Histone deacetylase inhibitors synergize with sildenafil to suppress purine metabolism and proliferation in pulmonary hypertension.

RATIONALE: Sildenafil, a well-known vasodilator known to interfere with purinergic signaling through effects on cGMP, is a mainstay in the treatment of pulmonary hypertension (PH). However, little is known regarding its effects on the metabolic reprogramming of vascular cells, which is a hallmark of PH. Purine metabolism, especially intracellular de novo purine biosynthesis is essential for vascular cell proliferation. Since adventitial fibroblasts are critical contributors to proliferative vascular remodeling in PH, in this study we aimed to investigate if sildenafil, beyond its well-known vasodilator role in smooth muscle cells, impacts intracellular purine metabolism and proliferation of fibroblasts derived from human PH patients. METHODS: Integrated omics approaches (plasma and cell metabolomics) and pharmacological inhibitor approaches were employed in plasma samples and cultured pulmonary artery fibroblasts from PH patients. MEASUREMENTS AND MAIN RESULTS: Plasma metabolome analysis of 27 PH patients before and after treatment with sildenafil, demonstrated a partial, but specific effect of sildenafil on purine metabolites, especially adenosine, adenine, and xanthine. However, circulating markers of cell stress, including lactate, succinate, and hypoxanthine were only decreased in a small subset of sildenafil-treated patients. To better understand potential effects of sildenafil on pathological changes in purine metabolism (especially purine synthesis) in PH, we performed studies on pulmonary fibroblasts from PAH patients (PH-Fibs) and corresponding controls (CO-Fibs), since these cells have previously been shown to demonstrate stable and marked PH associated phenotypic and metabolic changes. We found that PH-Fibs exhibited significantly increased purine synthesis. Treatment of PH-Fibs with sildenafil was insufficient to normalize cellular metabolic phenotype and only modestly attenuated the proliferation. However, we observed that treatments which have been shown to normalize glycolysis and mitochondrial abnormalities including a PKM2 activator (TEPP-46), and the histone deacetylase inhibitors (HDACi), SAHA and Apicidin, had significant inhibitory effects on purine synthesis. Importantly, combined treatment with HDACi and sildenafil exhibited synergistic inhibitory effects on proliferation and metabolic reprogramming in PH-Fibs. CONCLUSIONS: While sildenafil alone partially rescues metabolic alterations associated with PH, treatment with HDACi, in combination with sildenafil, represent a promising and potentially more effective strategy for targeting vasoconstriction, metabolic derangement and pathological vascular remodeling in PH.

Right ventricular function and cardiopulmonary performance among patients with heart failure supported by durable mechanical circulatory support devices.

BACKGROUND: Patients with continuous-flow left ventricular assist devices (CF-LVADs) experience limitations in functional capacity and frequently, right ventricular (RV) dysfunction. We sought to characterize RV function in the context of global cardiopulmonary performance during exercise in this population. METHODS: A total of 26 patients with CF-LVAD (aged 58 ± 11 years, 23 males) completed a hemodynamic assessment with either conductance catheters (Group 1, n = 13) inserted into the right ventricle to generate RV pressure‒volume loops or traditional Swan‒Ganz catheters (Group 2, n = 13) during invasive cardiopulmonary exercise testing. Hemodynamics were collected at rest, 2 sub-maximal levels of exercise, and peak effort. Breath-by-breath gas exchange parameters were collected by indirect calorimetry. Group 1 participants also completed an invasive ramp test during supine rest to determine the impact of varying levels of CF-LVAD support on RV function. RESULTS: In Group 1, pump speed modulations minimally influenced RV function. During upright exercise, there were modest increases in RV contractility during sub-maximal exercise, but there were no appreciable increases at peak effort. Ventricular‒arterial coupling was preserved throughout the exercise. In Group 2, there were large increases in pulmonary arterial, left-sided filling, and right-sided filling pressures during sub-maximal and peak exercises. Among all participants, the cardiac output‒oxygen uptake relationship was preserved at 5.8:1. Ventilatory efficiency was severely abnormal at 42.3 ± 11.6. CONCLUSIONS: Patients with CF-LVAD suffer from limited RV contractile reserve; marked elevations in pulmonary, left-sided filling, and right-sided filling pressures during exercise; and severe ventilatory inefficiency. These findings explain mechanisms for persistent reductions in functional capacity in this patient population.

New insights into resting and exertional right ventricular performance in the healthy heart through real-time pressure-volume analysis.

KEY POINTS: Despite growing interest in right ventricular form and function in diseased states, there is a paucity of data regarding characteristics of right ventricular function - namely contractile and lusitropic reserve, as well as ventricular-arterial coupling, in the healthy heart during rest, as well as submaximal and peak exercise. Pressure-volume analysis of the right ventricle, during invasive cardiopulmonary exercise testing, demonstrates that that the right heart has enormous contractile reserve, with a three- or fourfold increase in all metrics of contractility, as well as myocardial energy production and utilization. The healthy right ventricle also demonstrates marked augmentation in lusitropy, indicating that diastolic filling of the right heart is not passive. Rather, the right ventricle actively contributes to venous return during exercise, along with the muscle pump. Ventricular-arterial coupling is preserved during submaximal and peak exercise in the healthy heart. ABSTRACT: Knowledge of right ventricular (RV) function has lagged behind that of the left ventricle and historically, the RV has even been referred to as a 'passive conduit' of lesser importance than its left-sided counterpart. Pressure-volume (PV) analysis is the gold standard metric of assessing ventricular performance. We recruited nine healthy sedentary individuals free of any cardiopulmonary disease (42 ± 12 years, 78 ± 11 kg), who completed invasive cardiopulmonary exercise testing during upright ergometry, while using conductance catheters inserted into the RV to generate real-time PV loops. Data were obtained at rest, two submaximal levels of exercise below ventilatory threshold, to simulate real-world scenarios/activities of daily living, and maximal effort. Breath-by-breath oxygen uptake was determined by indirect calorimetry. During submaximal and peak exercise, there were significant increases in all metrics of systolic function by three- to fourfold, including cardiac output, preload recruitable stroke work, and maximum rate of pressure change in the ventricle (dP/dtmax ), as well as energy utilization as determined by stroke work and pressure-volume area. Similarly, the RV demonstrated a significant, threefold increase in lusitropic reserve throughout exercise. Ventricular-arterial coupling, defined by the quotient of end-systolic elastance and effective arterial elastance, was preserved throughout all stages of exercise. Maximal pressures increased significantly during exercise, while end-diastolic volumes were essentially unchanged. Overall, these findings demonstrate that the healthy RV is not merely a passive conduit, but actively participates in cardiopulmonary performance during exercise by accessing an enormous amount of contractile and lusitropic reserve, ensuring that VA coupling is preserved throughout all stages of exercise.

Extensive macrophage accumulation in young and old Niemann-Pick C1 model mice involves the alternative, M2, activation pathway and inhibition of macrophage apoptosis.

We have studied the pathophysiology of lung disease which occurs in two mouse models of Niemann-Pick C1 disease. We utilized Npc1(-/-) mice transgenic for normal gene expression in glia or neurons and glia at ages several fold the usual and a mouse model of the juvenile form of NPC1, a point mutation, at one age to confirm some findings. Lung weights, as per cent of body weight, increase much more than liver and spleen weights. Although pulmonary function parameters only vary for hysteresis between young and older Npc1(-/-) mice, they are markedly different than those found in normal control mice. Cholesterol accumulation continued in the older mice but sphingosine-1-phosphate was not increased. Bronchoalveolar lavage (BAL) showed a massive increase (26×) in the number of macrophages. Histologic examination from the older, transgenic Npc1(-/-) mice showed small foci of alveolar proteinosis and evidence of hemorrhage, as well as dense macrophage accumulation. A large subset of macrophages was immunopositive for Fizz1 or arginase-1, markers of the alternative activation pathway, while no Fizz1 or arginase-1 positive macrophages were found in wild-type mice. The percentage of marker positive macrophages was relatively stable at 5-10% at various ages and within the 2 transgenic models. Phosphohistone H3 and Ki67 showed low levels of proliferation of these macrophages. Apoptosis was prominent within lung capillary endothelial cells, but limited within macrophages. Thus, activation of the alternative pathway is involved in Niemann-Pick C1 associated pulmonary macrophage accumulation, with low proliferation of these cells balanced by low levels of apoptosis.

Pulmonary function and pathology in hydroxypropyl-beta-cyclodextin-treated and untreated Npc1⁻/⁻ mice.

Lung dysfunction is an important part of the pathology of the neurodegenerative disorder, Niemann-Pick C1 (NPC1). We have studied the pulmonary disease in the Npc1(NIH/NIH) mouse model. On histology, we find large numbers of alveolar foamy macrophages but no alveolar proteinosis. Lung weight as percent of body weight was markedly increased; using the flexiVent small animal ventilator (SCIREQ, Inc.), we find inspiratory capacity, elastance and hysterisivity to be increased while resistance was not changed. Cholesterol measurements show a doubling of lung cholesterol levels. Collagen is also increased. Treatment of Npc1(-/-) mice with hydroxypropyl-ß-cyclodextrin (HPBCD), despite efficacious effects in brain and liver, results in little difference from age-matched controls (using a CNS-expressed transgene to extend the life expectancy of the Npc1(-/-) mice) for these variables.