Ralph T. Schermuly

Reversal of experimental pulmonary hypertension by PDGF inhibition

Progression of pulmonary hypertension is associated with increased proliferation and migration of pulmonary vascular smooth muscle cells. PDGF is a potent mitogen and involved in this process. We now report that the PDGF receptor antagonist STI571 (imatinib) reversed advanced pulmonary vascular disease in 2 animal models of pulmonary hypertension. In rats with monocrotaline-induced pulmonary hypertension, therapy with daily administration of STI571 was started 28 days after induction of the disease. A 2-week treatment resulted in 100% survival, compared with only 50% in sham-treated rats. The changes in RV pressure, measured continuously by telemetry, and right heart hypertrophy were reversed to near-normal levels. STI571 prevented phosphorylation of the PDGF receptor and suppressed activation of downstream signaling pathways. Similar results were obtained in chronically hypoxic mice, which were treated with STI571 after full establishment of pulmonary hypertension. Moreover, expression of the PDGF receptor was found to be significantly increased in lung tissue from pulmonary arterial hypertension patients compared with healthy donor lung tissue. We conclude that STI571 reverses vascular remodeling and cor pulmonale in severe experimental pulmonary hypertension regardless of the initiating stimulus. This regimen offers a unique novel approach for antire-modeling therapy in progressed pulmonary hypertension.

Sildenafil for treatment of lung fibrosis and pulmonary hypertension: a randomised controlled trial

BACKGROUND Lung fibrosis can be complicated by pulmonary hypertension, limiting exercise tolerance and life expectancy. Furthermore, vasodilators might cause deterioration in gas exchange. Our aim was to compare acute effects of sildenafil, nitric oxide, and epoprostenol in individuals with pulmonary hypertension secondary to lung fibrosis. METHODS We did a randomised controlled, open-label trial, in 16 individuals admitted to our hospital with pulmonary hypertension secondary to lung fibrosis. After inhalation of nitric oxide (10-20 ppm), we assigned patients to either maximum tolerated dose of intravenous epoprostenol (mean 8.0 ng/kg per min; n=8) or oral sildenafil (50 mg; n=8). Our primary objective was to assess pulmonary vasodilative potency (decrease in pulmonary vascular resistance index) of sildenafil by comparison with inhaled nitric oxide and infused epoprostenol. Analyses were by intention to treat. FINDINGS Pulmonary vascular resistance index was reduced by nitric oxide (-21.9%, 95% CI -14.1 to -36.2), epoprostenol (-36.9%, -24.4 to -59.6), and sildenafil (-32.5%, -10.2 to -54.1). However, ratio of pulmonary to systemic vascular resistance decreased only in individuals who received nitric oxide and sildenafil. Baseline measurement of multiple-inert-gas elimination showed right-to-left shunt flow (4.8%, 0.0-28.2) and little perfusion of low ventilation(V)/perfusion(Q) areas (0.1%, 0.0-13.0). Prostacyclin increased V/Q mismatch (shunt 16.8%, 10.8-35.9; low V/Q 3.8%, 0.0-13.0) and decreased arterial oxygenation. By contrast, nitric oxide (4.5%, 0.0-18.0; 0.0%, 0.0-17.3) and sildenafil (3.3%, 0.0-11.3; 0.0%, 0.0-12.4) maintained V/Q matching, with raised arterial partial pressure of oxygen (14.3 mm Hg, -1.7 to 31.3) noted for sildenafil. We recorded no adverse events. INTERPRETATION Sildenafil causes preferential pulmonary vasodilation and improves gas exchange in patients with severe lung fibrosis and secondary pulmonary hypertension.

Inflammation, Growth Factors, and Pulmonary Vascular Remodeling

Inflammatory processes are prominent in various types of human and experimental pulmonary hypertension (PH) and are increasingly recognized as major pathogenic components of pulmonary vascular remodeling. Macrophages, T and B lymphocytes, and dendritic cells are present in the vascular lesions of PH, whether in idiopathic pulmonary arterial hypertension (PAH) or PAH related to more classical forms of inflammatory syndromes such as connective tissue diseases, human immunodeficiency virus (HIV), or other viral etiologies. Similarly, the presence of circulating chemokines and cytokines, viral protein components (e.g., HIV-1 Nef), and increased expression of growth (such as vascular endothelial growth factor and platelet-derived growth factor) and transcriptional (e.g., nuclear factor of activated T cells or NFAT) factors in these patients are thought to contribute directly to further recruitment of inflammatory cells and proliferation of smooth muscle and endothelial cells. Other processes, such as mitochondrial and ion channel dysregulation, seem to convey a state of cellular resistance to apoptosis; this has recently emerged as a necessary event in the pathogenesis of pulmonary vascular remodeling. Thus, the recognition of complex inflammatory disturbances in the vascular remodeling process offers potential specific targets for therapy and has recently led to clinical trials investigating, for example, the use of tyrosine kinase inhibitors. This paper provides an overview of specific inflammatory pathways involving cells, chemokines and cytokines, cellular dysfunctions, growth factors, and viral proteins, highlighting their potential role in pulmonary vascular remodeling and the possibility of future targeted therapy.

Combination Therapy with Oral Sildenafil and Inhaled Iloprost for Severe Pulmonary Hypertension

Context Common therapies for pulmonary hypertension have disadvantages: Continuous intravenous epoprostenol may cause sepsis, hypotension, and tachyphylaxis; nitric oxide requires continuous inhalation; and inhaled iloprost requires up to 12 doses per day. Phosphodiesterase in lung tissue (PDE-5) inhibits the action of these therapies by inactivating the second messengers of prostacyclin and nitric oxide (cyclic adenosine monophosphate and cyclic guanosine monophosphate). Because sildenafil blocks the action of PDE-5, thereby causing vascular dilatation, it could be useful in treating pulmonary hypertension. Contribution This randomized, controlled trial of low- or high-dose sildenafil, with or without inhaled iloprost, showed dose-dependent improvement in mean pulmonary artery pressure and hemodynamics with sildenafil alone. Iloprost amplified the effects. Implications Sildenafil may enhance the management of pulmonary hypertension. The Editors Severe pulmonary hypertension is a debilitating disease with short life expectancy that often affects young people. Continuous intravenous administration of epoprostenol has been used for pulmonary vasodilatation and was shown to improve exercise capacity and survival in patients with primary pulmonary hypertension (1). However, this therapy is limited by serious infectious complications of the intravenous line, systemic side effects due to the nonselectivity of the vasodilator response, and very high costs incurred because of tachyphylaxis with long-term administration. The vasodilatory effects of nitric oxide administered by inhalation are restricted to the pulmonary vasculature. Nitric oxide has a very short half-life and is used as a screening agent for lung vasoreactivity (2). Inhalation of aerosolized iloprost, a long-acting prostacyclin analogue, causes strong preferential pulmonary vasodilatation in both primary and secondary pulmonary hypertension (3-6). Long-term use of nebulized iloprost was shown to be beneficial in severe primary pulmonary hypertension and overt right-heart failure (7, 8), but because the drug wears off in about 60 minutes, patients must take 6 to 12 inhalations daily to achieve sustained relief of pulmonary hypertension. Phosphodiesterases are a superfamily of enzymes that inactivate cyclic adenosine monophosphate and cyclic guanosine monophosphate, the second messengers of prostacyclin and nitric oxide. The phosphodiesterases have different tissue distributions and substrate affinities (9); in particular, phosphodiesterase-5 is abundantly expressed in lung tissue (10). Inhibition of phosphodiesterase may augment and prolong prostanoid- and nitric oxiderelated vascular effects. The novel selective phosphodiesterase-5 inhibitor sildenafil has been approved for treatment of erectile dysfunction. Sildenafil causes only very minor systemic hemodynamic effects in healthy humans (11). Data from an experimental model of pulmonary hypertension (12) and two recent case reports (13, 14) suggested that sildenafil might be an effective pulmonary vasodilator. We compared the pulmonary vasodilatory effect of sildenafil with that of inhaled nitric oxide and aerosolized iloprost in 30 patients with severe pulmonary hypertension. We evaluated doseresponse characteristics of sildenafil alone and in combination with inhaled prostanoid during right-heart catheterization. Methods Patients Thirty patients (23 women and 7 men) with severe pulmonary hypertension (mean pulmonary arterial pressure > 40 mm Hg) were included. Sixteen patients had pulmonary arterial hypertension, as defined by the World Health Organization World Symposium on Primary Pulmonary Hypertension (15); of these patients, 10 had primary pulmonary hypertension and 6 had calcinosis, the Raynaud phenomenon, esophageal dysfunction, sclerodactyly, and telangiectasia (the CREST syndrome). One patient had aplasia of the left pulmonary artery. Thirteen patients had chronic thromboembolic pulmonary hypertension and were not candidates for surgery. All patients were admitted for testing of pulmonary vasoreactivity and evaluation of therapeutic options. Seventeen patients were tested for the first time and had not been previously treated with inhaled iloprost. Thirteen patients had been tested at least 3 months after previous vasodilator testing; 11 of these patients were receiving long-term therapy with inhaled iloprost. Exclusion criteria were pulmonary hypertension secondary to chronic obstructive pulmonary disease, pulmonary venous congestion, congenital heart disease, acute or chronic inflammatory lung disease, pregnancy or insufficient contraceptive measures, or previous treatment with phosphodiesterase inhibitors, including theophylline preparations. The individual response to vasodilators, including inhaled nitric oxide, was neither an inclusion nor an exclusion criterion. Treatment The study protocol was approved by the Justus-Liebig-University Ethics Committee, and each patient gave written informed consent. A 7.5-French SwanGanz fiberoptic thermodilution pulmonary artery catheter (93A-754H, Baxter Healthcare, Irvine, California) was inserted via the right jugular vein by using standard techniques. Each patient received short-term inhaled nitric oxide; the maximum vasodilator response to this agent required nitric oxide, 20 to 40 parts per million. If necessary, patients received nasal oxygen throughout the test to achieve arterial oxygen saturation greater than 88%. After hemodynamic values returned to baseline, inhaled iloprost was delivered by using an ultrasonic nebulizer (Multisonic compact [Otto Schill GmbH, Probstzella, Germany]; mass median aerodynamic diameter, 3.9 m; duration of inhalation, 4 minutes; total inhaled dose, 2.8 g) (16). Hemodynamic and gas exchange variables were measured at 5, 15, 30, 60, and 90 minutes after iloprost inhalation. At the end of a 2-hour observation period, when hemodynamics had returned to baseline values, patients were randomly assigned to one of four treatment groups: 12.5 mg of oral sildenafil alone (n = 7); 12.5 mg of oral sildenafil, followed by 2.8 g of inhaled iloprost 1 hour later (n = 7); 50 mg of oral sildenafil alone (n = 8); or 50 mg of oral sildenafil, followed by 2.8 g of inhaled iloprost 1 hour later (n = 8). Patients were assigned to the therapeutic regimens by using computerized randomization in groups of four; no more than two patients in a row were assigned to one group. Hemodynamic measurements and blood sampling were performed at 15, 30, 60, 90, and 120 minutes (and at 150 and 180 minutes in the group receiving 12.5 mg of sildenafil plus iloprost and in the group receiving 50 mg of sildenafil plus iloprost) after administration of oral sildenafil. To assess the overall vasodilator response, the area under the curve (AUC) of reduction in pulmonary vascular resistance was calculated as the integral of the difference between preintervention baseline values until pulmonary vascular resistance again reached 95% of baseline values or 120 minutes. Statistical Analysis Data are given as the mean and 95% CIs. For every variable, the response to vasodilator treatment was considered significant if the 95% (P < 0.05), 99% (P < 0.01), or 99.9% (P < 0.001) CI of the difference between pretreatment and post-treatment values did not overlap with zero. Two-way analysis of variance (ANOVA) was performed to test for significant differences and possible interaction of vasoreactivity to different vasodilators with underlying disease. One-way ANOVA with the Scheff post-test was used to determine hemodynamic responsiveness to the vasodilator regimens when two-way ANOVA indicated that underlying disease was a significant factor. Results Baseline Hemodynamics The Table shows baseline hemodynamics in all treatment groups. All patients had severe pulmonary hypertension and low cardiac index values. No pulmonary venous congestion was observed, as indicated by low pulmonary arterial occlusion pressure (8.7 0.6 mm Hg). On ANOVA, the groups did not differ significantly in any baseline characteristic. Table. Baseline Hemodynamic Data Inhaled Nitric Oxide Inhalation of nitric oxide decreased mean pulmonary arterial pressure (change, 7.0% [95% CI, 3.8% to 10.2%]), increased the cardiac index (change, 7.9% [CI, 3.4% to 12.5%]), and decreased pulmonary vascular resistance (change, 14.1% [CI, 19.1% to 9.2%]) (Figure 1). The pulmonary selectivity of the vasodilatory effect was indicated by a significant reduction in the ratio of pulmonary to systemic vascular resistance. The decrease in pulmonary vascular resistance in response to inhaled nitric oxide did not differ significantly among groups (Figure 2). The effects of nitric oxide wore off within 15 minutes. Figure 1. Hemodynamic responses to vasodilators. P P P P Figure 2. Time to decrease in pulmonary vascular resistance in response to vasodilator challenge. NO Inhaled Iloprost Aerosolized iloprost decreased the pulmonary vascular resistance by 27.1% (CI, 22.2% to 32.1%) and increased the cardiac index by 22.8% (CI, 17.6% to 27.9%) (Figure 1). The decrease in the ratio of pulmonary to systemic vascular resistance indicated preferential pulmonary vasodilatation. No significant differences among groups in the vasodilator response to inhaled iloprost were observed (Figure 2). The effects of iloprost wore off within 60 to 90 minutes. Oral Sildenafil Administration of 12.5 mg or 50 mg of sildenafil decreased mean pulmonary arterial pressure in a dose-dependent manner (change, 8.5% [CI, 15.1% to 1.9%] and 13.5% [CI, 23.8% to 3.2%], respectively) (Figure 1). The cardiac index increased by 5.0% (CI, 0.5% to 10.5%) and 13.2% (CI, 4.3% to 22.2%), and pulmonary vascular resistance decreased by 14.7% (CI, 6.6% to 22.7%) and 24.3% (CI, 16.7% to 31.8%). The decrease in the ratio of pulmonary to systemic vascular resistance indicated preferential pulmonary vasodilatation. Arterial oxygen did not decrease significantly (data not sh

Aerosolized prostacyclin and iloprost in severe pulmonary hypertension

Primary pulmonary hypertension is a progressive fatal disease of unknown cause, and patients with this condition have a median life expectancy of less than 3 years after diagnosis [1]. Death is most closely associated with an increase in pulmonary artery pressure and right atrial pressure and a decrease in cardiac output due to failure of the right side of the heart. The responsiveness of pulmonary hypertension to various vasodilator agents led to the speculation that, concomitant with vascular remodeling processes, persistent vasoconstriction is an important feature of the disease. Long-term use of calcium-channel blockers improves the survival rate in approximately 25% of patients; in such patients, the response to these drugs is a substantial decrease in pulmonary artery pressure and pulmonary vascular resistance [2]. The main hazards of this therapy are systemic hypotension and worsening of right ventricular function. Intravenous prostacyclin is a potent pulmonary vasodilator in patients with primary pulmonary hypertension, but it requires continuous intravenous access and, like calcium antagonists, lacks selectivity for the lung vasculature [3-6]. We recently used aerosol techniques for preferential distribution of prostacyclin to well-ventilated lung areas, thereby achieving selective pulmonary vasodilatation with a concomitant improvement of ventilation-perfusion matching in mechanically ventilated patients with the adult respiratory distress syndrome [7]. This approach has also been shown to be effective in hypoxia-induced pulmonary hypertension in dogs [8]. We extended this strategy by nebulizing prostacyclin and a stable analog, iloprost, in patients with severe pulmonary hypertension. Methods Four patients with primary pulmonary hypertension and two patients with severe pulmonary hypertension associated with calcinosis, the Raynaud phenomenon, esophageal dysfunction, sclerodactyly, and telangiectasia (the CREST syndrome), all of whom were classified as New York Heart Association class III or class IV. Patients gave informed consent to the test trial, which was approved by the institutional ethics committee of Justus-Liebig-University, Giessen, Germany. Two of the patients with primary pulmonary hypertension (patient A, 30 years of age, and patient B, 31 years of age) developed the disease several months after pregnancy. In the other two patients with pulmonary hypertension (patient C, a 37-year-old man, and patient D, a 56-year-old woman), no trigger was identified. The patients with the CREST syndrome (patient E, a 56-year-old woman who had received a diagnosis of the syndrome 2 years earlier, and patient F, a 54-year-old woman who had received this diagnosis 15 years earlier) had no involvement of the inner organs and had never received immunosuppressive therapy other than low-dose corticosteroids. Pulmonary embolism was excluded by pulmonary angiograms and ventilation and perfusion lung scans in each patient. Chest radiography and high-resolution computed tomography showed no lung fibrosis, and pulmonary function testing showed no evidence of obstructive or restrictive lung disease. Only one patient (patient B) responded to calcium antagonists according to the criteria given by Rich and colleagues [2]. Longterm therapy included anticoagulation and diuretics (in all patients) and low-dose steroids (in patients E and F). A fiberoptic thermodilution pulmonary artery catheter was used to measure central venous pressure, pulmonary artery pressure, pulmonary artery wedge pressure, cardiac output, and venous oxygen saturation; a femoral artery catheter was used to assess mean arterial pressure and arterial oxygen saturation. In each test trial, the following were administered: 1. Oxygen, 2 to 8 L/min. 2. Inhaled nitric oxide, 10 to 28 parts per million. 3. Intravenous prostacyclin (epoprostenol sodium, Wellcome Research Laboratories, Beckenham, Kent, United Kingdom), increased in increments of 2.5 ng/kg min1 until patients had discomfort (thoracic oppression, heat, or headache) or until mean arterial pressure decreased to less than 70 mm Hg. The highest tolerable dose (5 to 7.5 ng/kg min1) was continued for 20 minutes. 4. Aerosolized prostacyclin (25 to 50 g of glycine buffer per mL), jet-nebulized with room air at a pressure of 153 kPa (fluid flux, 0.15 mL/min; mass median aerodynamic diameter of particles, 2.9 m; geometric SD, 3.1, ascertained by impactor technique) and delivered to a spacer connected to the afferent limb of a y-valve mouthpiece for 15 minutes (total nebulized dose, 52 to 112 g). 5. Aerosolized iloprost (Ilomedin, Schering AG, Berlin, Germany) (5 to 10 g of saline per mL), administered according to prostacyclin (total nebulized dose, 9 to 21 g). Patients were tested on a separate day for the presence of calcium antagonists (data not shown). All other measurements were taken before, during, and after application of each test trial challenge. The time between the different maneuvers was sufficient for a new stable baseline period. The number of challenges required 10 hours for one entire trial sequence. These trials were done twice within a 1-month period in all patients, and the data were averaged (except for the data from patient B, who took part in only one test trial). One patient (patient E) subsequently began receiving long-term treatment with aerosolized iloprost (100 g/d, divided into six aerosol doses), which has now been continued for 1.5 years. Values before and after challenge in six patients are given for each challenge and are expressed as mean SD. We tested these values for significance using the two-tailed Student t-test for intrapair differences; these levels of significance are given. Results In all six patients, the high pulmonary artery pressure and pulmonary vascular resistance values, the low cardiac output and venous oxygen saturation values, and the increased central venous pressure value indicated advanced disease (Table 1). As we had anticipated, arterial and venous oxygen saturation values increased with oxygen, but hemodynamics improved only moderately. Inhaled nitric oxide substantially decreased pulmonary artery pressure, pulmonary vascular resistance, and central venous pressure and increased cardiac output and venous oxygen saturation. Inhalation only moderately decreased systemic vascular resistance. After cessation of nitric oxide therapy, values returned to baseline within 2 to 5 minutes. Intravenous prostacyclin substantially decreased pulmonary vascular resistance and increased cardiac output in all patients, resulting in a modest decline of pulmonary artery pressure but a substantial decrease in systemic arterial pressure and an increase in heart rate due to peripheral vasodilatation. Aerosolized prostacyclin achieved the same reduction in pulmonary vascular resistance with a smaller increase in cardiac output but a significant decline in pulmonary artery pressure. Moreover, arterial oxygen saturation values were increased with aerosolized prostacyclin but remained unchanged, on average, after intravenous prostanoid application. In all patients, the stable prostacyclin analog iloprost caused nearly identical changes in hemodynamics and gas exchange (example given in Figure 1, further data not shown). The effects of iloprost, however, lasted longer: After termination of aerosolization, prostacyclin-induced changes returned to baseline within 10 to 30 minutes and iloprost-induced changes, within 60 to 120 minutes. Table 1. Hemodynamics and Gas Exchange in Response to Acute Exposure to Vasoactive Agents* Figure 1. Short- and long-term effects of iloprost inhalation in one patient. Left. o Right. o arrows Because response to inhaled prostanoids was favorable, we initiated regular long-term therapy with aerosolized iloprost in one patient (patient E; see Figure 1). Daily inhalation of the prostanoid was well tolerated, and sustained responsiveness of the pulmonary vasculature was seen over a 1-year period. This patients New York Heart Association class improved from IV to III, and her CREST-related skin lesions healed. Discussion Prostacyclin is one of the most potent pulmonary vasodilatory agents available for clinical purposes. Moreover, in severe pulmonary hypertension, local deficiencies of prostacyclin may play a role in the genesis and progression of vascular remodeling [4]. Accordingly, long-term therapy with high doses of prostacyclin resulted in increased life expectancy and exercise tolerance [6]. We describe a technique of intra-alveolar deposition of prostacyclin in patients with severe pulmonary hypertension. The acute effects on pulmonary vascular resistance seen with this technique corresponded to those of intravenous prostacyclin, but selectivity for the pulmonary circulation was achieved, as indicated by a substantial decrease of pulmonary artery pressure and a smaller effect on systemic arterial pressure. Moreover, preferential distribution of the aerosolized vasodilator to the best-ventilated lung areas, which improved ventilation-perfusion matching, was suggested by an increase in arterial oxygen saturation. This effect was even superior to that of inhaled nitric oxide, which has previously been shown to cause selective pulmonary vasodilatation in patients with severe pulmonary hypertension [9, 10]. We found that the nitric oxide-evoked decrease in pulmonary vascular resistance was less than that seen in response to inhaled or intravenous prostacyclin in all patients. Iloprost showed an identical efficacy profile but a longer duration of pulmonary vasodilatation in comparison with prostacyclin. It was well tolerated in the patient who received long-term therapy with daily inhalations of this drug, and sustained responsiveness of the pulmonary vasculature was seen. Interestingly, the beneficial effects were achieved by a total aerosolized iloprost quantity of 100 g/d, which corresponds to a dosage of only approximately 1.1 ng/kg min1

Oral sildenafil as long-term adjunct therapy to inhaled iloprost in severe pulmonary arterial hypertension

OBJECTIVES We sought to investigate the impact of adjunct sildenafil on exercise capacity and hemodynamic parameters in patients with pulmonary arterial hypertension (PAH) who fulfilled predefined criteria of deterioration despite ongoing treatment with inhaled iloprost. BACKGROUND Inhaled iloprost is an effective therapy in PAH. The phosphodiesterase-5 inhibitor sildenafil exerts pulmonary vasodilation and may amplify prostanoid efficacy. METHODS Of 73 PAH patients receiving long-term inhaled iloprost treatment, 14 fulfilled criteria of deterioration unresponsive to conventional treatment. These patients received adjunct oral sildenafil over a period of nine to 12 months, leaving the inhalative iloprost regimen unchanged. RESULTS Before iloprost therapy, the baseline 6-min walking distance was 217 +/- 31 m (mean +/- SEM), with an improvement to 305 +/- 28 m within the first three months of iloprost treatment and a subsequent decline to 256 +/- 30 m after 18 +/- 4 months. Adjunct therapy with sildenafil reversed the deterioration and increased the 6-min walk distance to 346 +/- 26 m (p = 0.002, Wilcoxon test) at three months of combined therapy, with a sustained efficacy up to 12 months (349 +/- 32 m, p = 0.002). The distribution of New York Heart Association functional classes (IV/III/II) improved from September 9, 2000, before sildenafil, to January 8, 2003, after nine to 12 months with sildenafil. All hemodynamic variables changed favorably: pulmonary vascular resistance decreased from 2,494 +/- 256 before sildenafil to 1,950 +/- 128 dynes.s.cm(-5).m(2) after three months of adjunct sildenafil (p = 0.036). Two patients died of severe pneumonia during the period of combined therapy. No further serious adverse events occurred. CONCLUSIONS; In patients with severe PAH deteriorating despite ongoing prostanoid treatment, long-term adjunct oral sildenafil improves exercise capacity and pulmonary hemodynamics. A combination of prostanoids and sildenafil is an appealing concept for future treatment of pulmonary hypertension.

Mechanisms of disease: pulmonary arterial hypertension

Our understanding of, and approach to, pulmonary arterial hypertension has undergone a paradigm shift in the past decade. Once a condition thought to be dominated by increased vasoconstrictor tone and thrombosis, pulmonary arterial hypertension is now seen as a vasculopathy in which structural changes driven by excessive vascular cell growth and inflammation, with recruitment and infiltration of circulating cells, play a major role. Perturbations of a number of molecular mechanisms have been described, including pathways involving growth factors, cytokines, metabolic signaling, elastases, and proteases, that may underlie the pathogenesis of the disease. Elucidating their contribution to the pathophysiology of pulmonary arterial hypertension could offer new drug targets. The role of progenitor cells in vascular repair is also under active investigation. The right ventricular response to increased pressure load is recognized as critical to survival and the molecular mechanisms involved are attracting increasing interest. The challenge now is to integrate this new knowledge and explore how it can be used to categorize patients by molecular phenotype and tailor treatment more effectively.

Hypoxia-Dependent Regulation of Nonphagocytic NADPH Oxidase Subunit NOX4 in the Pulmonary Vasculature

Nonphagocytic NADPH oxidases have recently been suggested to play a major role in the regulation of physiological and pathophysiological processes, in particular, hypertrophy, remodeling, and angiogenesis in the systemic circulation. Moreover, NADPH oxidases have been suggested to serve as oxygen sensors in the lung. Chronic hypoxia induces vascular remodeling with medial hypertrophy leading to the development of pulmonary hypertension. We screened lung tissue for the expression of NADPH oxidase subunits. NOX1, NOXA1, NOXO1, p22phox, p47phox, p40phox, p67phox, NOX2, and NOX4 were present in mouse lung tissue. Comparing mice maintained for 21 days under hypoxic (10% O2) or normoxic (21% O2) conditions, an upregulation exclusively of NOX4 mRNA was observed under hypoxia in homogenized lung tissue, concomitant with increased levels in microdissected pulmonary arterial vessels. In situ hybridization and immunohistological staining for NOX4 in mouse lungs revealed a localization of NOX4 mRNA and protein predominantly in the media of small pulmonary arteries, with increased labeling intensities after chronic exposure to hypoxia. In isolated pulmonary arterial smooth muscle cells (PASMCs), NOX4 was localized primarily to the perinuclear space and its expression levels were increased after exposure to hypoxia. Treatment of PASMCs with siRNA directed against NOX4 decreased NOX4 mRNA levels and reduced PASMC proliferation as well as generation of reactive oxygen species. In lungs from patients with idiopathic pulmonary arterial hypertension (IPAH), expression levels of NOX4, which was localized in the vessel media, were 2.5-fold upregulated. These results support an important role for NOX4 in the vascular remodeling associated with development of pulmonary hypertension.

Relevant Issues in the Pathology and Pathobiology of Pulmonary Hypertension

Knowledge of the pathobiology of pulmonary hypertension (PH) continues to accelerate. However, fundamental gaps remain in our understanding of the underlying pathological changes in pulmonary arteries and veins in the different forms of this syndrome. Although PH primarily affects the arteries, venous disease is increasingly recognized as an important entity. Moreover, prognosis in PH is determined largely by the status of the right ventricle, rather than the levels of pulmonary artery pressures. It is increasingly clear that although vasospasm plays a role, PH is an obstructive lung panvasculopathy. Disordered metabolism and mitochondrial structure, inflammation, and dysregulation of growth factors lead to a proliferative, apoptosis-resistant state. These abnormalities may be acquired, genetically mediated as a result of mutations in bone morphogenetic protein receptor-2 or activin-like kinase-1, or epigenetically inherited (as a result of epigenetic silencing of genes such as superoxide dismutase-2). There is a pressing need to better understand how the pathobiology leads to severe disease in some patients versus mild PH in others. Recent recognition of a potential role of acquired abnormalities of mitochondrial metabolism in the right ventricular myocytes and pulmonary vascular cells suggests new therapeutic approaches, diagnostic modalities, and biomarkers. Finally, dissection of the role of pulmonary inflammation in the initiation and promotion of PH has revealed a complex yet fascinating interplay with pulmonary vascular remodeling, promising to lead to novel therapeutics and diagnostics. Emerging concepts are also relevant to the pathobiology of PH, including a role for bone marrow and circulating progenitor cells and microribonucleic acids. Continued interest in the interface of the genetic basis of PH and cellular and molecular pathogenetic links should further expand our understanding of the disease.

Classical transient receptor potential channel 6 (TRPC6) is essential for hypoxic pulmonary vasoconstriction and alveolar gas exchange

Regional alveolar hypoxia causes local vasoconstriction in the lung, shifting blood flow from hypoxic to normoxic areas, thereby maintaining gas exchange. This mechanism is known as hypoxic pulmonary vasoconstriction (HPV). Disturbances in HPV can cause life-threatening hypoxemia whereas chronic hypoxia triggers lung vascular remodeling and pulmonary hypertension. The signaling cascade of this vitally important mechanism is still unresolved. Using transient receptor potential channel 6 (TRPC6)-deficient mice, we show that this channel is a key regulator of acute HPV as this regulatory mechanism was absent in TRPC6−/− mice whereas the pulmonary vasoconstrictor response to the thromboxane mimetic U46619 was unchanged. Accordingly, induction of regional hypoventilation resulted in severe arterial hypoxemia in TRPC6−/− but not in WT mice. This effect was mirrored by a lack of hypoxia-induced cation influx and currents in smooth-muscle cells from precapillary pulmonary arteries (PASMC) of TRPC6−/− mice. In both WT and TRPC6−/− PASMC hypoxia caused diacylglycerol (DAG) accumulation. DAG seems to exert its action via TRPC6, as DAG kinase inhibition provoked a cation influx only in WT but not in TRPC6−/− PASMC. Notably, chronic hypoxia-induced pulmonary hypertension was independent of TRPC6 activity. We conclude that TRPC6 plays a unique and indispensable role in acute hypoxic pulmonary vasoconstriction. Manipulation of TRPC6 function may thus offer a therapeutic strategy for the control of pulmonary hemodynamics and gas exchange.