86 SURGERY APOPTOSIS PROVIDES A PHYSIOLOGIC MEANS for the clearance of tissue neutrophils (PMNs) and resolu- tion of inflammatory processes. 1,2 Conversely, dys- functional (delayed) PMN apoptosis has been implicated in numerous hyperinflammatory dis- ease states, including smoke inhalation injury, 3 the acute respiratory distress syndrome, 4 and the sys- temic inflammatory response syndrome. 5 Dysfunc- tional PMN apoptosis may also be involved in the pathogenesis of postinjury multiple organ failure (MOF). We 6 and others 7-9 have reported that trau- ma patients at risk for MOF have delayed PMN apoptosis in the early postinjury period. Trauma patients’ plasma delays the apoptosis of healthy donors’ PMNs, indicating the presence of circulat- ing antiapoptotic mediators. 6-8 We have recently implicated platelet-activating factor (PAF) and PAF-like phospholipids, including lyso-phos- phatidylcholine (LPC) as the key circulating anti- apoptotic mediators in trauma patients’ plasma. 10 Our previous epidemiologic studies identified red blood cell (RBC) transfusion as the most robust Clinically relevant hypertonicity prevents stored blood- and lipid-mediated delayed neutrophil apoptosis independent of p38 MAPK or caspase-3 activation Walter L. Biffl, MD, Rachel Carnaggio, MS, Ernest E. Moore, MD, David J. Ciesla, MD, Jeffrey L. Johnson, MD, and Christopher C. Silliman, MD, PhD, Providence, RI, and Denver, Colo Background. Delayed apoptosis of primed neutrophils (PMNs) may facilitate PMN-mediated tissue injury leading to postinjury multiple organ failure. Aged (42-day-old) stored red blood cells (RBC42) delay PMN apoptosis through proinflammatory phospholipids such as platelet-activating factor (PAF) and lyso-phosphatidylcholine (LPC). Hypertonic saline (HTS) attenuates PMN cytotoxic functions. We hypothesized that clinically relevant HTS would provoke PMN apoptosis, as well as prevent stored blood- and lipid-mediated delayed PMN apoptosis through activation of p38 mitogen-activated protein kinase (MAPK) and caspase-3. Methods. PMNs harvested from healthy volunteers were incubated (5% CO 2 , 37° C, 24 hr) with RBC42 plasma, PAF (20 μM), or LPC (4.5 μM), with or without the p38 MAPK inhibitor SB 203580, the caspase-3 inhibitor zDEVD-fmk (10 μmol/L) or the pan-caspase inhibitor zVAD-fmk (20 μmol/L). Duplicate samples were preincubated in HTS (Na [180 mM]). Apoptotic index (% PMNs undergoing apoptosis) was assessed morphologically. p38 MAPK activation was assessed by Western blotting. Caspase-3 activity was measured colorimetrically. Results. PAF, LPC, and RBC42 plasma delayed apoptosis; HTS increased apoptosis compared with con- trols. HTS prevented PAF, LPC, and RBC42-delayed apoptosis. p38 MAPK was not activated by HTS; its inhibition had no effect on the actions of HTS. Caspase inhibition attenuated the ability of HTS to increase apoptosis, but it did not affect the ability of HTS to restore healthy PMN apoptosis in the pres- ence of RBC42. Conclusion. HTS increases PMN apoptosis and prevents stored blood- and lipid-mediated delayed PMN apoptosis. HTS may activate caspase-3, but alternative signaling pathways appear to be involved in modulating the effects of lipids on PMN apoptosis. (Surgery 2003;134:86-91.) From the Departments of Surgery, Rhode Island Hospital/Brown Medical School, Providence, RI, and the Den- ver Health Medical Center, and Bonfils Blood Center and Department of Pediatrics, University of Colorado Health Sciences Center, Denver, Colo Accepted for publication March 23, 2003. Presented at the 1 st Joint Meeting of the Surgical Infection Soci- ety (22 nd Annual Meeting) and Surgical Infection Society- Europe (15 th Annual Meeting), Madrid, Spain, May, 2002. Supported in part by The Society of University Surgeons Foun- dation Junior Faculty Fellowship Award (WLB), a National Blood Foundation Scientific Research Grant (WLB), and National Institutes of Health grants P50GM4922 and T32GM08315 (EEM, WLB, CCS). Reprint Requests: Walter L. Biffl, MD, Division of Trauma and Surgical Critical Care, 593 Eddy Street, APC 110, Providence, RI 02903. © 2003 Mosby, Inc. All rights reserved. 0039-6060/2003/$30.00 + 0 doi:10:1067/msy.2003.178