Acute Effects of Rho-Kinase Inhibitor Fasudil on Pulmonary Arterial Hypertension in Patients With Congenital Heart Defects

Abstract

Background: We investigated the acute vasodilator effects of i.v. fasudil, a specific Rho-kinase inhibitor, on pulmonary circulation in patients with congenital heart defects (CHD) and severe pulmonary arterial hypertension (PAH).

Methods and Results: Thirty-five patients (34.23±12.10 years old) with CHD and severe PAH were consecutively enrolled. All patients underwent heart catheterization. At baseline and 30 min after initiation of i.v. fasudil, the following hemodynamic parameters were measured and calculated: right atrial pressure, pulmonary and systemic artery pressure (PAP and SAP), pulmonary and systemic vascular resistance, pulmonary-to-systemic blood pressure ratio (Pp/Ps), pulmonary-to-systemic blood flow ratio (Qp/Qs), cardiac index (CI) and artery oxygen saturation (SaO₂). After fasudil treatment, marked decrease in mean PAP (mPAP), pulmonary vascular resistance (PVR), total pulmonary resistance, pulmonary-to-systemic vascular resistance ratio (Rp/Rs) and mean Pp/Ps (mPp/Ps) was found, while Qp/Qs increased significantly without affecting CI and SAP. mPAP, PVR, Rp/Rs and Qp/Qs tended to be improved more significantly in the post-tricuspid shunt group compared with the pre-tricuspid shunt group.

Conclusions: Fasudil was well tolerated in patients with CHD and severe PAH, and significantly reduced PAP and PVR without affecting CI, SAP or SaO₂. (Circ J 2015; 79: 1342–1348)

Pulmonary arterial hypertension (PAH) is a rapidly progressive and fatal cardio-pulmonary vascular disease, characterized by pulmonary vascular endothelial dysfunction, hypercontraction, proliferation of smooth muscle cells, and migration of inflammatory cells. With the progressive elevation of pulmonary artery pressure (PAP) and vascular resistance, PAH leads to right heart failure and death.¹ Prognosis depends on disease subtype, sex, renal function, hemodynamics, World Health Organization functional class, exercise capacity, brain natriuretic peptide level, pericardial effusion on echocardiography, diffusion capacity for carbon monoxide, and vital parameters at diagnosis.^2,3 The severity of PAH in patients with congenital heart defects (CHD) is usually correlated with type and size of defect, hemodynamic characteristics, presence of extra-cardiac anomalies, and the status of the repair (unrepaired, palliated, or repaired).⁴ Theoretically, pre-tricuspid shunt, such as atrial septal defect (ASD), rarely develops to severe PAH, whereas the post-tricuspid shunts such as patent ductus arteriosus (PDA) and ventricular septal defect (VSD) are more likely to accompany severe PAH.

Editorial p 1213

In recent years, many studies have demonstrated that Rho-kinase is activated in animal models and patients with PAH, and is associated with enhanced pulmonary vascular constricting and proliferating responses, impaired endothelial function, and pulmonary vascular remodeling.^5,6 The Rho/Rho-kinase pathway has recently been found to play an important role in various cellular functions involved in the pathogenesis of a variety of cardiovascular diseases. Rho-kinase suppresses myosin phosphatase activity by phosphorylating the myosin-binding subunit of the enzyme and thus augments vascular smooth muscle cell (VSMC) contraction at a given intracellular calcium concentration. Moreover, Shimokawa demonstrated that Rho-kinase inhibition increases endothelial nitric oxide synthase (eNOS) expression and decreases inflammatory cell migration or anigiotensin II-induced mRNA expression of monocyte chemoattractant protein-1 and plasminogen activator inhibitor-1 in vivo and in vitro.⁷ This suggests that Rho-kinase may be involved in the major pathogenesis of PAH and may be a novel, and important therapeutic target of PAH in humans. Rho-kinase inhibitors are a promising new class of drugs for PAH. The present study investigated the acute hemodynamic effects of fasudil in patients with severe PAH and CHD.

Methods

Patients

The study was approved by the ethics committee of General Hospital of Shenyang Military Area Command, and written informed consent was obtained from all patients or guardians before cardiac catheterization and acute vasodilator tests. We consecutively enrolled 35 patients hospitalized with left-to-right shunt CHD due to severe PAH, who underwent cardiac catheterization between November 2012 and October 2013. All patients underwent cardiac evaluation, including a review of medical history, physical examination, electrocardiogram and echocardiogram before cardiac catheterization. Severe PAH was defined as mean PAP (mPAP) ≥46 mmHg, systolic PAP (sPAP) >two-thirds of the systemic level and pulmonary capillary wedge pressure (PCWP) <15 mmHg at catheterization at rest. The patients with idiopathic PAH or PAH due to connective tissue diseases, complex CHD, chronic thromboembolic pulmonary hypertension, heart failure (≥New York Heart Association class III), valvular heart disease or other system diseases and those who could not receive catheterization under local anesthesia were excluded. None of the patients received pulmonary vasodilators before cardiac catheterization, such as endothelin receptor antagonist, prostacyclin analogues, or phosphodiesterase type-5 inhibitors.

Cardiac Catheterization

All the patients underwent right heart catheterization, and acute vasodilator test under local anesthesia in the supine position, and breathing room air. A 6-F sheath was placed in the femoral vein, and a Swan-Ganz catheter (Edwards Life Sciences, Irvine, CA, USA) was advanced into the pulmonary artery under fluoroscopy. Transducers were positioned at the level of the anterior axillary line, and zeroed on atmospheric pressure. Hemodynamic parameters, including right atrial pressure (RAP), pulmonary and systemic artery pressure (PAP and SAP) and PCWP were recorded with a fluid-filled and catheter-connected pressure transducer. Mean RAP, mPAP, mean SAP and mean PCWP were then calculated. Blood samples were obtained from the pulmonary artery, right ventricle, right atrium, superior and inferior vena cava, and aorta during cardiac catheterization to detect oxygen saturation. Cardiac output, systemic vascular resistance (SVR), pulmonary vascular resistance (PVR) and total pulmonary resistance (TPR) were calculated from assumed oxygen consumption based on the Fick principle. Cardiac index (CI) was obtained by dividing left cardiac output by body surface area. Ratios of mean pulmonary to systemic pressure (mPp/Ps), systolic pulmonary to systemic pressure (sPp/Ps), pulmonary to systemic vascular resistance (Rp/Rs), and pulmonary to systemic blood flow (Qp/Qs) were calculated using the standard formulas from the Fick method.

Fasudil Treatment and Acute Vasodilator Test

After determination of baseline hemodynamic parameters, 60 mg fasudil hydrochloride (Chuanwei, Red Sun Pharmaceutical Company, Tianjin, China) was injected i.v. (dorsal superficial veins) using a portable infusion pump over 30 min.⁸ Hemodynamic parameters were measured and calculated again at 30 min after initiation of fasudil infusion. During fasudil treatment, SAP and heart rate were observed, as well as occurrence of symptoms due to adverse drug reaction such as headache, dyspnea, and dizziness. Based on the 2009 European Society of Cardiology/European Respiratory Society/International Society of Heart and Lung Transplantation guidelines on the management of PAH, all patients were divided into a pre-tricuspid shunt group or post-tricuspid shunts group according to anatomical-pathophysiological classification of congenital systemic to pulmonary shunts associated with PAH: (1) pre-tricuspid shunt group including ASD (n=21); and (2) post-tricuspid shunt group including VSD (n=8); PDA (n=5) and the combinations of ASD and VSD (n=1).

Statistical Analysis

Data are expressed as mean±SD and differences in hemodynamic parameters before and after intervention were assessed using paired Student t-test. Differences in hemodynamic parameters between the pre-tricuspid shunt group and the post-tricuspid shunt group were compared using unpaired Student t-test. Statistical analysis was performed with SPSS 13.0, and P<0.05 was considered statistically significant.

Results

Clinical Characteristics

A total of 35 patients (12 male, 23 female; age 34.93±12.87 years; range, 18–61 years; body weight, 53.80±7.56 kg; body height, 161.69±8.03 cm; body surface area, 1.53±0.14 m²) were respectively enrolled in the study. Baseline hemodynamics are listed in Table 1. There were no significant adverse reactions during fasudil treatment.

Table 1. Hemodynamic Data in Severe PAH-CHD (n=35)

Hemodynamic variables	Baseline	After fasudil infusion	P-value*
mPAP (mmHg)	65.37±13.55	57.28±13.97	<0.01
sPAP (mmHg)	104.51±17.87	89.77±16.04	<0.01
dPAP (mmHg)	46.09±13.14	40.34±13.27	<0.01
mRAP (mmHg)	6.91±2.32	4.87±2.53	<0.01
SAP (mmHg)	87.03±9.72	85.34±11.21	0.46
sPp/Ps	0.79±0.17	0.86±0.16	<0.01
mPp/Ps	0.75±0.18	0.70±0.18	<0.05
SaO2 (%)	91.29±4.17	91.42±4.80	0.75
PVR (Wood units)	10.66±5.93	8.36±5.02	<0.01
TPR (Wood units)	12.84±6.44	9.91±5.61	<0.01
CI (L·min−1·m−2)	3.03±0.75	3.07±0.76	0.68
Qp/Qs	1.20±0.33	1.45±0.58	<0.01
SVR (Wood units)	18.93±6.43	17.77±4.57	0.19
Rp/Rs	0.58±0.30	0.48±0.26	<0.01

Data given as mean±SD. *Paired t-test. CHD, congenital heart defect; CI, cardiac index; mPp/Ps, ratio of mean pulmonary to systemic pressure; mRAP, mean right atrial pressure; PAH, pulmonary arterial hypertension; PVR, pulmonary vascular resistance; Qp/Qs, ratio of pulmonary to systemic blood flow; Rp/Rs, ratio of pulmonary to systemic vascular resistance; SaO₂, oxygen saturation of femoral arterial blood; SAP, systemic arterial pressure; s/d/mPAP, systolic/diastolic/mean pulmonary artery pressure; sPp/Ps, ratio of systolic pulmonary to systemic pressure; SVR, systemic vascular resistance; TPR, total pulmonary resistance.

Hemodynamic Variables

After 30-min fasudil treatment, most of the subjects had marked decrease in mPAP, PVR, Rp/Rs and mPp/Ps. Decrease in mPAP, PVR, Rp/Rs and mPp/Ps were observed in 33 (94.3%), 30 (85.7%), 33 (94.3%) and 28 (80%) patients, respectively. Moreover, Qp/Qs increased in 28 patients (80%). Compared with baseline, both sPAP and mPAP decreased significantly: from 104.51±17.87 to 89.77±16.04 mmHg (P<0.01), and from 65.37±13.55 to 57.28±13.97 mmHg (P<0.01), respectively. Both PVR and TPR tended to reduce significantly: from 10.66±5.93 to 8.36±5.02 Wood units (P<0.01) and from 12.84±6.44 to 9.91±5.61 Wood units (P<0.01), respectively. mPp/Ps also decreased, from 0.75±0.18 to 0.70±0.18 (P<0.01), while Qp/Qs increased from 1.20±0.33 to 1.45±0.58 (P<0.05). And as shown in Figure 1, the changes of mPAP and PVR from baseline were not correlated with body weight or body surface area. No significant difference was observed in SAP, CI, SVR or SaO₂. The changes in hemodynamic parameters from baseline to 30 min after fasudil treatment, are listed in Table 1.

Figure 1.

Effects of 60-mg fixed dose fasudil vs. (A,B) body weight and (C,D) body surface area. (A,B) ΔMean pulmonary artery pressure (mPAP) and Δpulmonary vascular resistance (PVR) were not correlated with body weight (A, r=–0.017, P=0.923; B, r=0.136, P=0.437). (C,D) ΔmPAP and ΔPVR were not correlated with body surface area (C, r=–0.093, P=0.596; D, r=0.172, P=0.324).

All patients were divided into a pre-tricuspid shunt group (36.43±12.13 years old; 7 male, 14 female) or a post-tricuspid shunt group (30.93±11.69 years old; 5 male, 9 female). As shown in Table 2, mPAP, PVR, Rp/Rs and mPp/Ps at baseline were higher and SaO₂ at baseline was lower in the post-tricuspid shunt group than in the pre-tricuspid shunt group. There were significant statistical differences between the 2 groups, but not in age, gender, or Qp/Qs. As shown in Figure 2, no significant difference was found in CI, SaO₂, SAP or SVR between the 2 groups, and significantly decreased mPp/Ps, Qp/Qs, Rp/Rs and PVR were found only in the post-tricuspid shunt group. Only mPAP was significantly improved in both groups. As listed in Table 3, after fasudil treatment, mPAP, PVR, Rp/Rs and Qp/Qs tended to show a greater improvement in the post-tricuspid shunt group compared with the pre-tricuspid shunt group, although percent change from baseline was not significantly different between the 2 groups. Only percent change from baseline in SaO₂ was significantly improved in the post-tricuspid shunt group.

Table 2. Clinical Data and Baseline Hemodynamics vs. Shunt Type

	Pre-tricuspid group	Post-tricuspid group	P-value*
No. patients	21	14	–
F/M	7/14	5/9	0.885
Age (years)	36.43±12.13	30.93±11.69	0.192
SaO2 (%)	92.77±3.96	89.06±3.52	<0.01
CI (L·min−1·m−2)	3.09±0.71	2.94±0.83	0.56
mPAP (mmHg)	57.32±9.36	77.45±9.13	<0.01
PVR (Wood units)	8.43±4.24	14.02±6.65	<0.01
mPp/Ps	0.64±0.12	0.92±0.12	<0.01
Rp/Rs	0.47±0.23	0.74±0.34	<0.05
Qp/Qs	1.25±0.34	1.11±0.32	0.253

Data given as n or mean±SD. *Unpaired t-test. mPAP, mean pulmonary artery pressure. Other abbreviations as in Table 1.

Figure 2.

Comparison between patients with (––) pre-tricuspid and (- - -) post-tricuspid shunts with regard to changes in hemodynamic parameters after fasudil treatment, for cardiac index (CI), oxygen saturation of femoral arterial blood (SaO₂); systemic vascular resistance (SVR); systemic arterial pressure (SAP); ratio of mean pulmonary to systemic pressure (mPp/Ps); pulmonary vascular resistance (PVR); ratio of pulmonary to systemic blood flow (Qp/Qs); ratio of pulmonary to systemic vascular resistance (Rp/Rs); and mean pulmonary artery pressure (mPAP). NS, not significant.

Table 3. Change in Cardiac Hemodynamics After Fasudil Treatment

Parameters	Percent change from baseline		Between-group difference
	Pre-tricuspid shunt (n=21)	Post-tricuspid shunt (n=14)	Difference (95% confidence interval)	P-value*
ΔmPAP (%)	11.99±10.60	12.92±11.49	−0.93 (−8.62 to 6.77)	0.81
ΔmRAP (%)	30.48±25.67	24.76±40.77	5.71 (−17.08 to 28.51)	0.61
ΔPVR (%)	16.00±21.93	26.86±16.89	−10.85 (−24.96 to −3.25)	0.13
ΔTPR (%)	19.04±16.20	28.28±15.75	−9.24 (−20.49 to 2.01)	0.10
ΔRp/Rs (%)	10.14±27.57	21.90±22.99	−11.76 (−29.91 to 6.40)	0.20
ΔmPp/Ps (%)	5.88±15.23	7.98±11.55	−2.10 (−11.86 to 7.65)	0.66
ΔSaO2 (%)	0.89±1.90	−1.83±2.85	2.81 (1.18 to 4.44)	<0.01
ΔCI (%)	−7.78±23.84	1.50±15.99	−9.27 (−25.00 to 6.45)	0.24
ΔSAP (%)	2.50±6.24	3.90±9.12	−1.40 (−6.66 to 3.87)	0.59
ΔQp/Qs (%)	−15.5±40.2	−33.5±27.4	18.05 (−7.02 to 43.11)	0.15

Data given as mean±SD. *Unpaired t-test. Abbreviations as in Tables 1,2.

Discussion

Despite the advances in surgical repair, and the discovery of potential targeted therapies for PAH in the past 5 years, pulmonary hypertension still carries significant mortality and morbidity in patients with PAH-CHD. The anatomical, pathological, and structural abnormalities that exist in the pulmonary circulation of CHD are, to some extent, qualitatively similar to those observed in other forms of PAH. Beneficial effects on PAH-CHD with regard to the new classes of agents, however, are rarely reported; ideal drugs for the management of PAH should be highly selective in their actions on the pulmonary vascular bed, without affecting the systemic circulation.^9–11

The Rho/Rho-kinase pathway and Rho-kinase inhibitors have recently attracted much attention in the cardiovascular research fields, especially with regard to PAH. More and more evidence suggests that Rho/Rho kinase-mediated signaling pathways are involved in the pathogenesis of PAH. In experimental studies using animal models, Rho-kinase activity in the pulmonary arteries was enhanced, irrespective of the different etiologies.¹² Furthermore, Rho-kinase activation is observed in lung tissue, and in the pulmonary artery in rat models of chronic hypoxic, and monocrotaline (MCT)-induced pulmonary hypertension.^12–14 Other studies have also suggested that in addition to chronic thromboembolic pulmonary hypertension, Rho-kinase activity in circulating neutrophils and lung tissue was significantly enhanced in patients with PAH, especially idiopathic PAH. Furthermore, it is reported that the activity of Rho-kinase was significantly correlated with the severity of PAH, and exercise tolerance.^12,15,16 The Rho kinase pathway participates in vasoconstriction elicited by numerous agents involved in PAH, including 5-hydroxytryptamine (5-HT), endothelin-1 (ET-1) and thromboxane A2. Rho-kinase, an effector of the small GTP-binding protein Rho, suppresses myosin phosphatase activity by phosphorylating the myosin-binding subunit of the enzyme, thus augmenting VSMC contraction at a given intracellular calcium concentration.^17,18 Recent studies have demonstrated that the inhibition of Rho-kinase ameliorates endothelial dysfunction and suppresses the hypercontraction and proliferation of VSMC, and migration of inflammatory cells. Therefore it is observed that fasudil could decrease mPAP and PVR in many animal models of PAH, including MCT rat and chronic hypoxia rat.^12,18,19 In clinical studies, a Rho-kinase inhibitor, fasudil, acutely improved pulmonary hemodynamics in patients with PAH, which showed modest, immediate reductions in PVR in humans.^9,10 Fasudil, as a potent, and specific inhibitor of Rho-kinase, is the first approved agent for use in clinical practice. It exerts acute pulmonary vasodilative effects by inhibiting the phosphorylation of myosin light chain, which can directly affect excessive vasoconstriction, and by inhibiting proliferation of pulmonary vascular smooth muscle; it can disrupt the balance between endothelial contractive and dilative factors, and ameliorate pulmonary vascular remodeling, and inflammation.^5,13

There are, however, only a small number of studies on whether Rho-kinase is involved in the pathogenesis of PAH-CHD, and whether fasudil is useful in patients with PAH-CHD.^9–11 In the present study, we showed that i.v. treatment with fasudil, a Rho-kinase inhibitor, was beneficial in patients with severe PAH-CHD. Decreases in mPAP, PVR, TPR, Rp/Rs, and mPp/Ps were found in 94.3%, 85.7%, 94.3%, 82.9% and 74.3% of patients, respectively. Qp/Qs increased in 80%, suggesting that fasudil causes a significant reduction in mPAP, sPAP, PVR, mPp/Ps, Rp/Rs, and a marked increase in Qp/Qs, without changes in CI, SAP, SVR, and SaO₂. Moreover, the effect of the fixed dose fasudil on patients with PAH-CHD was not correlated with body weight or body surface area. Another advantage of fasudil is the pulmonary selectivity of fasudil’s vasodilatory effect, but the decrease in SAP was relatively small; and mPAP, PVR, and Rp/Rs tended to decrease, suggesting that fasudil seemed to have a less vasodilatory effect on systemic vasculature than on pulmonary vasculature in this study. The acute beneficial effect of Rho-kinase inhibitor on left-to-right shunt-induced PAH in this study, mainly its inhibitory effect on pulmonary vasoconstriction, could be ascribed to multiple mechanisms. The patients with CHD and PAH have unique hemodynamic characteristics. On the one hand, pulmonary vascular dilatation increases blood flow back to the left side of the heart. On the other hand, pulmonary vascular dilatation decreases PAP, which may increase the blood flow across the defects from left-to-right shunt and reduce cardiac output. Therefore, CI was not changed by fasudil in different types of CHD. These findings support the previous findings in animal models of PAH, and during right-heart cardiac catheterization in patients with PAH.^10,11,20 Thus, increased PVR may be caused, at least in part, by the activated Rho-kinase pathway.

Different from PAH associated with non-cardiac shunt, PAH-CHD has its own specific cardiac anatomy and pathophysiological basis. At earlier stages of PAH-CHD, pulmonary vascular lesions, which are mainly due to the tonic contraction of pulmonary vascular smooth muscle cells, are still reversible. At the final stage, the patient has low pulmonary blood flow, cyanosis with reverted shunt, and high PVR. Those patients will not benefit from surgical closure, which is even contraindicated, but they will potentially benefit from new targeted therapies for PAH.²¹ This situation should be avoided, and intervention conducted while pulmonary vascular reactivity is still present. Therefore, it is crucial to select drugs for patients with severe PAH and CHD on a patient-by-patient basis, to decrease PAP and PVR. This study determined whether acute inhibition of Rho-kinase would have beneficial effects on abnormal pulmonary hemodynamics in patients with CHD and PAH. In the present study, after i.v. fasudil treatment, patients with PAH-CHD had significantly increased acute pulmonary vasodilation, without significant changes in SAP and CI. The results indicate that Rho-kinase pathway is involved in the pathogenesis of PAH-CHD, and that fasudil is a novel and useful agent for managing patients with PAH-CHD. It is also consistent with a previous clinical study in which acute inhibition of Rho-kinase improved pulmonary hemodynamics in PAH patients. I.v. fasudil treatment rapidly blocks the Rho-kinase pathway to reduce inflammatory factors, including angiotensin, ET-1, and 5-HT. Blocking of those factors could weaken the sustained contraction of pulmonary vascular smooth muscle and induce increased expression and activity of the promoter of eNOS and prolong the half-life of eNOS mRNA, which inhibits pulmonary vascular hypercontraction and significantly improves hemodynamics parameters in patients with PAH-CHD.^22,23 Therefore, fasudil is an alternative drug for the treatment of PAH-CHD.

In the present study, we also compared the acute effects of the Rho-kinase inhibitor fasudil in PAH-CHD between patients with pre-tricuspid and post-tricuspid shunts. According to the conventional indicators of the severity of PAH (mPAP, PVR, Rp/Rs ratio, and mPp/Ps) at baseline, pulmonary vascular injury in the post-tricuspid shunt group was more severe. After fasudil treatment, however, the improvement in the post-tricuspid shunt group, according to CI, Qp/Qs, Rp/Rs and PVR, tended to be greater than in the pre-tricuspid shunt group; and SaO₂ percent change from baseline was significantly improved in the post-tricuspid shunt group compared with the pre-tricuspid shunt group. It was presumed that patients with post-tricuspid shunts have enhanced pulmonary vasoconstriction and reaction of pulmonary hypertension due to largely left-to-right flow. After fasudil treatment, right-to-left shunt reduced significantly in the post-tricuspid shunt group compared with the pre-tricuspid shunt group, and SaO₂ improved dramatically. Indeed, it is still unclear why the status of some patients with PAH-CHD progresses from reversible to irreversible. So far, most experts agree that a small cardiac shunt, especially a small ASD, is not a cause of PAH. ASD is very similar to idiopathic PAH. ASD and PAH may coexist or the small defect may initiate and activate a mechanism that causes sustained damage to the vascular endothelial system, ultimately causing irreversible pulmonary vascular remodeling.^5,24 I.v. fasudil may rapidly inhibit pulmonary vascular smooth muscle hypercontraction, while the reactivity of fasudil was poor with regard to the chronic changes of pulmonary vascular remodeling such as the proliferation and angiogenesis of neonatal VSMC and the muscularization of non-muscle vessels, leading to different reactivity for fasudil between the patients with pre-tricuspid and post-tricuspid shunt.

Given that the evolution of pulmonary vascular disease differs markedly between patients with pre-tricuspid and post-tricuspid shunts, the patients could respond differently to fasudil. This also prompts us to better define PAH-CHD, including the classification, diagnostic criteria, and treatment methods.

Study Limitations

The present study, however, had several limitations. Due to the differences in the types of CHD and the severity of PAH, as well as vasodilator reactivity, the safety of fasudil in terms of the systemic hemodynamics of severe PAH-CHD patients, and its long-term efficacy, should be further investigated in randomized controlled studies.

Conclusions

The present study suggested that i.v. fasudil, a Rho-kinase inhibitor, is effective and well tolerated in patients with PAH-CHD, and might be a novel therapeutic approach.

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