Original Contribution POLYNITROXYL-ALBUMIN (PNA) PLUS TEMPOL ATTENUATE LUNG CAPILLARY LEAK ELICITED BY PROLONGED INTESTINAL ISCHEMIA AND REPERFUSION SHU ZHANG,* HAIQUAN LI, † LI MA, ‡ CHARLES E. TRIMBLE, ‡ PERIANNAN KUPPUSAMY, † CARLETON J. C. HSIA, ‡ and DONNA L. CARDEN* *Departments of Internal Medicine and Physiology and Biophysics, Louisiana State University Medical Center, Shreveport, LA, USA; † Division of Cardiology and the EPR Center, Johns Hopkins Medical Institutions, Baltimore, MD, USA; and ‡ SynZyme Technologies LLC, Irvine, CA, USA (Received 27 January 2000; Revised 13 April 2000; Accepted 20 April 2000) Abstract—Stable nitroxyl radicals (nitroxides) are potential antioxidant drugs, and we have previously reported that linking nitroxide to biological macromolecules can improve therapeutic activity in at least two ways. First, polyni- troxylated compounds such as polynitroxyl human serum albumin (PNA) are a novel class of high molecular weight, extracellular antioxidants. Second, compounds such as PNA can prolong the half-life of free (unbound, low molecular weight) nitroxides such as 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (Tempol) in vivo. Unlike PNA, Tempol can readily access the intracellular compartment. Thus PNA can act alone in the extracellular compartment, or in concert with Tempol, to provide additional antioxidant protection within cells. In this study, we compared the abilities of PNA, Tempol, and the combination of PNA + Tempol to prevent lung microvascular injury secondary to prolonged gut ischemia (I, 120 min) and reperfusion (R, 20 min) in the rat. Pulmonary capillary filtration coefficient (K f,c ) and lung neutrophil retention (tissue myeloperoxidase activity, MPO) were measured in normal, isolated rat lungs perfused with blood harvested from I/R rats. Blood donor rats were treated with drug during ischemia. Gut I/R resulted in a marked increase in pulmonary capillary coefficient and lung MPO. PNA + Tempol, but not PNA alone or Tempol alone, at the doses used, prevented the development of lung leak. None of the treatments had an effect on lung neutrophil retention. Anti-inflammatory therapeutic activity appeared to correlate with blood Tempol level: in the presence of PNA, blood Tempol levels were maintained in the 50 –100 M range vs. essentially undetectable levels shortly after Tempol was administered alone. In this model of lung injury secondary to prolonged gut I/R, lung capillary leak was prevented when the membrane-permeable compound Tempol was maintained in its active, free radical state by PNA. © 2000 Elsevier Science Inc. Keywords—ARDS, Capillary permeability, Nitroxide, Polynitroxyl albumin, Intestinal ischemia/reperfusion, Free radicals INTRODUCTION Adult respiratory distress syndrome (ARDS) is charac- terized by the acute onset of hypoxemia and bilateral lung infiltrates, and occurs as a serious complication of lung injury or systemic insults including shock, trauma, burn, and sepsis. ARDS is associated with high mortal- ity. However, death in ARDS patients is generally not due to respiratory insufficiency, but rather tends to result from multiple organ failure. Therefore, it seems likely that ARDS is the pulmonary manifestation of a systemic inflammatory syndrome, which includes widespread in- jury to the vascular endothelium [1]. For this reason, and because ARDS remains difficult to treat, the ideal ap- proach may be to prevent ARDS in at-risk patients [2]. In this view, successful prevention of ARDS may also prevent multiple organ failure. The development of lung capillary leak in ARDS can be blocked at a number of points. For example, inhibition of complement activation [3], immunoneutralization of This work was presented in part at the Fourth International Shock Congress, Philadelphia, Pennsylvania, June 12-16, 1999 (Shock, Vol. 11 (Suppl.), Abstract #62). Address correspondence to: Dr. Carleton J. C. Hsia, SynZyme Tech- nologies LLC, 1 Technology Drive, Suite E-309, Irvine, CA 92630, USA; Tel: (949) 453-1072; Fax: (949) 453-1074; E-Mail: synzyme@ synzyme.com. Free Radical Biology & Medicine, Vol. 29, No. 1, pp. 42–50, 2000 Copyright © 2000 Elsevier Science Inc. Printed in the USA. All rights reserved 0891-5849/00/$–see front matter PII S0891-5849(00)00295-1 42