Hossein Ardeschir Ghofrani

Publications of Hossein Ardeschir Ghofrani

• Role of Src Tyrosine Kinases in Experimental Pulmonary Hypertension.

  Authors: Soni Savai Pullamsetti, Eva Maria Berghausen, Swati Dabral, Alexandra Tretyn, Elsa Butrous, Rajkumar Savai, Ghazwan Butrous, Bhola Kumar Dahal, Ralf P Brandes, Hossein Ardeschir Ghofrani, Norbert Weissmann, Friedrich Grimminger, Werner Seeger, Stephan Rosenkranz, Ralph Theo Schermuly

  Arteriosclerosis, thrombosis, and vascular biology. 04/2012;

  OBJECTIVE: Pulmonary arterial hypertension is a progressive pulmonary vascular disorder with high morbidity and mortality. Compelling evidence suggests that receptor tyrosine kinases, such asOBJECTIVE: Pulmonary arterial hypertension is a progressive pulmonary vascular disorder with high morbidity and mortality. Compelling evidence suggests that receptor tyrosine kinases, such as platelet-derived growth factor (PDGF) are closely involved in the pathogenesis of pulmonary arterial hypertension. We investigated the effects of 2 novel PDGF inhibitors, nilotinib/AMN107 (Abl kinases/PDGF receptor inhibitor) and dasatinib/BMS-354825 (Abl kinases/PDGF receptor/Src inhibitor), on the proliferation and migration of pulmonary arterial smooth muscle cells (PASMCs) and on the hemodynamics and pulmonary vascular remodeling in experimental pulmonary hypertension, and determined the expression and regulation of Src family kinases. METHODS AND RESULTS: Human PASMCs were stimulated by PDGF alone or multiple growth factors to induce proliferation and migration in vitro. Dasatinib (0.03 μmol/L), nilotinib (0.3 μmol/L), and imatinib (1 μmol/L) potently inhibited PDGF-induced Human PASMCs were stimulated by PDGF alone or multiple growth factors to induce proliferation and migration in vitro. Dasatinib (0.03 μmol/L), nilotinib (0.3 μmol/L), and imatinib (1 μmol/L) potently inhibited PDGF-induced signal transducer and activator of transcription 3 and Akt phosphorylation. All 3 inhibitors decreased PDGF-induced proliferation, cell cycle gene regulation, and migration. In contrast, only dasatinib inhibited multiple growth factor-induced PASMC proliferation, and this was associated with the inhibition of Src phosphorylation. Combination of specific Src inhibitors (phosphoprotein phosphatase 1, phosphoprotein phosphatase 2) with either imatinib or nilotinib reduced multiple growth factor-induced proliferation to a similar extent as dasatinib. Importantly, Src phosphorylation increased in pulmonary arterial hypertension PASMCs compared with control PASMCs. Finally, in vivo dasatinib (15 mg/kg per bw) treatment caused a complete reversal of pulmonary vascular remodeling and achieved similar effectiveness as imatinib (100 mg/kg per bw) in both monocrotaline- and hypoxia-induced pulmonary hypertension models. phosphatase 1, phosphoprotein phosphatase 2) with either imatinib or nilotinib reduced multiple growth factor-induced proliferation to a similar extent as dasatinib. Importantly, Src phosphorylation increased in pulmonary arterial hypertension PASMCs compared with control PASMCs. Finally, in vivo dasatinib (15 mg/kg per bw) treatment caused a complete reversal of pulmonary vascular remodeling and achieved similar effectiveness as imatinib (100 mg/kg per bw) in both monocrotaline- and hypoxia-induced pulmonary hypertension models. CONCLUSIONS: We suggest that dual inhibition of PDGF receptor and Src kinases potently inhibits mitogenic and motogenic responses to growth factors in PASMCs and pulmonary vascular remodeling in vivo so that dual inhibition may represent an alternative therapeutic approach for pulmonary arterial hypertension.

• The role of dimethylarginine dimethylaminohydrolase in idiopathic pulmonary fibrosis.

  Authors: Soni Savai Pullamsetti, Rajkumar Savai, Rio Dumitrascu, Bhola Kumar Dahal, Jochen Wilhelm, Melanie Konigshoff, Dariusz Zakrzewicz, Hossein Ardeschir Ghofrani, Norbert Weissmann, Oliver Eickelberg, Andreas Guenther, James Leiper, Werner Seeger, Friedrich Grimminger, Ralph Theo Schermuly

  Science translational medicine. 06/2011; 3(87):87ra53.

  Idiopathic pulmonary fibrosis (IPF) is a progressive, dysregulated response to alveolar injury that culminates in compromised lung function from excess extracellular matrix production. AssociatedIdiopathic pulmonary fibrosis (IPF) is a progressive, dysregulated response to alveolar injury that culminates in compromised lung function from excess extracellular matrix production. Associated with high morbidity and mortality, IPF is generally refractory to current pharmacological therapies. We examined fibrotic lungs from mice and from patients with IPF and detected increased expression of dimethylarginine dimethylaminohydrolases (DDAHs)--key enzymes that metabolize asymmetric dimethylarginine (ADMA), which is an endogenous inhibitor of nitric oxide synthase, to form l-citrulline and dimethylamine. DDAHs are up-regulated in primary alveolar epithelial type II cells from these mice and patients where they are colocalized with inducible nitric oxide synthase. In cultured alveolar epithelial type II cells from bleomycin-induced fibrotic mouse lungs, inhibition of DDAH suppressed proliferation and induced apoptosis in an ADMA-dependent manner. In addition, DDAH inhibition reduced collagen production by fibroblasts in an ADMA-independent but transforming growth factor/SMAD-dependent manner. In mice with bleomycin-induced pulmonary fibrosis, the DDAH inhibitor L-291 reduced collagen deposition and normalized lung function. In bleomycin-induced fibrosis, inducible nitric oxide synthase inhibition decreased fibrosis, but an even stronger reduction was observed after inhibition of DDAH. Thus, DDAH inhibition reduces fibroblast-induced collagen deposition in an ADMA-independent manner and reduces abnormal epithelial proliferation in an ADMA-dependent manner, offering a possible therapeutic avenue for attenuation of pulmonary fibrosis.

• cAMP phosphodiesterase inhibitors increases nitric oxide production by modulating dimethylarginine dimethylaminohydrolases.

  Authors: Soni Savai Pullamsetti, Rajkumar Savai, Martina Barbara Schaefer, Jochen Wilhelm, Hossein Ardeschir Ghofrani, Norbert Weissmann, Christian Schudt, Ingrid Fleming, Konstantin Mayer, James Leiper, Werner Seeger, Friedrich Grimminger, Ralph Theo Schermuly

  Circulation. 03/2011; 123(11):1194-204.

  Pulmonary arterial hypertension is characterized by a progressive increase in pulmonary vascular resistance caused by endothelial dysfunction, inward vascular remodeling, and severe loss ofPulmonary arterial hypertension is characterized by a progressive increase in pulmonary vascular resistance caused by endothelial dysfunction, inward vascular remodeling, and severe loss of precapillary pulmonary vessel cross-sectional area. Asymmetrical dimethylarginine (ADMA), an endogenous nitric oxide synthase inhibitor, and its metabolizing enzyme dimethylarginine dimethylaminohydrolase (DDAH) play important roles in endothelial dysfunction. We investigated whether combined phosphodiesterase (PDE) 3 and 4 inhibition ameliorates endothelial function by regulating the ADMA-DDAH axis. We investigated the effects of the PDE3/4 inhibitor tolafentrine in vitro on endothelial cell survival, proliferation, and apoptosis. Effects of tolafentrine on the endothelial nitric oxide synthase/nitric oxide pathway, DDAH expression, DDAH promoter activity, and cytokine release from endothelial cells and their subsequent influence on DDAH expression were investigated. In monocrotaline-induced pulmonary arterial hypertension in rats, the effects of inhaled tolafentrine on DDAH expression and activity were investigated. Real-time-polymerase chain reaction, immunocytochemistry, and PDE activity assays suggested high expression of PDE3 and PDE4 isoforms in endothelial cells. Treatment of endothelial cells with PDE3/4 inhibitor significantly decreased ADMA-induced apoptosis via a cAMP/PKA-dependent pathway by induction of DDAH2. Chronic nebulization of PDE3/4 inhibitor significantly attenuated monocrotaline-induced hemodynamic, gas exchange abnormalities, vascular remodeling, and right heart hypertrophy. Interestingly, PDE3/4 inhibitor treatment reduced ADMA and elevated nitric oxide/cGMP levels. Mechanistically, this could be attributed to direct modulatory effects of cAMP on the promoter region of DDAH2, which was consequently found to be increased in expression and activity. Furthermore, PDE3/4 inhibitor suppressed apoptosis in endothelial cells and increased vascularization in the lung. Combined inhibition of PDE3 and 4 regresses development of pulmonary hypertension and promotes endothelial regeneration by modulating the ADMA-DDAH axis.

• Riociguat for the treatment of pulmonary hypertension.

  Authors: Ralph T Schermuly, Wiebke Janssen, Norbert Weissmann, Johannes-Peter Stasch, Friedrich Grimminger, Hossein Ardeschir Ghofrani

  Expert opinion on investigational drugs. 03/2011; 20(4):567-76.

  INTRODUCTION: Pulmonary hypertension (PH) is a severe condition with a poor prognosis despite recent treatment advances. Therapies with new mechanisms of action are needed. AREAS COVERED: This reviewINTRODUCTION: Pulmonary hypertension (PH) is a severe condition with a poor prognosis despite recent treatment advances. Therapies with new mechanisms of action are needed. AREAS COVERED: This review will help readers understand the mechanism of action of the soluble guanylate cyclase (sGC) stimulator riociguat (BAY 63-2521) and will provide a comprehensive summary regarding efficacy and safety of this drug in the management of PH. The most relevant publications up to December 2010 were used as sources for this review. EXPERT OPINION: Cyclic guanosine monophosphate (cGMP) is an important mediator of the preferential perfusion of well-ventilated regions throughout the lung. Drugs that increase cGMP levels could promote pulmonary vasorelaxation while maintaining optimal gas exchange. cGMP is generated by sGC, which can be stimulated by nitric oxide (NO). Riociguat stimulates sGC independently of NO and increases the sensitivity of sGC to NO, resulting in increased cGMP levels. Results to date suggest rapid, potent and prolonged efficacy and good tolerability in different types of PH. Phase III clinical trials are evaluating the long-term safety and clinical effectiveness of riociguat in pulmonary arterial hypertension (PAH) and chronic thromboembolic PH. Riociguat has the potential to become an important drug for the treatment of patients with PH.

• Glycogen synthase kinase 3beta contributes to proliferation of arterial smooth muscle cells in pulmonary hypertension.

  Authors: Piotr Sklepkiewicz, Ralph Theo Schermuly, Xia Tian, Hossein Ardeschir Ghofrani, Norbert Weissmann, Daniel Sedding, Tarek Kashour, Werner Seeger, Friedrich Grimminger, Soni Savai Pullamsetti

  PloS one. 01/2011; 6(4):e18883.

  Pulmonary arterial hypertension (PAH) is a rare progressive pulmonary vascular disorder associated with vascular remodeling and right heart failure. Vascular remodeling involves numerous signalingPulmonary arterial hypertension (PAH) is a rare progressive pulmonary vascular disorder associated with vascular remodeling and right heart failure. Vascular remodeling involves numerous signaling cascades governing pulmonary arterial smooth muscle cell (PASMC) proliferation, migration and differentiation. Glycogen synthase kinase 3beta (GSK3ß) is a serine/threonine kinase and can act as a downstream regulatory switch for numerous signaling pathways. Hence, we hypothesized that GSK3ß plays a crucial role in pulmonary vascular remodeling. All experiments were done with lung tissue or isolated PASMCs in a well-established monocrotaline (MCT)-induced PAH rat model. The mRNA expression of Wnt ligands (Wnt1, Wnt3a, Wnt5a), upstream Wnt signaling regulator genes (Frizzled Receptors 1, 2 and secreted Frizzled related protein sFRP-1) and canonical Wnt intracellular effectors (GSK3ß, Axin1) were assessed by real-time polymerase chain reaction and protein levels of GSK3ß, phospho-GSK3ß (ser 9) by western blotting and localization by immunohistochemistry. The role of GSK3ß in PASMCs proliferation was assessed by overexpression of wild-type GSK3ß (WT) and constitutively active GSK3ß S9A by [(3)H]-thymidine incorporation assay. Increased levels of total and phosphorylated GSK3ß (inhibitory phosphorylation) were observed in lungs and PASMCs isolated from MCT-induced PAH rats compared to controls. Further, stimulation of MCT-PASMCs with growth factors induced GSK3ß inactivation. Most importantly, treatment with the PDGFR inhibitor, Imatinib, attenuated PDGF-BB and FCS induced GSK3ß phosphorylation. Increased expression of GSK3ß observed in lungs and PASMC isolated from MCT-induced PAH rats was confirmed to be clinically relevant as the same observation was identified in human iPAH lung explants. Overexpression of GSK3ß significantly increased MCT-PASMCs proliferation by regulating ERK phosphorylation. Constitutive activation of GSK3ß (GSK3ß S9A, 9th serine replaced to alanine) inhibited MCT-PASMCs proliferation by decreasing ERK phosphorylation. This study supports a central role for GSK3ß in vascular remodeling processes and suggests a novel therapeutic opportunity for the treatment of PAH.

• Phosphodiesterase 10A upregulation contributes to pulmonary vascular remodeling.

  Authors: Xia Tian, Christina Vroom, Hossein Ardeschir Ghofrani, Norbert Weissmann, Ewa Bieniek, Friedrich Grimminger, Werner Seeger, Ralph Theo Schermuly, Soni Savai Pullamsetti

  PloS one. 01/2011; 6(4):e18136.

  Phosphodiesterases (PDEs) modulate the cellular proliferation involved in the pathophysiology of pulmonary hypertension (PH) by hydrolyzing cAMP and cGMP. The present study was designed to determinePhosphodiesterases (PDEs) modulate the cellular proliferation involved in the pathophysiology of pulmonary hypertension (PH) by hydrolyzing cAMP and cGMP. The present study was designed to determine whether any of the recently identified PDEs (PDE7-PDE11) contribute to progressive pulmonary vascular remodeling in PH. All in vitro experiments were performed with lung tissue or pulmonary arterial smooth muscle cells (PASMCs) obtained from control rats or monocrotaline (MCT)-induced pulmonary hypertensive (MCT-PH) rats, and we examined the effects of the PDE10 inhibitor papaverine (Pap) and specific small interfering RNA (siRNA). In addition, papaverine was administrated to MCT-induced PH rats from day 21 to day 35 by continuous intravenous infusion to examine the in vivo effects of PDE10A inhibition. We found that PDE10A was predominantly present in the lung vasculature, and the mRNA, protein, and activity levels of PDE10A were all significantly increased in MCT PASMCs compared with control PASMCs. Papaverine and PDE10A siRNA induced an accumulation of intracellular cAMP, activated cAMP response element binding protein and attenuated PASMC proliferation. Intravenous infusion of papaverine in MCT-PH rats resulted in a 40%-50% attenuation of the effects on pulmonary hypertensive hemodynamic parameters and pulmonary vascular remodeling. The present study is the first to demonstrate a central role of PDE10A in progressive pulmonary vascular remodeling, and the results suggest a novel therapeutic approach for the treatment of PH.

• Redox signaling and reactive oxygen species in hypoxic pulmonary vasoconstriction.

  Authors: Beate Fuchs, Natascha Sommer, Alexander Dietrich, Ralph Theo Schermuly, Hossein Ardeschir Ghofrani, Friedrich Grimminger, Werner Seeger, Thomas Gudermann, Norbert Weissmann

  Respiratory physiology & neurobiology. 12/2010; 174(3):282-91.

  Hypoxic pulmonary vasoconstriction (HPV) is an essential physiological mechanism of the lung that matches blood perfusion with alveolar ventilation to optimize gas exchange. Perturbations of HPV, asHypoxic pulmonary vasoconstriction (HPV) is an essential physiological mechanism of the lung that matches blood perfusion with alveolar ventilation to optimize gas exchange. Perturbations of HPV, as may occur in pneumonia or adult respiratory distress syndrome, can cause life-threatening hypoxemia. Despite intensive research for decades, the molecular mechanisms of HPV have not been fully elucidated. Reactive oxygen species (ROS) and changes in the cellular redox state are proposed to link O2 sensing and pulmonary arterial smooth muscle cell contraction underlying HPV. In this regard, mitochondria and NAD(P)H oxidases are discussed as sources of ROS. However, there is controversy whether ROS levels decrease or increase during hypoxia. With this background we summarize the current knowledge on the role of ROS and redox state in HPV.

• Pulmonary hypertension: updated classification and management of pulmonary hypertension.

  Authors: Holger M Nef, Helge Möllmann, Christian Hamm, Friedrich Grimminger, Hossein-Ardeschir Ghofrani

  Heart (British Cardiac Society). 04/2010; 96(7):552-9.

• Riociguat for pulmonary hypertension.

  Authors: Hossein-Ardeschir Ghofrani, Robert Voswinckel, Henning Gall, Ralph Schermuly, Norbert Weissmann, Werner Seeger, Friedrich Grimminger

  Future cardiology. 03/2010; 6(2):155-66.

  Pulmonary hypertension (PH) encompasses a group of diseases associated with progressively increasing pulmonary vascular resistance, right heart failure and premature death. Riociguat is a novel,Pulmonary hypertension (PH) encompasses a group of diseases associated with progressively increasing pulmonary vascular resistance, right heart failure and premature death. Riociguat is a novel, first-in-class oral drug that directly stimulates soluble guanylate cyclase, both independently of the endogenous vasodilator nitric oxide (NO) and in synergy with NO. Single oral doses of riociguat were well tolerated in a Phase I study of healthy volunteers. They had a favorable safety profile, and improved pulmonary hemodynamics and cardiac index to a greater extent than inhaled NO in a proof-of-concept study in patients with moderate-to-severe PH. In a 12-week Phase II trial in patients with chronic thromboembolic PH or pulmonary arterial hypertension, pulmonary hemodynamics and exercise capacity improved following individual dose titration with oral riociguat, which was generally well tolerated. Further trials in PH have been initiated.

• Long-term therapy with inhaled iloprost in patients with pulmonary hypertension.

  Authors: Horst Olschewski, Marius M Hoeper, Juergen Behr, Ralf Ewert, Andreas Meyer, Mathias M Borst, Jörg Winkler, Michael Pfeifer, Heinrike Wilkens, Hossein Ardeschir Ghofrani, Sylvia Nikkho, Werner Seeger

  Respiratory medicine. 02/2010; 104(5):731-40.

  To investigate the long-term safety of inhaled iloprost in patients with pulmonary hypertension (pH), including idiopathic PAH (IPAH group) and other forms of pulmonary hypertensionTo investigate the long-term safety of inhaled iloprost in patients with pulmonary hypertension (pH), including idiopathic PAH (IPAH group) and other forms of pulmonary hypertension (PHother). Sixty-three patients (IPAH group, n=40, PHother n=23) were enrolled to receive inhaled iloprost either from baseline or after 3 months in a prospective, open-label 2-year study. Iloprost was inhaled 6-9 times daily with a night pause employing a jet nebulizer delivering an inhaled single dose of 4microg at the mouthpiece. In the case of side effects the single dose was reduced to 2microg. Sixty patients received at least 1 dose of inhaled iloprost. Thirty-six patients completed at least 630 days of therapy (25 IPAH, 11 PHother), 19 patients dropped out prematurely and 8 patients died (3 IPAH, 5 PHother). There were no drug-induced toxicities and only mild to moderate side effects. The most common side effects were coughing and flushing. Two-year survival was estimated at 85% (IPAH group 91%, PHother 78%). A modified analysis was performed to correct for differential drop-out. It included follow-up data from the premature discontinuations and revealed a 2-year survival of 87% [95% CI, 76%-98%] in the IPAH group while the predicted survival was 63%. The iloprost dose increased by 16% over 2 years. Inhaled iloprost is well tolerated as long-term therapy and no substantial dose increase is required. Although uncontrolled, the data suggest a long-term clinical benefit from continued therapy with inhaled iloprost.

• Targeting cancer with phosphodiesterase inhibitors.

  Authors: Rajkumar Savai, Soni Savai Pullamsetti, Gamal-Andre Banat, Norbert Weissmann, Hossein Ardeschir Ghofrani, Friedrich Grimminger, Ralph Theo Schermuly

  Expert opinion on investigational drugs. 01/2010; 19(1):117-31.

  Importance of the field: For many cancers, there has been a shift from management with traditional, nonspecific cytotoxic chemotherapies to treatment with molecule-specific targeted therapies thatImportance of the field: For many cancers, there has been a shift from management with traditional, nonspecific cytotoxic chemotherapies to treatment with molecule-specific targeted therapies that are used either alone or in combination with traditional chemotherapy and radiation therapy. Accumulating data suggest that multi-targeted agents may produce greater benefits than those observed with single-targeted therapies, may have acceptable tolerability profiles, and may be active against a broader range of tumour types. Thus, regulation of cyclic nucleotide signalling is properly regarded as a composite of multiple component pathways involved in diverse aspects of tumour cell function. The impairment of cAMP and/or cGMP generation by overexpression of PDE isoforms that has been described in various cancer pathologies, and the effects of PDE inhibitors in tumour models in vitro and in vivo, may offer promising insight into future cancer treatments because of the numerous advantages of PDE inhibitors. Areas covered in this review: In this review, we focus on the expression and regulation of cyclic nucleotide phosphodiesterases (PDEs) in tumour progression and provide evidence that PDE inhibitors may be effective agents for treating cancer; the review covers literature from the past several years. What the reader will gain: PDEs have been studied in a variety of tumours; data have suggested that the levels of PDE activity are elevated and, therefore, the ratio of cGMP to cAMP is affected. In addition, PDE inhibitors may be potential targets for tumour cell growth inhibition and induction of apoptosis. This review explores the prospects of targeting PDEs with therapeutic agents for cancer, as well as the shortcomings of this approach such as dose-limiting side effects, toxicity/efficacy ratio and selectivity towards tumour tissue. In addition, it includes opinions and suggestion for developing PDE inhibition for cancer treatment from initial concept to potential therapeutic application and final relevance in clinical use. Take home message: Impaired cAMP and/or cGMP generation upon overexpression of PDE isoforms has been described in various cancer pathologies. Inhibition of selective PDE isoforms, which raises the levels of intracellular cAMP and/or cGMP, induces apoptosis and cell cycle arrest in a broad spectrum of tumour cells and regulates the tumour microenvironment. Therefore, the development and clinical application of inhibitors specific for individual PDE isoenzymes may selectively restore normal intracellular signalling, providing antitumour therapy with reduced adverse effects.

• Role of Epidermal Growth Factor Inhibition in Experimental Pulmonary Hypertension.

  Authors: Bhola Kumar Dahal, Teodora Cornitescu, Aleksandra Tretyn, Soni Savai Pullamsetti, Djuro Kosanovic, Rio Dumitrascu, Hossein Ardeschir Ghofrani, Norbert Weissmann, Robert Voswinckel, Gamal-Andre Banat, Werner Seeger, Friedrich Grimminger, Ralph Theo Schermuly

  American journal of respiratory and critical care medicine. 10/2009;

  RATIONALE: Epidermal growth factor (EGF) and its receptors play a role in cell proliferation and survival and are implicated in the pathobiology of pulmonary arterial hypertension (PAH). ObjectiveRATIONALE: Epidermal growth factor (EGF) and its receptors play a role in cell proliferation and survival and are implicated in the pathobiology of pulmonary arterial hypertension (PAH). Objective and methods: We investigated a) the effects of three clinically approved EGF receptor (EGFR) antagonists in vitro on rat pulmonary arterial smooth muscle cell (PASMC) proliferation and in vivo on experimental pulmonary hypertension (PH) induced by monocrotaline injection in rats and by chronic hypoxia in mice, and b) the expression of EGFR in the lung tissues from experimental and clinical PH. MEASUREMENTS AND MAIN RESULTS: The EGFR inhibitors, gefitinib, erlotinib and lapatinib inhibited the EGF-induced proliferation of PASMCs. In rats with established PH, gefitinib and erlotinib significantly reduced right ventricular systolic pressure (RVSP) and right ventricular hypertrophy (RVH). In addition, the medial wall thickness and muscularization of pulmonary arteries were improved. In contrast, lapatinib did not provide therapeutic benefit. These EGFR antagonists at their highest tolerable dose did not yield significant improvement in RVSP, RVH and pulmonary vascular remodeling in mice with chronic hypoxic PH. Moreover, no significant alteration in the EGFR expression was detected in the lung tissues from patients with idiopathic PAH (IPAH). CONCLUSION: The partial therapeutic efficacy of the EGFR antagonists in animal models of pulmonary hypertension and the absence of significant alteration in EGFR expression in the lungs from IPAH patients suggest that the EGFR do not represent a promising target for the treatment of pulmonary hypertension.

• Expression and Activity of Phosphodiesterase Isoforms during Epithelial Mesenchymal Transition - The Role of PDE4.

  Authors: Ewa Kolosionek, Rajkumar Savai, Hossein Ardeschir Ghofrani, Norbert Weissmann, Andreas Guenther, Friedrich Grimminger, Werner Seeger, Gamal-Andre Banat, Ralph Theo Schermuly, Soni Savai Pullamsetti

  Molecular biology of the cell. 09/2009;

  Monitoring Editor: Keith E. Mostov Epithelial-to-mesenchymal transition (EMT) has emerged as a critical event in the pathogenesis of organ fibrosis and cancer, and is typically induced by theMonitoring Editor: Keith E. Mostov Epithelial-to-mesenchymal transition (EMT) has emerged as a critical event in the pathogenesis of organ fibrosis and cancer, and is typically induced by the multifunctional cytokine transforming growth factor-beta1 (TGF-beta1). The present study was undertaken to evaluate the potential role of phosphodiesterases (PDEs) in TGF-beta1 induced EMT in the human alveolar epithelial type-II cell line-A549. Stimulation of A549 with TGF-beta1 induced EMT by morphological alterations and by expression changes of the epithelial phenotype markers, E-cadherin, cytokeratin-18, zona occludens-1, and the mesenchymal phenotype markers, collagen I, fibronectin, alpha-smooth muscle actin. Interestingly, TGF-beta1 stimulation caused twofold increase in total cAMP-PDE activity, contributed mostly by PDE4. Further, mRNA and protein expression demonstrated upregulation of PDE4A and PDE4D isoforms in TGF-beta1 stimulated cells. Most importantly, treatment of TGF-beta1 stimulated epithelial cells with the PDE4 selective inhibitor Rolipram or PDE4 siRNA potently inhibited EMT changes in a Smad-independent manner by decreasing reactive oxygen species (ROS), p38 and ERK phosphorylation. In contrast, the ectopic overexpression of PDE4A and/or PDE4D resulted in a significant loss of epithelial marker E-cadherin, but did not result in changes of mesenchymal markers. Additionally, Rho kinase signaling activated by TGF-beta1 during EMT demonstrated to be a positive regulator of PDE4. Collectively, the findings presented herein suggest that TGF-beta1 mediated up-regulation of PDE4 promotes EMT in alveolar epithelial cells. Thus, targeting PDE4 isoforms may be a novel approach to attenuating EMT-associated lung diseases such as pulmonary fibrosis and lung cancer.

• The soluble guanylate cyclase activator HMR1766 reverses hypoxia-induced experimental pulmonary hypertension in mice.

  Authors: Norbert Weissmann, Sascha Hackemack, Bhola Kumar Dahal, Soni Savai Pullamsetti, Rajkumar Savai, Manish Mittal, Beate Fuchs, Thomas Medebach, Rio Dumitrascu, Martin van Eickels, Hossein Ardeschir Ghofrani, Werner Seeger, Friedrich Grimminger, Ralph Theo Schermuly

  American journal of physiology. Lung cellular and molecular physiology. 08/2009;

  Severe pulmonary hypertension (PH) is a disabling disease with high mortality, characterized by pulmonary vascular remodeling and right heart hypertrophy. In mice with PH induced by chronic hypoxia,Severe pulmonary hypertension (PH) is a disabling disease with high mortality, characterized by pulmonary vascular remodeling and right heart hypertrophy. In mice with PH induced by chronic hypoxia, we examined the acute and chronic effects of the soluble guanylate cyclase (sGC) activator HMR1766 on hemodynamics and pulmonary vascular remodeling. In isolated perfused mouse lungs from control animals, HMR1766 dose-dependently inhibited the pressor response of acute hypoxia. This dose response curve was shifted leftward when investigating the effects of HMR1766 in isolated lungs from chronic hypoxic animals for 21 days at 10% oxygen. Mice exposed for 21 or 35 days to chronic hypoxia developed pulmonary hypertension, right heart hypertrophy and pulmonary vascular remodeling. Treatment with HMR1766 (10mg/kg/day), after full establishment of pulmonary hypertension from day 21 to day 35, significantly reduced pulmonary hypertension, as measured continuously by telemetry. In addition, right ventricular (RV) hypertrophy and structural remodeling of the lung vasculature were reduced. Pharmacological activation of oxidized sGC partially reverses hemodynamic and structural changes in chronic hypoxia-induced experimental PH. Key words: Pulmonary hypertension, soluble guanylate cyclase (sGC), Hypoxia, Vascular remodeling.

• Heme Oxygenase-2 and Large-conductance Ca2+ activated K+ Channels: Lung Vascular Effects of Hypoxia.

  Authors: Markus Roth, Markus Rupp, Simone Hofmann, Manish Mittal, Beate Fuchs, Natascha Sommer, Nirmal Parajuli, Karin Quanz, Dominic Schubert, Eva Dony, Ralph Theo Schermuly, Hossein Ardeschir Ghofrani, Ulrike Sausbier, Katrin Rutschmann, Sarah Wilhelm, Werner Seeger, Peter Ruth, Friedrich Grimminger, Matthias Sausbier, Norbert Weissmann

  American journal of respiratory and critical care medicine. 07/2009;

  RATIONALE: Hypoxic pulmonary vasoconstriction (HPV) is an important mechanism by which pulmonary gas exchange is optimized by the adaptation of blood flow to alveolar ventilation. In chronic hypoxia,RATIONALE: Hypoxic pulmonary vasoconstriction (HPV) is an important mechanism by which pulmonary gas exchange is optimized by the adaptation of blood flow to alveolar ventilation. In chronic hypoxia, in addition to HPV a vascular remodeling process leads to pulmonary hypertension. Recently, a complex of heme oxygenase-2 (HO-2) and the BK channel has been suggested as a universal oxygen sensor system. OBJECTIVE: We investigated, whether this complex serves as an oxygen sensor for the vascular effects of alveolar hypoxia in the lung. METHODS and RESULTS: Immunohistochemical analysis of mouse lungs identified HO-2 mainly in pulmonary arteries, the bronchial epithelium and alveolar epithelial cells. BK channel alpha-subunit (BKalpha) immunoreactivity was found primarily in the bronchial and vascular smooth muscle layer. Immunofluorescence staining and co-immunoprecipitation suggested only a weak complexation of HO-2 and BKalpha in pulmonary arterial smooth muscle cells. The strength of acute and sustained HPV, determined in isolated perfused and ventilated lungs, was not different between the wild-type, HO-2- and BKalpha-deficient mice. Exposure of mice to three weeks of chronic hypoxia resulted in a slight downregulation of HO-2 and no alteration in BKalpha expression. The degree of pulmonary hypertension which developed, quantified from right ventricular pressure, right heart hypertrophy, and the degree of muscularization of precapillary pulmonary arteries, was not different between the wild-type, HO-2- or BKalpha-deficient mice. CONCLUSION: It is demonstrated that neither deletion of HO-2 nor BK channels affects the acute, sustained and chronic vascular responses to alveolar hypoxia in the lung.

• Modulating cGMP to Treat Lung Diseases.

  Authors: Hossein Ardeschir Ghofrani, Friedrich Grimminger

  Handbook of experimental pharmacology. 02/2009;

  Background: Nitric oxide (NO) is constitutively produced in the lung by NO-synthases. The main cellular sources of lung NO production are the vascular endothelium and the airway epithelia (Bohle etBackground: Nitric oxide (NO) is constitutively produced in the lung by NO-synthases. The main cellular sources of lung NO production are the vascular endothelium and the airway epithelia (Bohle et al. 2000; German et al. 2000; Ide et al. 1999). Local NO production contributes to regulation of pulmonary perfusion depending on alveolar ventilation to assure optimized ventilation/perfusion distribution (Grimminger et al. 1995). NO-synthase activity is regulated on transcriptional and post-translational redox-based modulation level. The common signaling pathway of endogenous vasodilators, such as nitric oxide, prostaglandins, and natriuretic peptides, engage cyclic nucleotides (cAMP and cGMP). These second messengers are mainly produced by activation of adenylate- and guanylate-cyclases, both membrane-bound and soluble (Beavo 1995). Phosphodiesterases (PDEs) represent a superfamily of enzymes, with PDE1 through PDE11 being currently known, that inactivate cyclic AMP and cyclic GMP, with different tissue distribution and substrate specificities (Ahn et al. 1991; Von Euler and Liljestrand. 1946). Because of stabilization of these second messengers, PDE inhibitors differentially regulate levels of cAMP and/or cGMP, depending on their selectivity profile. Recently, direct activators and stimulators of the sGC have been suggested as new therapeutic tools for the treatment of lung vascular disorders that might have even higher potency than PDE inhibitors or exogenously applied NO.

• Novel soluble guanylyl cyclase stimulator BAY 41-2272 attenuates ischemia/reperfusion induced lung injury.

  Authors: Bakytbek Egemnazarov, Akylbek Sydykov, Ralph Theo Schermuly, Norbert Weissmann, Johannes-Peter Stasch, Akpai S Sarybaev, Werner Seeger, Friedrich Grimminger, Hossein Ardeschir Ghofrani

  American journal of physiology. Lung cellular and molecular physiology. 01/2009;

  The protective effects of nitric oxide (NO), a physiological activator of soluble guanylyl cyclase (sGC), have been reported in ischemia/reperfusion (I/R) syndrome of the lung. Therefore, we studiedThe protective effects of nitric oxide (NO), a physiological activator of soluble guanylyl cyclase (sGC), have been reported in ischemia/reperfusion (I/R) syndrome of the lung. Therefore, we studied the effects of BAY 41-2272, a novel sGC stimulator, on I/R injury of the lung in an isolated intact organ model. Lung injury was assessed by measuring weight gain and microvascular permeability (capillary filtration coefficient, Kfc). Release of reactive oxygen species (ROS) into the perfusate was measured during early reperfusion by ESR spectroscopy. Rabbit lungs were treated with BAY 41-2272, L-NMMA 400, or NO to evaluate the effects on I/R induced lung injury. In untreated lungs a dramatic rise in Kfc values and weight gain during reperfusion were observed, these results were associated with increased ROS production. Both, BAY 41-2272 and L-NMMA significantly attenuated vascular leakage and suppressed ROS release. Additional experiments showed that BAY 41-2272 diminished phorbol-12-myristrate-13-acetate induced ROS production by NADPH-oxidase. A pharmacological inhibition of the enzyme with consequent reduction in ROS levels decreased I/R injury. Nitric oxide had only marginal effect on I/R injury. Thus, BAY 41-2272 protects against I/R induced lung injury by interfering with the activation of NADPH-oxidases. Key words: acute lung injury, nitric oxide, oxidative stress, ROS.

• Pulmonary vascular disease in the developing world.

  Authors: Ghazwan Butrous, Hossein Ardeschir Ghofrani, Friedrich Grimminger

  Circulation. 11/2008; 118(17):1758-66.

• Role of the prostanoid EP4 receptor in iloprost-mediated vasodilatation in pulmonary hypertension.

  Authors: Ying-Ju Lai, Soni Savai Pullamsetti, Eva Dony, Norbert Weissmann, Ghazwan Butrous, Gamal-Andre Banat, Hossein Ardeschir Ghofrani, Werner Seeger, Friedrich Grimminger, Ralph Theo Schermuly

  American journal of respiratory and critical care medicine. 08/2008; 178(2):188-96.

  RATIONALE: Iloprost is effective for the treatment of pulmonary hypertension. It acts through elevation of cAMP by binding to the prostacyclin receptor (IP receptor). However, there is evidence thatRATIONALE: Iloprost is effective for the treatment of pulmonary hypertension. It acts through elevation of cAMP by binding to the prostacyclin receptor (IP receptor). However, there is evidence that patients with severe pulmonary hypertension have decreased expression of the IP receptor in the remodeled pulmonary arterial smooth muscle. OBJECTIVES: We hypothesized that prostanoid receptors other than the IP receptor are involved in signal transduction by iloprost. METHODS: Immunoblotting was used to detect the IP and prostanoid EP4 receptor in lung tissue from patients with idiopathic pulmonary arterial hypertension, and immunohistochemistry was used to detect these receptors in lung sections from rats treated with monocrotaline (MCT28d). Protein and mRNA were isolated from pulmonary arterial smooth muscle cells (PASMCs) from control and MCT28d rats treated with AH6809 (an EP2 receptor antagonist) and AH23848 (an EP4 receptor antagonist) in combination with iloprost. Intracellular cAMP was also assessed in these tissues. Measurements and Main Results: IP receptor expression was reduced in idiopathic pulmonary arterial hypertension patient lung samples and MCT28d rat lungs compared with the controls. Reverse transcriptase-polymerase chain reaction and immunoblotting of MCT28d rat PASMC extracts revealed scant expression of the IP receptor but stable expression of EP4 receptor, compared with controls. Iloprost-induced elevation in intracellular cAMP in PASMCs was dose-dependently reduced by AH23848, but not by AH6809. CONCLUSIONS: Iloprost mediates vasodilatory functions via the EP4 receptor in the case of low IP receptor expression associated with pulmonary arterial hypertension. This is a previously unrecognized mechanism for iloprost, and illustrates that the EP4 receptor may be a novel therapeutic approach for the treatment of pulmonary arterial hypertension.

• Hypoxia-induced pulmonary hypertension: different impact of iloprost, sildenafil, and nitric oxide.

  Authors: Norbert Weissmann, Boris Gerigk, Ozlem Kocer, Matthias Nollen, Sascha Hackemack, Hossein Ardeschir Ghofrani, Ralph Theo Schermuly, Ghazwan Butrous, Andreas Schulz, Markus Roth, Werner Seeger, Friedrich Grimminger

  Respiratory medicine. 11/2007; 101(10):2125-32.

  OBJECTIVES: Chronic alveolar hypoxia induces pulmonary hypertension, evident from elevated pulmonary artery pressure (PAP), pulmonary vascular resistance, right ventricular hypertrophy (RVH), andOBJECTIVES: Chronic alveolar hypoxia induces pulmonary hypertension, evident from elevated pulmonary artery pressure (PAP), pulmonary vascular resistance, right ventricular hypertrophy (RVH), and increased muscularization of the pulmonary vasculature. Additionally, the vasoconstrictor response to acute hypoxia (HPV) may be reduced in the remodeled vasculature. However, no direct comparison of different treatments on the various parameters characterizing pulmonary hypertension has been performed yet. Against this background, we compared the effects of inhaled NO, infused iloprost, a stable prostacyclin analogue, and oral sildenafil, a phosphodiesterase 5 inhibitor, on hypoxia-induced pulmonary hypertension. METHODS: Exposure of rabbits to chronic hypoxia (FiO(2)=0.10) for 42 days. Treatment with infused iloprost, oral sildenafil, and inhaled nitric oxide. RESULTS: We quantified PAP, pulmonary vascular resistance, RVH, vascular remodeling, vasoreactivity, and the strength of HPV. Chronic hypoxia resulted in an increase in (a) the right ventricle/(left ventricle+septum) ratio from 0.26+/-0.01 to 0.44+/-0.01, (b) PAP, and (c) the degree of muscularization from 14.0+/-4.0% to 43.5+/-5.3%. Treatment with iloprost and sildenafil, but not with NO, prevented the increase in muscularization. In contrast, RVH was strongly inhibited by sildenafil, whereas NO had some minor, and iloprost had no effect. Only iloprost reduced PAP compared to NO and sildenafil. The downregulation of HPV was abrogated only by NO. CONCLUSION: We demonstrated (a) that the parameters characterizing hypoxia-induced pulmonary hypertension are not functionally linked, (b) that the downregulation of HPV under chronic hypoxia can be prevented by inhaled NO but not by sildenafil and iloprost, and (c) that iloprost is particularly effective in preventing vascular remodeling and sildenafil in preventing RVH.