No abstract

C ytokines/chemokines and growth factors regulate pulmonary endothelial function and influence the development of pulmonary arterial hypertension (PAH).1 PAH is characterized by pulmonary vascular remodeling and perivascular inflammation, leading to right ventricular (RV) failure and premature death. [2][3][4][5] Endothelial dysfunction is a crucial pathogenic status that triggers a variety of vascular disorders, such as PAH. 6,7 Endothelial dysfunction is also considered a key underlying feature in most forms of clinical and experimental PAH, which is enhanced by inflammatory cytokines/ chemokines and growth factors.1,8 Indeed, we experience rapid progression and worsening of PAH, especially when complicated with infectious diseases, such as pneumonia and catheter-related infection.8 Pulmonary endothelial dysfunction in patients with PAH enhances pulmonary vascular remodeling through impaired release of vasodilators, such as nitric oxide (NO) and prostacyclin. [9][10][11] AMP-activated protein kinase (AMPK) is a heterotrimeric complex consisting of a catalytic subunit α and 2 regulatory subunits β and γ, and it is expressed in various tissues and Molecular Medicine© 2016 American Heart Association, Inc. ). In contrast, development of hypoxia-induced PH was accelerated in eAMPK -/-mice compared with controls. Furthermore, the exacerbation of PH in eAMPK -/-mice was accompanied by reduced endothelial function, upregulation of growth factors, and increased proliferation of pulmonary artery smooth muscle cells. Importantly, conditioned medium from endothelial cells promoted pulmonary artery smooth muscle cell proliferation, which was further enhanced by the treatment with AMPK inhibitor. Serum levels of inflammatory cytokines, including tumor necrosis factor-α and interferon-γ were significantly increased in patients with PAH compared with healthy controls. Consistently, endothelial AMPK and cell proliferation were significantly reduced by the treatment with serum from patients with PAH compared with controls. Importantly, long-term treatment with metformin, an AMPK activator, significantly attenuated hypoxiainduced PH in mice. Conclusions:

Background: Right ventricular (RV) function is the major determinant for both functional capacity and survival in patients with pulmonary arterial hypertension (PAH). Despite the recognized clinical importance of preserving RV function, the subcellular mechanisms that govern the transition from a compensated to a decompensated state remain poorly understood and as a consequence there are no clinically established treatments for RV failure and a paucity of clinically useful biomarkers. Accumulating evidence indicates that long noncoding RNAs are powerful regulators of cardiac development and disease. Nonetheless, their implication in adverse RV remodeling in PAH is unknown. Methods: Expression of the long noncoding RNA H19 was assessed by quantitative PCR in plasma and RV from patients categorized as control RV, compensated RV or decompensated RV based on clinical history and cardiac index. The impact of H19 suppression using GapmeR was explored in 2 rat models mimicking RV failure, namely the monocrotaline and pulmonary artery banding. Echocardiographic, hemodynamic, histological, and biochemical analyses were conducted. In vitro gain- and loss-of-function experiments were performed in rat cardiomyocytes. Results: We demonstrated that H19 is upregulated in decompensated RV from PAH patients and correlates with RV hypertrophy and fibrosis. Similar findings were observed in monocrotaline and pulmonary artery banding rats. We found that silencing H19 limits pathological RV hypertrophy, fibrosis and capillary rarefaction, thus preserving RV function in monocrotaline and pulmonary artery banding rats without affecting pulmonary vascular remodeling. This cardioprotective effect was accompanied by E2F transcription factor 1-mediated upregulation of enhancer of zeste homolog 2. In vitro, knockdown of H19 suppressed cardiomyocyte hypertrophy induced by phenylephrine, while its overexpression has the opposite effect. Finally, we demonstrated that circulating H19 levels in plasma discriminate PAH patients from controls, correlate with RV function and predict long-term survival in 2 independent idiopathic PAH cohorts. Moreover, H19 levels delineate subgroups of patients with differentiated prognosis when combined with the NT-proBNP (N-terminal pro-B-type natriuretic peptide) levels or the risk score proposed by both REVEAL (Registry to Evaluate Early and Long-Term PAH Disease Management) and the 2015 European Pulmonary Hypertension Guidelines. Conclusions: Our findings identify H19 as a new therapeutic target to impede the development of maladaptive RV remodeling and a promising biomarker of PAH severity and prognosis.

These results indicate that SeP promotes the development of PH, suggesting that it is a novel biomarker and therapeutic target of the disorder.

Aims/hypothesis Nitric oxide (NO) is synthesised not only from L-arginine by NO synthases (NOSs), but also from its inert metabolites, nitrite and nitrate. Green leafy vegetables are abundant in nitrate, but whether or not a deficiency in dietary nitrite/nitrate spontaneously causes disease remains to be clarified. In this study, we tested our hypothesis that long-term dietary nitrite/nitrate deficiency would induce the metabolic syndrome in mice. Methods To this end, we prepared a low-nitrite/nitrate diet (LND) consisting of an amino acid-based low-nitrite/nitrate chow, in which the contents of L-arginine, fat, carbohydrates, protein and energy were identical with a regular chow, and potable ultrapure water. Nitrite and nitrate were undetectable in both the chow and the water. Results Three months of the LND did not affect food or water intake in wild-type C57BL/6J mice compared with a regular diet (RD). However, in comparison with the RD, 3 months of the LND significantly elicited visceral adiposity, dyslipidaemia and glucose intolerance. Eighteen months of the LND significantly provoked increased body weight, hypertension, insulin resistance and impaired endothelium-dependent relaxations to acetylcholine, while 22 months of the LND significantly led to death mainly due to cardiovascular disease, including acute Mika Kina-Tanada and Mayuko Sakanashi contributed equally to this work.

Although postcapillary pulmonary hypertension (PH) is an important prognostic factor for patients with heart failure (HF), its pathogenesis remains to be fully elucidated. To elucidate the different roles of Rho-kinase isoforms, ROCK1 and ROCK2, in cardiomyocytes in response to chronic pressure overload, we performed transverse aortic constriction (TAC) in cardiac-specific ROCK1-deficient () and ROCK2-deficient () mice. Cardiomyocyte-specific ROCK1 deficiency promoted pressure-overload-induced cardiac dysfunction and postcapillary PH, whereas cardiomyocyte-specific ROCK2 deficiency showed opposite results. Histological analysis showed that pressure-overload-induced cardiac hypertrophy and fibrosis were enhanced in mice compared with controls, whereas cardiac hypertrophy was attenuated in mice after TAC. Consistently, the levels of oxidative stress were up-regulated in hearts and down-regulated in hearts compared with controls after TAC. Furthermore, cyclophilin A (CyPA) and basigin (Bsg), both of which augment oxidative stress, enhanced cardiac dysfunction and postcapillary PH in mice, whereas their expressions were significantly lower in mice. In clinical studies, plasma levels of CyPA were significantly increased in HF patients and were higher in patients with postcapillary PH compared with those without it. Finally, high-throughput screening demonstrated that celastrol, an antioxidant and antiinflammatory agent, reduced the expressions of CyPA and Bsg in the heart and the lung, ameliorating cardiac dysfunction and postcapillary PH induced by TAC. Thus, by differentially affecting CyPA and Bsg expressions, ROCK1 protects and ROCK2 jeopardizes the heart from pressure-overload HF with postcapillary PH, for which celastrol may be a promising agent.

Rationale: Pulmonary arterial hypertension (PAH) is characterized by pulmonary vascular remodeling with aberrant pulmonary artery smooth muscle cells (PASMCs) proliferation, endothelial dysfunction, and extracellular matrix remodeling. Objective: Right ventricular (RV) failure is an important prognostic factor in PAH. Thus, we need to elucidate a novel therapeutic target in both PAH and RV failure. Methods and Results: We performed microarray analysis in PASMCs from patients with PAH (PAH-PASMCs) and controls. We found a ADAMTS8 (disintegrin and metalloproteinase with thrombospondin motifs 8), a secreted protein specifically expressed in the lung and the heart, was upregulated in PAH-PASMCs and the lung in hypoxia-induced pulmonary hypertension (PH) in mice. To elucidate the role of ADAMTS8 in PH, we used vascular smooth muscle cell-specific ADAMTS8-knockout mice (ADAMTS ΔSM22 ). Hypoxia-induced PH was attenuated in ADAMTS ΔSM22 mice compared with controls. ADAMTS8 overexpression increased PASMC proliferation with downregulation of AMPK (AMP-activated protein kinase). In contrast, deletion of ADAMTS8 reduced PASMC proliferation with AMPK upregulation. Moreover, deletion of ADAMTS8 reduced mitochondrial fragmentation under hypoxia in vivo and in vitro. Indeed, PASMCs harvested from ADAMTS ΔSM22 mice demonstrated that phosphorylated DRP-1 (dynamin-related protein 1) at Ser637 was significantly upregulated with higher expression of profusion genes (Mfn1 and Mfn2) and improved mitochondrial function. Moreover, recombinant ADAMTS8 induced endothelial dysfunction and matrix metalloproteinase activation in an autocrine/paracrine manner. Next, to elucidate the role of ADAMTS8 in RV function, we developed a cardiomyocyte-specific ADAMTS8 knockout mice (ADAMTS8 ΔαMHC ). ADAMTS8 ΔαMHC mice showed ameliorated RV failure in response to chronic hypoxia. In addition, ADAMTS8 ΔαMHC mice showed enhanced angiogenesis and reduced RV ischemia and fibrosis. Finally, high-throughput screening revealed that mebendazole, which is used for treatment of parasite infections, reduced ADAMTS8 expression and cell proliferation in PAH-PASMCs and ameliorated PH and RV failure in PH rodent models. Conclusions: These results indicate that ADAMTS8 is a novel therapeutic target in PAH.

Rationale: Pulmonary hypertension (PH) due to left heart disease (LHD), or group 2 PH, is the most prevalent form of PH worldwide. PH due to LHD is often associated with metabolic syndrome (MetS). In 12% to 13% of cases, patients with PH due to LHD display vascular remodeling of pulmonary arteries (PAs) associated with poor prognosis. Unfortunately, the underlying mechanisms remain unknown; PH-targeted therapies for this group are nonexistent, and the development of a new preclinical model is crucial. Among the numerous pathways dysregulated in MetS, inflammation plays also a critical role in both PH and vascular remodeling. Objective: We hypothesized that MetS and inflammation may trigger the development of vascular remodeling in group 2 PH. Methods and Results: Using supracoronary aortic banding, we induced diastolic dysfunction in rats. Then we induced MetS by a combination of high-fat diet and olanzapine treatment. We used metformin treatment and anti–IL-6 (interleukin-6) antibodies to inhibit the IL-6 pathway. Compared with sham conditions, only supracoronary aortic banding+MetS rats developed precapillary PH, as measured by both echocardiography and right/left heart catheterization. PH in supracoronary aortic banding+MetS was associated with macrophage accumulation and increased IL-6 production in lung. PH was also associated with STAT3 (signal transducer and activator of transcription 3) activation and increased proliferation of PA smooth muscle cells, which contributes to remodeling of distal PA. We reported macrophage accumulation, increased IL-6 levels, and STAT3 activation in the lung of group 2 PH patients. In vitro, IL-6 activates STAT3 and induces human PA smooth muscle cell proliferation. Metformin treatment decreased inflammation, IL-6 levels, STAT3 activation, and human PA smooth muscle cell proliferation. In vivo, in the supracoronary aortic banding+MetS animals, reducing IL-6, either by anti–IL-6 antibody or metformin treatment, reversed pulmonary vascular remodeling and improve PH due to LHD. Conclusions: We developed a new preclinical model of group 2 PH by combining MetS with LHD. We showed that MetS exacerbates group 2 PH. We provided evidence for the importance of the IL-6–STAT3 pathway in our experimental model of group 2 PH and human patients.