- Email: [email protected]
The pivotal role of eNOS uncoupling in vascular endothelial dysfunction in patients with heart failure with preserved ejection fraction
International Journal of Cardiology 190 (2015) 335–337
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
The pivotal role of eNOS uncoupling in vascular endothelial dysfunction in patients with heart failure with preserved ejection fraction Eiichiro Yamamoto a,⁎, Yoshihiro Hirata a, Takanori Tokitsu a, Hiroaki Kusaka a, Kenji Sakamoto a, Megumi Yamamuro a, Koichi Kaikita a, Hiroshi Watanabe b, Seiji Hokimoto a, Seigo Sugiyama a,c, Toru Maruyama b, Hisao Ogawa a a b c
Faculty of Life Sciences, Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan Cardiovascular Division Diabetes Care Center, Jinnouchi Hospital, Kumamoto, Japan
a r t i c l e
i n f o
Article history: Received 9 February 2015 Accepted 18 April 2015 Available online 21 April 2015 Keywords: Heart failure with preserved left ventricular function Vascular endothelial dysfunction eNOS uncoupling
Vascular endothelial dysfunction correlates with cardiovascular diseases and plays an important role in all stages of atherosclerosis, leading to various cardiovascular diseases [1]. Furthermore, heart failure (HF) with preserved left ventricular ejection fraction (LVEF) (HFpEF) is one of the cardiovascular diseases of which the optimal treatment has not been established [2], because the precise pathophysiological mechanism underlying HFpEF still remains unclear. Recently, we reported that peripheral endothelial function, assessed by fingertrip digital reactive hyperemia peripheral arterial tonometry (RH-PAT), is significantly impaired in HFpEF patients than in non-HF patients [3]. Furthermore, using Dahl salt-sensitive hypertensive rats (DS rats), which are useful experimental models of HFpEF, we have previously elucidated molecular mechanism underlying endothelial dysfunction in HFpEF. We also demonstrated the association of endothelial nitric oxide synthase (eNOS) uncoupling, one of the sources of reactive oxygen species (ROS), with the progression of endothelial dysfunction-induced HFpEF [4]. ROS generally oxidizes tetrahydrobiopterin (BH4), an essential cofactor for eNOS, to its oxidative form 7,8-dihydrobiopterin (BH2) in the endothelial cells, leading to eNOS uncoupling which is at least partly involved in cardiovascular diseases. To our knowledge, however, there is no clinical information on correlations between endothelial ⁎ Corresponding author at: Department of Cardiovascular Medicine, Faculty of Life Sciences, Graduate School of Medical Science, Kumamoto University, 1-1-1 Honjo, Chuo-ku, Kumamoto 860-8556, Japan. E-mail address: [email protected] (E. Yamamoto).
http://dx.doi.org/10.1016/j.ijcard.2015.04.162 0167-5273/© 2015 Elsevier Ireland Ltd. All rights reserved.
dysfunction and pteridine such as BH4 and BH2 in HFpEF patients. In this study, hence, we examined the significance of plasma BH4 and BH2 in endothelial dysfunction in HFpEF. We recruited consecutive HFpEF patients hospitalized in Kumamoto University Hospital, and examined peripheral endothelial function by RH-PAT using the Endo-PAT2000 (Itamar Medical, Caesarea, Israel), and biomarkers (plasma pteridine and serum derivatives of reactive oxidative metabolites [DROM, Diacron srl, Grosseto, Italy]; normal range: 250–300 unit called the Carratelli unit [U.CARR], a new biomarker of ROS) in blood vein at stable condition after the optimal therapy. We defined HFpEF clinically according to the criteria of the European Working Group to HFpEF [5]: 1) symptoms of HF; 2) normal or mildly reduced LV systolic function (LVEF N 50% and LV end-diastolic volume index b 97 ml/m2); and 3) evidence of abnormal LV diastolic distensibility. Plasma BH4 and BH2 levels were measured directly by highperformance liquid chromatography (HPLC) with the electrochemical detection method, as previously described [6]. Baseline characteristics of patients are shown in Table 1. Compared to risk factor (number of patients, age, sex, and equal incidence of hypertension, diabetes mellitus, dyslipidemia and coronary artery disease)-matched non-HF patients, peripheral endothelial dysfunction, indicated by reduced RHI values, significantly occurred in risk factormatched HFpEF patients (2.01 [1.64–2.42] vs. 1.70 [1.55–1.88], P b 0.001), consistent of our previous clinical report [3]. In association with a reduction in RHI, plasma BH4 levels tended to decrease (33.4 ± 24.1 nM vs. 26.3 ± 15.9 nM, P = 0.051), BH2 levels significantly increased (13.4 ± 9.5 nM vs. 16.8 ± 10.1 nM, P = 0.047) and subsequently the BH4/BH2 ratio greatly decreased (3.21 ± 2.05 vs. 2.05 ± 1.62, P b 0.001) in HFpEF patients. Furthermore, serum DROM levels were significantly increased in HFpEF patients than in non-HF patients (312.5 [294.8–396.3] U.CARR vs. 384.0 [348.5–426.0] U.CARR, P b 0.001), accompanied by inverse reduction in BH4/BH2 ratio. Moreover, plasma BH4/BH2 ratios had significant and negative correlation with DROM values (P = 0.04, r = − 0.181, Fig. 1A) and the ratio of early-transmitral flow velocity to tissue doppler early-diastolic mitral annular velocity (E/e′) (P = 0.003, r = −0.26, Fig. 1C), and strong positive correlation with ln-RHI (P = 0.009, r = 0.233, Fig. 1B) in all enrolled patients. These data indicated that pteridine metabolism disorder induced-ROS/NO imbalance (eNOS uncoupling) could be involved
336
E. Yamamoto et al. / International Journal of Cardiology 190 (2015) 335–337
Table 1 Baseline characteristics of 64 risk factor-matched non-HF patients and 64 risk factor-matched HFpEF patients.
Age (years) Sex (male, %) BMI (kg/m2) CAD (yes, %) Hypertension (yes, %) DM (yes, %) Current smoking (yes, %) Dyslipidemia (yes, %) BH4 (nM) BH2 (nM) BH4/BH2 DROM (U.CARR) RHI BNP (pg/ml) eGFR (ml/min/1.73 m2) Hs-CRP (mg/l) LVEF (%) E/e′ β-Blockers (%) ACEI or ARB (%) CCB (%) HMG-CoA RI (%) MR blockers (%) Loop diuretics (%)
All patients (n = 128)
Matched non-HF (n = 64)
Matched HFpEF (n = 64)
P valuea
68.1 (9.3) 58.6 23.6 (3.5) 62.5 85.2 32.8 10.2 87.5 29.9 (20.7) 15.1 (9.9) 2.63 (1.93) 362.3 (294.8–396.3) 1.8 (1.62–2.15) 37.5 (19.8–80.9) 64.2 (19.0) 0.6 (0.3–0.9) 62.3 (5.7) 14.0 (4.9) 64.1 66.1 53.9 68.0 14.1 10.2
68.0 (8.3) 60.9 22.8 (3.0) 62.5 84.4 31.3 9.5 87.5 33.5 (24.1) 13.4 (9.5) 3.21 (2.05) 312.5 (277.3–376.0) 2.01 (1.64–2.42) 28.9 (13.2–41.8) 69.4 (15.3) 0.4 (0.2–0.7) 63.3 (3.4) 11.2 (3.4) 53.1 62.5 51.6 67.1 9.4 4.7
68.2 (10.3) 56.3 24.4 (3.8) 62.5 85.2 34.4 10.9 87.5 26.4 (15.9) 16.8 (10.1) 2.05 (1.62) 384.0 (348.5–426.0) 1.70 (1.55–1.88) 75.4 (30.3–123.2) 58.9 (21.0) 0.8 (0.4–1.5) 61.3 (7.1) 16.7 (4.7) 75.0 69.8 56.3 68.8 18.8 15.6
0.92 0.59 0.01 1.0 0.80 0.71 0.79 1.0 0.051 0.047 b0.001 b0.001 b0.001 0.008 0.002 0.006 0.047 b0.001 0.01 0.38 0.60 0.85 0.13 0.04
Data are mean (standard deviation), median (25th to 75th percentile range), or number (percentage). HF: heart failure, HFpEF: heart failure with preserved left ventricular ejection fraction, BMI: body mass index, CAD: coronary artery disease, DM: diabetes mellitus, BH4: tetrahydrobiopterin, BH2: 7,8-dihydrobiopterin, DROM: derivatives of reactive oxidative metabolites, RHI: reactive hyperemia peripheral arterial tonometry index, BNP: B-type natriuretic peptide, eGFR: estimated glomerular filtration rate, hs-CRP: high-sensitivity C-reactive protein, LVEF: left ventricular ejection fraction, E/e′: the ratio of early transmitral flow velocity to tissue doppler early diastolic mitral annular velocity, ACEI: angiotensin-converting enzyme inhibitors, ARB: angiotensin II receptor blockers, CCB: calcium channel blockers, HMG-CoA RI: hydroxymethylglutaryl coenzyme A reductase inhibitors, MR: mineralocorticoid receptor. a Compared between risk factor-matched non-HF patients and risk factor-matched HFpEF patients.
induced cardiovascular diseases. Actually, we recently demonstrated the clinical significance of DROM in HFpEF patients [8]. In the present study, we also investigated one other oxidative marker, urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG) levels, and found that they
in vascular endothelial dysfunction and cardiac diastolic dysfunction in HFpEF. Several studies reported that ROS were closely associated with the pathophysiology of vascular endothelial dysfunction and its-
(B)
r=-0.181 P=0.04
700 600
1.4
r=0.233 P=0.009
1.2 1
500
Ln-RHI
DROM (U.CARR)
(A)
400 300 200
0.8 0.6 0.4 0.2
100 0
0 0
2
4
6
8
10
0
2
(C)
6
8
10
40
r=-0.26 P=0.003
30
E/e’
4
BH4/BH2
BH4/BH2
20
10
0 0
2
4
6
8
10
BH4/BH2 Fig. 1. Correlations of the plasma tetrahydrobiopterin (BH4)/7,8-dihydrobiopterin (BH2) ratios with serum DROM levels (A), ln-reactive hyperemia peripheral arterial tonometry index (RHI) (B), and the ratio of early transmitral flow velocity to tissue doppler early diastolic mitral annular velocity (E/e′) (C), respectively.
E. Yamamoto et al. / International Journal of Cardiology 190 (2015) 335–337
were not significantly correlated with RHI (data not shown). Therefore, it is possible that plasma BH4/BH2 ratio and serum DROM levels could serve as more sensitive and specific markers of ROS in HFpEF than commonly used conventional ROS markers such as urinary 8-OHdG. In this study, we further examined plasma pteridine levels and peripheral endothelial function in HFpEF patients, and clearly showed that endothelial function was correlated with plasma pteridine levels. When BH4 is relatively deficient by downregulating its synthesis or upregulating its oxidation, eNOS becomes uncoupled, leading to ROS overproduction. Therefore, measurements of BH4/BH2 ratio and oxidative status might be useful biomarkers for estimating pteridine metabolism disorder and its-induced eNOS uncoupling. In this study, plasma BH4 levels decreased, BH2 levels increased and subsequently the BH4/BH2 ratio decreased in association with endothelial dysfunction. As well as a few previous reports which showed that plasma BH4/BH2 ratio was associated with endothelial dysfunction in various cardiovascular diseases [9], we firstly demonstrated that BH4/BH2 ratio was closely associated with endothelial dysfunction in HFpEF. ROS have been reported to be one of major risk factors for the development of HFpEF [10], and a variety of enzymes including NADPH oxidase, xanthine oxidase and uncoupled eNOS produce ROS in cardiovascular tissues. However, the relative relationship among these enzymes regarding the role in vascular injury in HFpEF is not fully elucidated. In previous basic research, we reported that angiotensin II-induced eNOS uncoupling, rather than NADPH oxidase activation, contributed to vascular injury in DS rats [4]. The present study clinically demonstrated that vascular eNOS uncoupling, suggested by the relative decrease in BH4/BH2 ratio and the inverse increase in DROM values, significantly occurred in HFpEF patients. Patients with HFpEF have a poor prognosis equivalent to patients with HF with reduced LVEF. Therefore, identification of effective therapeutic strategy for HFpEF has great clinical importance. Because the significance of ROS overproduction caused by eNOS uncoupling in HFpEF was demonstrated by clinical and basic reports including ours, any interventions for vascular eNOS uncoupling could be a novel therapeutic strategy for HFpEF. Hence, further clinical studies conducted with a larger population are required to elucidate whether the inhibition of eNOS uncoupling by BH4 supplementation can improve endothelial dysfunction and subsequent occurrence of HFpEF. In conclusion, the present study firstly suggested that eNOS uncoupling, indicated by relative decrease in BH4/BH2 ratio, was associated with endothelial dysfunction in HFpEF patients. Conflict of interest statement Dr. Ogawa received lecture fees and research grants from Astellas, AstraZeneca, Bayer, Boehringer Ingelheim, Chugai, Daiichi
337
Sankyo, Dainippon Sumitomo Pharma, Eisai, Kowa, Kyowa Hakko Kirin, Mitsubishi Tanabe, MSD, Novartis, Otsuka, Pfizer, Sanofi, Shionogi, Takeda, and Mochida. Funding/support This work was supported in part by Grants-in Aid for Scientific Research (grant number: B24790770 to E. Yamamoto) from the Japanese Ministry of Education, Culture, Sports, Science, and Technology, Suzuken Memorial Foundation (to E. Yamamoto), Japan Research Foundation for Clinical Pharmacology (to E. Yamamoto), Salt Science Research Foundation (no. 1237 to E. Yamamoto), Takeda Science Foundation (to E. Yamamoto) and Japan Cardiovascular Research Foundation (to H. Ogawa). References [1] V.J. Dzau, E.M. Antman, H.R. Black, D.L. Hayes, J.E. Manson, J. Plutzky, J.J. Popma, W. Stevenson, The cardiovascular disease continuum validated: clinical evidence of improved patient outcomes: part I: pathophysiology and clinical trial evidence (risk factors through stable coronary artery disease), Circulation 114 (25) (2006) 2850–2870. [2] R.S. Vasan, M.G. Larson, E.J. Benjamin, J.C. Evans, C.K. Reiss, D. Levy, Congestive heart failure in subjects with normal versus reduced left ventricular ejection fraction: prevalence and mortality in a population-based cohort, J. Am. Coll. Cardiol. 33 (1999) 1948–1955. [3] E. Akiyama, S. Sugiyama, Y. Matsuzawa, et al., Incremental prognostic significance of peripheral endothelial dysfunction in patients with heart failure with normal left ventricular ejection fraction, J. Am. Coll. Cardiol. 60 (2012) 1778–1786. [4] E. Yamamoto, K. Kataoka, H. Shintaku, et al., Novel mechanism and role of angiotensin II induced vascular endothelial injury in hypertensive diastolic heart failure, Arterioscler. Thromb. Vasc. Biol. 27 (2007) 2569–2575. [5] W.J. Paulus, C. Tschöpe, J.E. Sanderson, et al., How to diagnose diastolic heart failure: a consensus statement on the diagnosis of heart failure with normal left ventricular ejection fraction by the Heart Failure and Echocardiography Associations of the European Society of Cardiology, Eur. Heart J. 28 (2007) 2539–2550. [6] H. Shintaku, Disorders of tetrahydrobiopterin metabolism and their treatment, Curr. Drug Metab. 3 (2002) 123–131. [8] Y. Hirata, E. Yamamoto, T. Tokitsu, et al., Reactive oxidative metabolites are associated with the severity of heart failure and predict future cardiovascular events in heart failure with preserved left ventricular ejection fraction, Int. J. Cardiol. 179 (2015) 305–308. [9] M. Takeda, T. Yamashita, M. Shinohara, N. Sasaki, T. Takaya, K. Nakajima, et al., Plasma tetrahydrobiopterin/dihydrobiopterin ratio: a possible marker of endothelial dysfunction, Circ. J. 73 (5) (2009) 955–962. [10] W.J. Paulus, C. Tschöpe, A novel paradigm for heart failure with preserved ejection fraction: comorbidities drive myocardial dysfunction and remodeling through coronary microvascular endothelial inflammation, J. Am. Coll. Cardiol. 62 (2013) 263–271.
Contents lists available at ScienceDirect
International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard
Letter to the Editor
The pivotal role of eNOS uncoupling in vascular endothelial dysfunction in patients with heart failure with preserved ejection fraction Eiichiro Yamamoto a,⁎, Yoshihiro Hirata a, Takanori Tokitsu a, Hiroaki Kusaka a, Kenji Sakamoto a, Megumi Yamamuro a, Koichi Kaikita a, Hiroshi Watanabe b, Seiji Hokimoto a, Seigo Sugiyama a,c, Toru Maruyama b, Hisao Ogawa a a b c
Faculty of Life Sciences, Department of Cardiovascular Medicine, Graduate School of Medical Sciences, Kumamoto University, Kumamoto, Japan Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Kumamoto University, Kumamoto, Japan Cardiovascular Division Diabetes Care Center, Jinnouchi Hospital, Kumamoto, Japan
a r t i c l e
i n f o
Article history: Received 9 February 2015 Accepted 18 April 2015 Available online 21 April 2015 Keywords: Heart failure with preserved left ventricular function Vascular endothelial dysfunction eNOS uncoupling
Vascular endothelial dysfunction correlates with cardiovascular diseases and plays an important role in all stages of atherosclerosis, leading to various cardiovascular diseases [1]. Furthermore, heart failure (HF) with preserved left ventricular ejection fraction (LVEF) (HFpEF) is one of the cardiovascular diseases of which the optimal treatment has not been established [2], because the precise pathophysiological mechanism underlying HFpEF still remains unclear. Recently, we reported that peripheral endothelial function, assessed by fingertrip digital reactive hyperemia peripheral arterial tonometry (RH-PAT), is significantly impaired in HFpEF patients than in non-HF patients [3]. Furthermore, using Dahl salt-sensitive hypertensive rats (DS rats), which are useful experimental models of HFpEF, we have previously elucidated molecular mechanism underlying endothelial dysfunction in HFpEF. We also demonstrated the association of endothelial nitric oxide synthase (eNOS) uncoupling, one of the sources of reactive oxygen species (ROS), with the progression of endothelial dysfunction-induced HFpEF [4]. ROS generally oxidizes tetrahydrobiopterin (BH4), an essential cofactor for eNOS, to its oxidative form 7,8-dihydrobiopterin (BH2) in the endothelial cells, leading to eNOS uncoupling which is at least partly involved in cardiovascular diseases. To our knowledge, however, there is no clinical information on correlations between endothelial ⁎ Corresponding author at: Department of Cardiovascular Medicine, Faculty of Life Sciences, Graduate School of Medical Science, Kumamoto University, 1-1-1 Honjo, Chuo-ku, Kumamoto 860-8556, Japan. E-mail address: [email protected] (E. Yamamoto).
http://dx.doi.org/10.1016/j.ijcard.2015.04.162 0167-5273/© 2015 Elsevier Ireland Ltd. All rights reserved.
dysfunction and pteridine such as BH4 and BH2 in HFpEF patients. In this study, hence, we examined the significance of plasma BH4 and BH2 in endothelial dysfunction in HFpEF. We recruited consecutive HFpEF patients hospitalized in Kumamoto University Hospital, and examined peripheral endothelial function by RH-PAT using the Endo-PAT2000 (Itamar Medical, Caesarea, Israel), and biomarkers (plasma pteridine and serum derivatives of reactive oxidative metabolites [DROM, Diacron srl, Grosseto, Italy]; normal range: 250–300 unit called the Carratelli unit [U.CARR], a new biomarker of ROS) in blood vein at stable condition after the optimal therapy. We defined HFpEF clinically according to the criteria of the European Working Group to HFpEF [5]: 1) symptoms of HF; 2) normal or mildly reduced LV systolic function (LVEF N 50% and LV end-diastolic volume index b 97 ml/m2); and 3) evidence of abnormal LV diastolic distensibility. Plasma BH4 and BH2 levels were measured directly by highperformance liquid chromatography (HPLC) with the electrochemical detection method, as previously described [6]. Baseline characteristics of patients are shown in Table 1. Compared to risk factor (number of patients, age, sex, and equal incidence of hypertension, diabetes mellitus, dyslipidemia and coronary artery disease)-matched non-HF patients, peripheral endothelial dysfunction, indicated by reduced RHI values, significantly occurred in risk factormatched HFpEF patients (2.01 [1.64–2.42] vs. 1.70 [1.55–1.88], P b 0.001), consistent of our previous clinical report [3]. In association with a reduction in RHI, plasma BH4 levels tended to decrease (33.4 ± 24.1 nM vs. 26.3 ± 15.9 nM, P = 0.051), BH2 levels significantly increased (13.4 ± 9.5 nM vs. 16.8 ± 10.1 nM, P = 0.047) and subsequently the BH4/BH2 ratio greatly decreased (3.21 ± 2.05 vs. 2.05 ± 1.62, P b 0.001) in HFpEF patients. Furthermore, serum DROM levels were significantly increased in HFpEF patients than in non-HF patients (312.5 [294.8–396.3] U.CARR vs. 384.0 [348.5–426.0] U.CARR, P b 0.001), accompanied by inverse reduction in BH4/BH2 ratio. Moreover, plasma BH4/BH2 ratios had significant and negative correlation with DROM values (P = 0.04, r = − 0.181, Fig. 1A) and the ratio of early-transmitral flow velocity to tissue doppler early-diastolic mitral annular velocity (E/e′) (P = 0.003, r = −0.26, Fig. 1C), and strong positive correlation with ln-RHI (P = 0.009, r = 0.233, Fig. 1B) in all enrolled patients. These data indicated that pteridine metabolism disorder induced-ROS/NO imbalance (eNOS uncoupling) could be involved
336
E. Yamamoto et al. / International Journal of Cardiology 190 (2015) 335–337
Table 1 Baseline characteristics of 64 risk factor-matched non-HF patients and 64 risk factor-matched HFpEF patients.
Age (years) Sex (male, %) BMI (kg/m2) CAD (yes, %) Hypertension (yes, %) DM (yes, %) Current smoking (yes, %) Dyslipidemia (yes, %) BH4 (nM) BH2 (nM) BH4/BH2 DROM (U.CARR) RHI BNP (pg/ml) eGFR (ml/min/1.73 m2) Hs-CRP (mg/l) LVEF (%) E/e′ β-Blockers (%) ACEI or ARB (%) CCB (%) HMG-CoA RI (%) MR blockers (%) Loop diuretics (%)
All patients (n = 128)
Matched non-HF (n = 64)
Matched HFpEF (n = 64)
P valuea
68.1 (9.3) 58.6 23.6 (3.5) 62.5 85.2 32.8 10.2 87.5 29.9 (20.7) 15.1 (9.9) 2.63 (1.93) 362.3 (294.8–396.3) 1.8 (1.62–2.15) 37.5 (19.8–80.9) 64.2 (19.0) 0.6 (0.3–0.9) 62.3 (5.7) 14.0 (4.9) 64.1 66.1 53.9 68.0 14.1 10.2
68.0 (8.3) 60.9 22.8 (3.0) 62.5 84.4 31.3 9.5 87.5 33.5 (24.1) 13.4 (9.5) 3.21 (2.05) 312.5 (277.3–376.0) 2.01 (1.64–2.42) 28.9 (13.2–41.8) 69.4 (15.3) 0.4 (0.2–0.7) 63.3 (3.4) 11.2 (3.4) 53.1 62.5 51.6 67.1 9.4 4.7
68.2 (10.3) 56.3 24.4 (3.8) 62.5 85.2 34.4 10.9 87.5 26.4 (15.9) 16.8 (10.1) 2.05 (1.62) 384.0 (348.5–426.0) 1.70 (1.55–1.88) 75.4 (30.3–123.2) 58.9 (21.0) 0.8 (0.4–1.5) 61.3 (7.1) 16.7 (4.7) 75.0 69.8 56.3 68.8 18.8 15.6
0.92 0.59 0.01 1.0 0.80 0.71 0.79 1.0 0.051 0.047 b0.001 b0.001 b0.001 0.008 0.002 0.006 0.047 b0.001 0.01 0.38 0.60 0.85 0.13 0.04
Data are mean (standard deviation), median (25th to 75th percentile range), or number (percentage). HF: heart failure, HFpEF: heart failure with preserved left ventricular ejection fraction, BMI: body mass index, CAD: coronary artery disease, DM: diabetes mellitus, BH4: tetrahydrobiopterin, BH2: 7,8-dihydrobiopterin, DROM: derivatives of reactive oxidative metabolites, RHI: reactive hyperemia peripheral arterial tonometry index, BNP: B-type natriuretic peptide, eGFR: estimated glomerular filtration rate, hs-CRP: high-sensitivity C-reactive protein, LVEF: left ventricular ejection fraction, E/e′: the ratio of early transmitral flow velocity to tissue doppler early diastolic mitral annular velocity, ACEI: angiotensin-converting enzyme inhibitors, ARB: angiotensin II receptor blockers, CCB: calcium channel blockers, HMG-CoA RI: hydroxymethylglutaryl coenzyme A reductase inhibitors, MR: mineralocorticoid receptor. a Compared between risk factor-matched non-HF patients and risk factor-matched HFpEF patients.
induced cardiovascular diseases. Actually, we recently demonstrated the clinical significance of DROM in HFpEF patients [8]. In the present study, we also investigated one other oxidative marker, urinary 8-hydroxy-2′-deoxyguanosine (8-OHdG) levels, and found that they
in vascular endothelial dysfunction and cardiac diastolic dysfunction in HFpEF. Several studies reported that ROS were closely associated with the pathophysiology of vascular endothelial dysfunction and its-
(B)
r=-0.181 P=0.04
700 600
1.4
r=0.233 P=0.009
1.2 1
500
Ln-RHI
DROM (U.CARR)
(A)
400 300 200
0.8 0.6 0.4 0.2
100 0
0 0
2
4
6
8
10
0
2
(C)
6
8
10
40
r=-0.26 P=0.003
30
E/e’
4
BH4/BH2
BH4/BH2
20
10
0 0
2
4
6
8
10
BH4/BH2 Fig. 1. Correlations of the plasma tetrahydrobiopterin (BH4)/7,8-dihydrobiopterin (BH2) ratios with serum DROM levels (A), ln-reactive hyperemia peripheral arterial tonometry index (RHI) (B), and the ratio of early transmitral flow velocity to tissue doppler early diastolic mitral annular velocity (E/e′) (C), respectively.
E. Yamamoto et al. / International Journal of Cardiology 190 (2015) 335–337
were not significantly correlated with RHI (data not shown). Therefore, it is possible that plasma BH4/BH2 ratio and serum DROM levels could serve as more sensitive and specific markers of ROS in HFpEF than commonly used conventional ROS markers such as urinary 8-OHdG. In this study, we further examined plasma pteridine levels and peripheral endothelial function in HFpEF patients, and clearly showed that endothelial function was correlated with plasma pteridine levels. When BH4 is relatively deficient by downregulating its synthesis or upregulating its oxidation, eNOS becomes uncoupled, leading to ROS overproduction. Therefore, measurements of BH4/BH2 ratio and oxidative status might be useful biomarkers for estimating pteridine metabolism disorder and its-induced eNOS uncoupling. In this study, plasma BH4 levels decreased, BH2 levels increased and subsequently the BH4/BH2 ratio decreased in association with endothelial dysfunction. As well as a few previous reports which showed that plasma BH4/BH2 ratio was associated with endothelial dysfunction in various cardiovascular diseases [9], we firstly demonstrated that BH4/BH2 ratio was closely associated with endothelial dysfunction in HFpEF. ROS have been reported to be one of major risk factors for the development of HFpEF [10], and a variety of enzymes including NADPH oxidase, xanthine oxidase and uncoupled eNOS produce ROS in cardiovascular tissues. However, the relative relationship among these enzymes regarding the role in vascular injury in HFpEF is not fully elucidated. In previous basic research, we reported that angiotensin II-induced eNOS uncoupling, rather than NADPH oxidase activation, contributed to vascular injury in DS rats [4]. The present study clinically demonstrated that vascular eNOS uncoupling, suggested by the relative decrease in BH4/BH2 ratio and the inverse increase in DROM values, significantly occurred in HFpEF patients. Patients with HFpEF have a poor prognosis equivalent to patients with HF with reduced LVEF. Therefore, identification of effective therapeutic strategy for HFpEF has great clinical importance. Because the significance of ROS overproduction caused by eNOS uncoupling in HFpEF was demonstrated by clinical and basic reports including ours, any interventions for vascular eNOS uncoupling could be a novel therapeutic strategy for HFpEF. Hence, further clinical studies conducted with a larger population are required to elucidate whether the inhibition of eNOS uncoupling by BH4 supplementation can improve endothelial dysfunction and subsequent occurrence of HFpEF. In conclusion, the present study firstly suggested that eNOS uncoupling, indicated by relative decrease in BH4/BH2 ratio, was associated with endothelial dysfunction in HFpEF patients. Conflict of interest statement Dr. Ogawa received lecture fees and research grants from Astellas, AstraZeneca, Bayer, Boehringer Ingelheim, Chugai, Daiichi
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Sankyo, Dainippon Sumitomo Pharma, Eisai, Kowa, Kyowa Hakko Kirin, Mitsubishi Tanabe, MSD, Novartis, Otsuka, Pfizer, Sanofi, Shionogi, Takeda, and Mochida. Funding/support This work was supported in part by Grants-in Aid for Scientific Research (grant number: B24790770 to E. Yamamoto) from the Japanese Ministry of Education, Culture, Sports, Science, and Technology, Suzuken Memorial Foundation (to E. Yamamoto), Japan Research Foundation for Clinical Pharmacology (to E. Yamamoto), Salt Science Research Foundation (no. 1237 to E. Yamamoto), Takeda Science Foundation (to E. Yamamoto) and Japan Cardiovascular Research Foundation (to H. Ogawa). References [1] V.J. Dzau, E.M. Antman, H.R. Black, D.L. Hayes, J.E. Manson, J. Plutzky, J.J. Popma, W. Stevenson, The cardiovascular disease continuum validated: clinical evidence of improved patient outcomes: part I: pathophysiology and clinical trial evidence (risk factors through stable coronary artery disease), Circulation 114 (25) (2006) 2850–2870. [2] R.S. Vasan, M.G. Larson, E.J. Benjamin, J.C. Evans, C.K. Reiss, D. Levy, Congestive heart failure in subjects with normal versus reduced left ventricular ejection fraction: prevalence and mortality in a population-based cohort, J. Am. Coll. Cardiol. 33 (1999) 1948–1955. [3] E. Akiyama, S. Sugiyama, Y. Matsuzawa, et al., Incremental prognostic significance of peripheral endothelial dysfunction in patients with heart failure with normal left ventricular ejection fraction, J. Am. Coll. Cardiol. 60 (2012) 1778–1786. [4] E. Yamamoto, K. Kataoka, H. Shintaku, et al., Novel mechanism and role of angiotensin II induced vascular endothelial injury in hypertensive diastolic heart failure, Arterioscler. Thromb. Vasc. Biol. 27 (2007) 2569–2575. [5] W.J. Paulus, C. Tschöpe, J.E. Sanderson, et al., How to diagnose diastolic heart failure: a consensus statement on the diagnosis of heart failure with normal left ventricular ejection fraction by the Heart Failure and Echocardiography Associations of the European Society of Cardiology, Eur. Heart J. 28 (2007) 2539–2550. [6] H. Shintaku, Disorders of tetrahydrobiopterin metabolism and their treatment, Curr. Drug Metab. 3 (2002) 123–131. [8] Y. Hirata, E. Yamamoto, T. Tokitsu, et al., Reactive oxidative metabolites are associated with the severity of heart failure and predict future cardiovascular events in heart failure with preserved left ventricular ejection fraction, Int. J. Cardiol. 179 (2015) 305–308. [9] M. Takeda, T. Yamashita, M. Shinohara, N. Sasaki, T. Takaya, K. Nakajima, et al., Plasma tetrahydrobiopterin/dihydrobiopterin ratio: a possible marker of endothelial dysfunction, Circ. J. 73 (5) (2009) 955–962. [10] W.J. Paulus, C. Tschöpe, A novel paradigm for heart failure with preserved ejection fraction: comorbidities drive myocardial dysfunction and remodeling through coronary microvascular endothelial inflammation, J. Am. Coll. Cardiol. 62 (2013) 263–271.