Emanuele Cotroneo

Histone Deacetylation Inhibition in Pulmonary Hypertension Therapeutic Potential of Valproic Acid and Suberoylanilide Hydroxamic Acid

Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension.Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension. # Clinical Perspective {#article-title-50}

Heterogeneity in Lung 18 FDG Uptake in Pulmonary Arterial Hypertension Potential of Dynamic 18 FDG Positron Emission Tomography With Kinetic Analysis as a Bridging Biomarker for Pulmonary Vascular Remodeling Targeted Treatments

Background— Pulmonary arterial hypertension (PAH) is a disease of progressive vascular remodeling, characterized by dysregulated growth of pulmonary vascular cells and inflammation. A prevailing view is that abnormal cellular metabolism, notably aerobic glycolysis that increases glucose demand, underlies the pathogenesis of PAH. Increased lung glucose uptake has been reported in animal models. Few data exist from patients with PAH. Methods and Results— Dynamic positron emission tomography imaging with fluorine-18–labeled 2-fluoro-2-deoxyglucose (18FDG) ligand with kinetic analysis demonstrated increased mean lung parenchymal uptake in 20 patients with PAH, 18 with idiopathic PAH (IPAH) (FDG score: 3.27±1.22), and 2 patients with connective tissue disease (5.07 and 7.11) compared with controls (2.02±0.71; P<0.05). Further compartment analysis confirmed increased lung glucose metabolism in IPAH. Lung 18FDG uptake and metabolism varied within the IPAH population and within the lungs of individual patients, consistent with the recognized heterogeneity of vascular pathology in this disease. The monocrotaline rat PAH model also showed increased lung 18FDG uptake, which was reduced along with improvements in vascular pathology after treatment with dicholoroacetate and 2 tyrosine kinase inhibitors, imatinib and sunitinib. Hyperproliferative pulmonary vascular fibroblasts isolated from IPAH patients exhibited upregulated glycolytic gene expression, along with increased cellular 18FDG uptake; both were reduced by dicholoroacetate and imatinib. Conclusions— Some patients with IPAH exhibit increased lung 18FDG uptake. 18FDG positron emission tomography imaging is a tool to investigate the molecular pathology of PAH and its response to treatment.

The zinc transporter ZIP12 regulates the pulmonary vascular response to chronic hypoxia

The typical response of the adult mammalian pulmonary circulation to a low oxygen environment is vasoconstriction and structural remodelling of pulmonary arterioles, leading to chronic elevation of pulmonary artery pressure (pulmonary hypertension) and right ventricular hypertrophy. Some mammals, however, exhibit genetic resistance to hypoxia-induced pulmonary hypertension. We used a congenic breeding program and comparative genomics to exploit this variation in the rat and identified the gene Slc39a12 as a major regulator of hypoxia-induced pulmonary vascular remodelling. Slc39a12 encodes the zinc transporter ZIP12. Here we report that ZIP12 expression is increased in many cell types, including endothelial, smooth muscle and interstitial cells, in the remodelled pulmonary arterioles of rats, cows and humans susceptible to hypoxia-induced pulmonary hypertension. We show that ZIP12 expression in pulmonary vascular smooth muscle cells is hypoxia dependent and that targeted inhibition of ZIP12 inhibits the rise in intracellular labile zinc in hypoxia-exposed pulmonary vascular smooth muscle cells and their proliferation in culture. We demonstrate that genetic disruption of ZIP12 expression attenuates the development of pulmonary hypertension in rats housed in a hypoxic atmosphere. This new and unexpected insight into the fundamental role of a zinc transporter in mammalian pulmonary vascular homeostasis suggests a new drug target for the pharmacological management of pulmonary hypertension.

Iron Homeostasis and Pulmonary Hypertension: Iron Deficiency Leads to Pulmonary Vascular Remodeling in the Rat

RATIONALE Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. OBJECTIVES We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. METHODS AND RESULTS Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1, lung fluorine-18-labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. CONCLUSIONS These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension.Rationale: Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. Objectives: We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. Methods and Results: Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1 , lung fluorine-18–labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. Conclusions: These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension. # Novelty and Significance {#article-title-52}

Iron Homeostasis and Pulmonary Hypertension

RATIONALE Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. OBJECTIVES We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. METHODS AND RESULTS Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1, lung fluorine-18-labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. CONCLUSIONS These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension.Rationale: Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. Objectives: We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. Methods and Results: Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1 , lung fluorine-18–labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. Conclusions: These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension. # Novelty and Significance {#article-title-52}

Histone Deacetylation Inhibition in Pulmonary Hypertension: Therapeutic Potential of Valproic Acid (VPA) and Suberoylanilide Hydroxamic Acid (SAHA)

Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension.Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension. # Clinical Perspective {#article-title-50}

Response to Letter Regarding Article, “Histone Deacetylation Inhibition in Pulmonary Hypertension: Therapeutic Potential of Valproic Acid and Suberoylanilide Hydroxamic Acid”

In their letter, Bogaard et al appreciate that HDAC inhibitors “may mitigate pulmonary vascular remodeling through effects on lung endothelial cells or vascular smooth muscle cells”1,2 but are concerned about the possibility that “HDAC inhibitors threaten the heart’s adaptive response to pressure overload.” These concerns, which were addressed in our articles,1,2 arise from their own study using trichostatin A, a broad-spectrum HDAC inhibitor, and valproic acid, which is not a broad-spectrum HDAC inhibitor, in a rat pulmonary artery banding model.3 In their study, trichostatin A (but not valproic acid) worsened right ventricular function and was associated with exaggerated right ventricular fibrosis and capillary rarefaction. Although these observations …

Iron Homeostasis and Pulmonary HypertensionNovelty and Significance

RATIONALE Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. OBJECTIVES We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. METHODS AND RESULTS Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1, lung fluorine-18-labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. CONCLUSIONS These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension.Rationale: Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. Objectives: We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. Methods and Results: Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1 , lung fluorine-18–labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. Conclusions: These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension. # Novelty and Significance {#article-title-52}

Iron Homeostasis and Pulmonary HypertensionNovelty and Significance: Iron Deficiency Leads to Pulmonary Vascular Remodeling in the Rat

RATIONALE Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. OBJECTIVES We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. METHODS AND RESULTS Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1, lung fluorine-18-labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. CONCLUSIONS These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension.Rationale: Iron deficiency without anemia is prevalent in patients with idiopathic pulmonary arterial hypertension and associated with reduced exercise capacity and survival. Objectives: We hypothesized that iron deficiency is involved in the pathogenesis of pulmonary hypertension and iron replacement is a possible therapeutic strategy. Methods and Results: Rats were fed an iron-deficient diet (IDD, 7 mg/kg) and investigated for 4 weeks. Iron deficiency was evident from depleted iron stores (decreased liver, serum iron, and ferritin), reduced erythropoiesis, and significantly decreased transferrin saturation and lung iron stores after 2 weeks IDD. IDD rats exhibited profound pulmonary vascular remodeling with prominent muscularization, medial hypertrophy, and perivascular inflammatory cell infiltration, associated with raised pulmonary artery pressure and right ventricular hypertrophy. IDD rat lungs demonstrated increased expression of hypoxia-induced factor-1α and hypoxia-induced factor-2α, nuclear factor of activated T cells and survivin, and signal transducers and activators of transcription-3 activation, which promote vascular cell proliferation and resistance to apoptosis. Biochemical examination showed reduced mitochondrial complex I activity and mitochondrial membrane hyperpolarization in mitochondria from IDD rat pulmonary arteries. Along with upregulation of the glucose transporter, glucose transporter 1, and glycolytic genes, hk1 and pdk1 , lung fluorine-18–labeled 2-fluoro-2-deoxyglucose ligand uptake was significantly increased in IDD rats. The hemodynamic and pulmonary vascular remodeling were reversed by iron replacement (ferric carboxymaltose, 75 mg/kg) and attenuated in the presence of iron deficiency by dichloroacetate and imatinib, 2 putative treatments explored for pulmonary arterial hypertension that target aerobic glycolysis and proliferation, respectively. Conclusions: These data suggest a major role for iron in pulmonary vascular homeostasis and support the clinical evaluation of iron replacement in patients with pulmonary hypertension. # Novelty and Significance {#article-title-52}

Histone Deacetylation Inhibition in Pulmonary HypertensionClinical Perspective: Therapeutic Potential of Valproic Acid and Suberoylanilide Hydroxamic Acid

Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension.Background— Epigenetic programming, dynamically regulated by histone acetylation, is a key mechanism regulating cell proliferation and survival. Little is known about the contribution of histone deacetylase (HDAC) activity to the development of pulmonary arterial hypertension, a condition characterized by profound structural remodeling of pulmonary arteries and arterioles. Methods and Results— HDAC1 and HDAC5 protein levels were elevated in lungs from human idiopathic pulmonary arterial hypertension and in lungs and right ventricles from rats exposed to hypoxia. Immunohistochemistry localized increased expression to remodeled vessels in the lung. Both valproic acid, a class I HDAC inhibitor, and suberoylanilide hydroxamic acid (vorinostat), an inhibitor of class I, II, and IV HDACs, mitigated the development of and reduced established hypoxia-induced pulmonary hypertension in the rat. Both valproic acid and suberoylanilide hydroxamic acid inhibited the imprinted highly proliferative phenotype of fibroblasts and R-cells from pulmonary hypertensive bovine vessels and platelet-derived growth factor–stimulated growth of human vascular smooth muscle cells in culture. Exposure to valproic acid and suberoylanilide hydroxamic acid was associated with increased levels of p21 and FOXO3 and reduced expression of survivin. The significantly higher levels of expression of cKIT, monocyte chemoattractant protein-1, interleukin-6, stromal-derived factor-1, platelet-derived growth factor-b, and S100A4 in R-cells were downregulated by valproic acid and suberoylanilide hydroxamic acid treatment. Conclusions— Increased HDAC activity contributes to the vascular pathology of pulmonary hypertension. The effectiveness of HDAC inhibitors, valproic acid, and suberoylanilide hydroxamic acid, in models of pulmonary arterial hypertension supports a therapeutic strategy based on HDAC inhibition in pulmonary arterial hypertension. # Clinical Perspective {#article-title-50}