PH and then compared the patient plasma amino acid levels with those of age- and gender-matched healthy control subjects. Aminograms revealed that the plasma concentrations of many amino acids were significantly different between patients with PH and healthy control subjects. We focused on the Fischer ratio (branched-chain amino acids/aromatic amino acids) as an integrated parameter. In all enrolled patients, Fischer ratio was negatively correlated with New York Heart Association functional class (ρ = −0.37, p <0.001), plasma B-type natriuretic peptide (ρ = −0.35, p <0.001), and pulmonary vascular resistance (ρ = −0.27, p = 0.002) and positively correlated with venous oxygen saturation (ρ = 0.27, p = 0.002) and 6-minute walk distance (ρ = 0.23, p = 0.016). Time course changes in Fischer ratio and in cardiac output were significantly correlated (ρ = 0.39, p = 0.024). The aminogram is changed in patients with PH, and in these patients, Fischer ratio decreases in proportion to the clinical severity of PH.
The relation between amino acids and pathologic conditions in the cardiovascular field has been recently clarified. For example, homoarginine, an amino acid derivative that increases nitric oxide availability and enhances endothelial function, was demonstrated to have a crucial role in cardiovascular risk and mortality, whereas amino acid l -arginine protects against ischemia/reperfusion-induced endothelial dysfunction in patients with type 2 diabetes mellitus and coronary artery disease. Furthermore, plasma l -arginine levels were strongly correlated with disease severity of idiopathic pulmonary arterial hypertension, and oral administration of l -arginine improves hemodynamics and exercise capacity in patients with precapillary pulmonary hypertension (PH). These findings raise the possibility that altered plasma amino acid levels are related to disease severity of PH and right-sided cardiac failure and, thus, could directly affect the biologic causal path. However, changes in plasma amino acid levels remain to be comprehensively examined in patients with PH. Therefore, the objectives of this study were to (1) explore plasma levels of amino acids in patients with PH and (2) identify potential noninvasive, amino acid–related biomarkers for predicting disease severity in PH.
Methods
In this study, we enrolled 140 consecutive patients with PH who attended Kyorin University Hospital, Japan, from April 2009 to August 2013. The group comprised 57 patients with chronic thromboembolic pulmonary hypertension (CTEPH), 28 with idiopathic pulmonary arterial hypertension, 27 with collagen-associated PH, 10 with PH related to congenital heart disease, 3 with pulmonary veno-occlusive disease, and 15 with other diseases. CTEPH was diagnosed by demonstration of organized pulmonary thromboembolism using contrast-enhanced lung computed tomography, perfusion lung scintigraphy, and pulmonary angiography; idiopathic pulmonary arterial hypertension by ruling out collagen vascular disease, pulmonary disease, and congenital heart disease; and pulmonary veno-occlusive disease by blood tests, pulmonary function tests, echocardiography, and high-resolution computed tomography. All patients provided informed consent, and the hospitals’ institutional review boards approved the study protocol. To compare characteristics and serum amino acid concentration against control subjects, age- and gender-matched healthy subjects (n = 140) were recruited for a cross-sectional study of prevalent cases on therapy from December 1, 2005, to April 1, 2006, from the Center for Multiphasic Health Testing and Services, Mitsui Memorial Hospital, Tokyo, Japan.
Fasting blood sampling for aminogram was performed basically at early morning of the same day as right-sided cardiac catheterization. The in-hospital diet was generally unified. Blood samples were collected into blood collection tubes with EDTA-2Na and then centrifuged to remove protein as a precipitate. The supernatant plasma was collected and stored at −80°C until analysis. The equal volume of blood was collected from each patient, and then, the equal volume of plasma was obtained in each sample. Plasma amino acid concentrations were measured by high-performance liquid chromatography using a cation-exchange column, as described previously.
Right atrial pressure (RAP), pulmonary arterial pressure (PAP), and pulmonary artery wedge pressure were measured at right-sided cardiac catheterization. Cardiac output (CO) was determined by the Fick technique using assumed oxygen consumption. Pulmonary vascular resistance (PVR) was calculated by subtracting pulmonary artery wedge pressure from mean PAP and dividing by CO. Patients underwent repeated right-sided cardiac catheterization every several months or years at the time of need to assess each condition. Six-minute walk distance (6MWD), plasma B-type natriuretic peptide (BNP) level, and other serologic markers were measured. New York Heart Association (NYHA) functional class was assessed for classification of severity.
Significant differences between the control and patient groups were determined using the Student t test or Mann-Whitney U test, as appropriate. Correlation between Fischer ratios and variables such as parameters related to severity of PH and those related to liver function was measured by calculating Pearson or Spearman correlation coefficients, as appropriate. Differences in frequencies were analyzed using Fisher’s exact probability test or chi-square test, as appropriate. All data are presented as mean ± SD. A value of p <0.05 was considered statistically significant.
Results
Supplementary Table 1 lists the baseline characteristics of the enrolled patients with PH and the age- and gender-matched healthy subjects (n = 140 each). Several parameters included in Supplementary Table 1 were significantly different between patients with PH and healthy control subjects, but the differences might be due to the pathologic characteristics of PH. The serum concentrations of amino acids measured by aminogram were then compared between these groups ( Supplementary Table 2 and Supplementary Figure 1 ). Intriguingly, most measured amino acid levels were significantly higher in patients with PH than in healthy control subjects, although the concentrations of arginine and tryptophan were significantly lower in patients with PH and only tyrosine and histidine were comparable between patients and control subjects. Because 40.7% of the enrolled patients in this study had CTEPH and a higher mean age than that of the other patients (60 ± 12 years in CTEPH vs 47 ± 15 years in other patients), the patients with CTEPH were analyzed individually. Table 1 lists the baseline characteristics of patients with CTEPH and other patients with PH. Supplementary Table 3 lists the baseline characteristics of enrolled patients with CTEPH and age- and gender-matched healthy subjects (n = 57 each). Similar to the results of all enrolled patients, levels of most measured amino acids were significantly higher in patients with CTEPH compared with their age- and gender-matched healthy control subjects ( Supplementary Table 4 , Supplementary Figure 2 ). The relation between all amino acids measured by aminogram including Fischer ratio, the molar concentration ratio of branched-chain amino acids (BCAAs) (valine + leucine + isoleucine) to aromatic amino acids (AAAs) (tyrosine + phenylalanine), and NYHA functional class was analyzed exhaustively. The Fischer ratio and 8 amino acid types (histidine, citrulline, leucine, α-aminobutyric acid, threonine, valine, essential amino acids, and BCAAs) had statistically significant relations with NYHA functional class. Importantly, Fischer ratio had the strongest correlation among them and was therefore the focus of further analysis in this study. BCAA and AAA levels in patients with PH were significantly higher than those in healthy control subjects, and Fischer ratio (BCAA/AAA) was significantly different between patients with PH and healthy control subjects ( Figure 1 ). Similar findings regarding BCAA, AAA, and Fischer ratio were also recognized in patients with CTEPH.
| Variable | All enrolled patients (n = 140) | Comparison of etiological difference | ||
|---|---|---|---|---|
| CTEPH patients (n = 57) | Other PH patients (n = 83) | p -value | ||
| male/female | 33/ 107 | 14/ 43 | 19/ 64 | 0.819 |
| Age (years) | 52 ± 15 | 60 ± 12 | 47 ± 15 | < 0.001 |
| NYHA functional class (I/II/III/IV) | 20/ 80/ 36/ 4 | 4/ 35/ 15/ 3 | 16/ 45/ 21/ 1 | 0.121 |
| Mean right atrial pressure (mmHg) | 5.5 ± 3.3 | 4.9 ± 3.1 | 5.7 ± 3.3 | 0.072 |
| Mean pulmonary arterial pressure (mmHg) | 37.1 ± 13.8 | 33.6 ± 12.0 | 39.2 ± 14.6 | 0.017 |
| Pulmonary vascular resistance (wood units) | 7.9 ± 5.7 | 7.5 ± 5.8 | 8.3 ± 5.6 | 0.176 |
| Cardiac output (l/min) | 4.3 ± 1.5 | 4.2 ± 1.3 | 4.4 ± 1.6 | 0.392 |
| Arterial oxygen saturation (%) | 93.6 ± 6.3 | 93.4 ± 4.0 | 93.7 ± 7.5 | 0.109 |
| B-type natriuretic peptide (pg/ml) | 113 ± 212 | 108 ± 218 | 117 ± 208 | 0.570 |
| six-minute-walk distance (m) | 435 ± 103 (n = 114) | 417 ± 103 (n = 50) | 449 ± 101 (n = 64) | 0.122 |
Fischer ratios decreased as NYHA functional class increased (ρ = −0.37, p <0.001), and the correlation was also recognized in patients with CTEPH (ρ = −0.42, p = 0.004; Figure 2 ). Accordingly, we also investigated the relations between Fischer ratio and other parameters correlated with severity and prognosis of PH including plasma BNP level and mixed venous oxygen saturation (SvO 2 ) ( Figure 3 ). In enrolled patients with PH, Fischer ratio was significantly correlated with 6MWD (ρ = 0.23, p = 0.016), BNP (ρ = −0.35, p <0.001), and SvO 2 (ρ = 0.27, p = 0.002). Moreover, in patients with CTEPH, Fischer ratio was significantly correlated with BNP (ρ = −0.46, p <0.001) and SvO 2 (ρ = 0.46, p <0.001).
Stay updated, free articles. Join our Telegram channel
Full access? Get Clinical Tree
