0031-3998/06/5901-0147 PEDIATRIC RESEARCH Vol. 59, No. 1, 2006 Copyright © 2005 International Pediatric Research Foundation, Inc. Printed in U.S.A. Tezosentan Decreases Pulmonary Artery Pressure and Improves Survival Rate in an Animal Model of Meconium Aspiration RALF GEIGER, WERNER PAJK, NIKOLAUS NEU, STEPHAN MAIER, AXEL KLEINSASSER, SOHRAB FRATZ, SALVADOR NAVARRO-PSIHA, VIKTORIA FISCHER, BENEDIKT TREML, AND ALEXANDER LOECKINGER Clinical Divisions of Pediatric Cardiology [R.G., V.F.], Pediatric Intensive Care [N.N.], Neonatology [S.N.], Anesthesiology and Critical Care Medicine [W.P., S.M., A.K., B.T., A.L.], Innsbruck Medical University, 6020 Innsbruck, Austria; Department of Pediatric Cardiology and Congenital Heart Disease [S.F.], German Heart Center Munich, 80636 Munich, Germany ABSTRACT: Acute pulmonary arterial hypertension in acute lung prostacyclin, along with increased production of vasoconstric- injury aggravates the clinical course and complicates treatment. tive endothelin-1 (ET-1) are regarded as key factors in the Increased release and turnover of endogenous endothelin-1 is known pathophysiology of pulmonary hypertensive disorders (9). to be a major determinant in the pathophysiology of pulmonary ET-1 is a polypeptide, which is released by vascular endothe- arterial hypertension of various etiologies. We tested whether intra- lial cells in response to hypoxic and acute or chronic toxic venous tezosentan, a dual endothelin receptor antagonist, reduced lung injury (10,11). Induction of transcription of ET-1 mRNA pulmonary artery pressure in a pig model of acute lung injury induced by meconium aspiration. Acute pulmonary arterial hyper- and synthesis and secretion of ET-1 takes place within min- tension was induced in 12 anesthetized and instrumented pigs by utes after promoting stimuli by, for example, hypoxia, cyto- instillation of human pooled meconium in a 20% solution. Hemody- kines, adhesion molecules, and catecholamines (12). ET-1 namic and gas exchange parameters were recorded every 30 min. Six exerts a direct and sustained vasoconstrictive effect via ETA animals received tezosentan 5 mg/kg after 0 and 90 min; six animals receptors, expressed on vascular smooth muscle cells. Acti- served as controls. Tezosentan led to a decrease of mean pulmonary vation of ETB receptors, expressed on vascular endothelial artery pressure (PAP) from 33.4 Ϯ 4.0 mm Hg to 24.7 Ϯ 2.1 mm Hg cells, mediates pulmonary endothelin clearance and induces and pulmonary vascular resistance (PVR) from 7.8 Ϯ 1.4 mm Hg · production of nitric oxide and prostacyclin, which both exert –1 2 Ϯ –1 2 L · min · m to 5.2 0.7 mm Hg · L · min · m . All animals vasodilative effects. In addition, ET-1 stimulates proliferation treated with tezosentan survived, whereas in the control group four of vascular smooth muscle cells and acts as a proinflammatory out of six animals died. Tezosentan improved survival and decreased mediator (13). Indeed, up-regulation of ET-1 gene expression pulmonary artery pressure in a porcine model of acute pulmonary arterial hypertension after meconium aspiration. Tezosentan has the and increased levels of circulating ET-1 have been demon- potential for effective pharmacological treatment of pulmonary arte- strated in animal models of meconium aspiration (14,15) and rial hypertension following acute lung injury. (Pediatr Res 59: in hypoxia-induced pulmonary arterial hypertension (16). 147–150, 2006) Therefore, blocking the effects of ET-1 should attenuate the concomitant rise of pulmonary artery pressure in acute lung injury and thus diminish stress imposed on the right heart. The cute pulmonary arterial hypertension accompanies acute orally active combined ETA and ETB receptor antagonist A lung injury (ALI) and acute respiratory distress syn- bosentan has been shown to be of benefit in the treatment of drome. Aspiration of meconium induces acute parenchymal pulmonary arterial hypertension in humans (17,18). In the lung disease with diffuse inflammation of the alveolar- clinical situation of profound hemodynamic impairment after capillary membrane. Obliteration of the pulmonary capillary ALI, where pharmacodynamics might be influenced by vari- bed and subsequent pulmonary vasoconstriction induces pul- able enteral drug uptake, an injectable agent would be of monary arterial hypertension, which may be severe and may advantage. lead to right ventricular failure and subsequent multiorgan Recently, a water-soluble combined ETA and ETB receptor failure (1–4). Effective pharmacological therapy is limited. antagonist for parenteral use, tezosentan (Actelion Pharma- Endothelial dysfunction of the small pulmonary arteries is ceuticals, Allschwil, Switzerland), has been shown to exert known to play a key role in the pathophysiology of pulmonary pulmonary vasodilative effects in oleic acid–induced pulmo- arterial hypertension of various etiologies (5–8). Impaired nary arterial hypertension in dogs and in lambs with acute and production of vasodilative mediators, such as nitric oxide and chronic pulmonary arterial hypertension (19,20). The aim of Received March 17, 2005; accepted July 14, 2005. Abbreviations: CVP, central venous pressure; FiO2, inspired oxygen frac- Correspondence: Ralf Geiger, M.D., Clinical Division of Pediatric Cardiology, Inns- tion; PAP, mean pulmonary artery pressure; PCWP, mean pulmonary cap- bruck Medical University, Anichstrasse 35, 6020 Innsbruck, Austria; e-mail: ralf.geiger@ uibk.ac.at illary wedge pressure; PVR, pulmonary vascular resistance; PVR/SVR, pul- This research was supported by Grant 110 from MFF/TILAK, General Hospital, monary to systemic vascular resistance ratio Innsbruck, Austria. Tezosentan was provided by Actelion Pharmaceuticals, Allschwil, Switzerland. DOI: 10.1203/01.pdr.0000191813.60977.bf 147 148 GEIGER ET AL. this study was to test the hypothesis whether tezosentan Respirator settings were not changed during the study period in none of the reduces mean PAP in a pig model of acute pulmonary arterial animals. At the end of the study, animals were killed using pentobarbital. Thereafter, the heart together with the proximal parts of the great arteries was hypertension secondary to meconium aspiration. removed and inspected to rule out any anatomical intra- or extracardial shunt connection. Pulmonary and vascular resistance was calculated using standard METHODS formulas. Values were related to body surface area by converting factor 0.0947 · kg2/3 (21). Experimental preparation. The Austrian Federal Animal Investigational Statistical analysis. A two-way ANOVA was used to determine inter- and Committee approved all experiments, and animals were managed in accor- intragroup differences. Significant results were analyzed post hoc with the Ϯ dance with the National Institutes of Health guidelines. First-pass human Newman-Keuls and Fisher’s exact tests. Data are presented as mean SD, a Ͻ meconium was obtained, suspended in physiologic sodium chloride at a value of p 0.05 was considered significant. concentration of 20%, filtered, and stored in 20 mL syringes at –20°C. The meconium was thawed in hot water immediately before use. The study was RESULTS performed on healthy, 4-wk-old white farm pigs weighing 17–23 kg. Anes- thesia was induced with ketamine (50 mg/kg i.m.) and atropine (0.01 mg/kg Changes from baseline to min 0. Meconium instillation led i.m.), followed by i.v. propofol (2–4 mg/kg). After the trachea had been to an increase of PAP from mean 19.8 Ϯ 2.0 mm Hg to 33.4 intubated, lungs were ventilated in volume-controlled mode (Evita 4, Dra¨ger, Ϯ 3.4 mm Hg (70%) and PVR from mean 3.8 Ϯ 1.0 mm Hg Telford, PA) at an FiO of 0.4 and a tidal volume (VT) of 10 mL/kg at 20 2 –1 2 Ϯ –1 2 breaths/min, positive end-expiratory pressure set at 5 mm Hg. VT was then ·L ·min·m to 8.1 1.6 mm Hg · L ·min·m (113%), adjusted to achieve a PaCO2 between 35 and 40 mm Hg, resulting in a minute without any significant intergroup differences (Table 1). ventilation of 150–170 mL/kg/min. Anesthesia was maintained with propofol Changes from min 0 to min 210. None of the animals of the (10–15 mg/kg/h) and piritramide boluses (15 mg each). Ringer’s solution (6 mL/kg/h) and a 3% gelatin solution (4 mL/kg/h) were administered through- tezosentan group died during this observation period in con- out the procedure. A standard lead II ECG was used to monitor cardiac trast to the control group, where four out of six animals died rhythm. Body temperature was maintained between 38°C and 39°C by using (p ϭ 0.03). Death had occurred before 60 min (two animals), an electric heating blanket. A 4F catheter was advanced into a femoral artery for withdrawal of arterial 90 min, and 180 min (one animal, respectively) (Fig. 1). blood and for measuring arterial blood pressure. A 7F pulmonary artery Immediately after bolus injection of tezosentan, animals had a catheter was advanced from the internal jugular vein into the pulmonary significant decrease of mean PAP (33.4 Ϯ 4.0 mm Hg to 24.7 artery to measure mean PAP, PCWP, CVP, and cardiac output by the Ϯ ϭ Ϯ thermodilution technique (10 mL saline in triplicate) and to withdraw mixed 2.1 mm Hg, p 0.001) and PVR (from 7.8 1.4 mm Hg venous blood. All catheters were filled with saline and connected to pressure ·L–1 ·min·m2 to 5.2 Ϯ 0.7 mm Hg · L –1 ·min·m2, p ϭ transducers zeroed to ambient pressure at the level of the right atrium. 0.0003). These effects were continual throughout the study Experimental protocol. Twelve animals where randomly assigned to receive either tezosentan or to serve as controls. After preparation of the period. The second dose of tezosentan did not significantly animals a stabilization phase of 30 min was allowed, thereafter baseline alter the parameters measured. In striking contrast, PAP and measurements (hemodynamics, blood gases) were taken.