The upper panels show representative lesions from an untreated rabbit (that received LPS but no antibody: LPS/0). the antibody is usually administered after initial exposures to LPS. In contrast, anti-rabbit tumor necrosis factor mAb treatment fails to protect when administered after LPS injections. These results support the concept that anti-CD14 treatment provides a new therapeutic windows for the prevention of pathophysiologic changes that result from cumulative exposures to LPS during septic shock in man. Gram-negative (GN) bacterial infections are associated with the development of septic shock, a serious clinical problem estimated to result in 100,000 deaths annually in the United States (1C3). Endotoxin (lipopolysaccharide, LPS), a glycolipid component of the outer membrane of all GN organisms, is generally Rabbit Polyclonal to VPS72 acknowledged to play a central role in the development of septic shock (4C6). This syndrome is characterized by hypotension, coagulopathy, organ failure, and death (1, 4, 7, 8). The prevailing view is that the underlying mechanisms responsible for these complex pathophysiologic changes involve cellular injury caused by an array of proinflammatory mediators released from cells of the innate immune system (9C12). The innate immune system has evolved mechanisms to recognize trace amounts of LPS via a cell surface receptor that initiates cell activation after binding LPS. Numerous studies provide support for the concept that two proteins, LPS-binding protein (LBP) and CD14 are essential for LPS-induced cellular activation (13). Mice with targeted deletions of the gene encoding either CD14 or LBP are resistant to the lethal effects of LPS supporting the contention that LBP and CD14 are essential elements of a nonredundant LPS recognition system (14C16). Effective approaches for the treatment of septic shock have lagged behind the substantive advances Nesbuvir made in understanding host mechanisms involved in the response to contamination (17). It is likely that the complexity of host responses in sepsis makes the relationship between the timing of delivery and the efficacy of potential therapeutics for septic shock a critical issue. For example, such considerations may serve to limit the effectiveness of treatments involving neutralization of single mediators such as tumor necrosis factor (TNF) or interleukin-1 that might only appear at certain stages in the sequence of events leading to septic shock (17). Here, we have investigated the effects of mAbs to either rabbit (rab) CD14 or rabbit TNF in a model of endotoxin Nesbuvir shock in rabbits in which three sequential exposures to LPS administered during a 24-hr period results in progressive injury marked by death between 24 and 48 hr. The characteristics of this model contrast significantly with acute lethal bolus injection models (18, 19). It is likely that this model used here more closely reflects the development of septic shock in patients occurring over time as a consequence of repeated exposures to LPS. Herein, we show that a mAb to rabbit CD14 that blocks LPS binding and subsequent cell activation offers remarkable protection even when administered after several exposures to LPS. In contrast, a mAb to rabbit TNF that neutralizes the biological activity of Nesbuvir this mediator fails to demonstrate the same degree of protection. These data suggest that therapy by using anti-CD14 monoclonal treatment in patients with GN septicemia and shock may be useful even after the patient has been uncovered LPS and in settings where Nesbuvir other therapies have failed to be efficacious. MATERIALS AND METHODS Experimental Model. A three-injection endotoxemic model was used as described by Mathison (20). In brief, all of the rabbits (= 108), except for the control group (= 15), received i.v. injections of Re595 LPS (5 g/kg) via the marginal ear vein at times 0, 5, and 24 hr. The rough form of LPS designated Re595 was purified from bacteria (20). Stock samples of Re595 at 5 mg/ml were kept frozen at ?20C. Just before an i.v. injection, an aliquot was thawed, sonicated, and diluted with 0.9% NaCl to a concentration of 10 g/ml. New Zealand White rabbits were obtained from Western Oregon Rabbit Company, Philomath, OR, and allowed 10 days for equilibration before use. Three days before the start of experiments, a 3.5 French polyurethane cannula (MRE-040, Braintree Scientific) with an accessible subcutaneous port (Jelco Intermittent injection cap, Critikon, Tampa, FL) was placed into the aortic arch via the left carotid artery by using aseptic technique and halothane anesthesia. Animals had free access to food and water and were unrestrained throughout the length of each study. Blood samples (1.3 ml total) were obtained, and mean arterial pressures (MAPs) were measured by using a model P10EZ transducer (Viggo-Spectramed, Oxnard, CA) at various time points throughout each 48-hr experiment. Blood was collected in.