Although studies have shown increased evidence of death receptor-driven apoptosis in intestinal lymphoid cells, splenocytes, and the liver following the onset of polymicrobial sepsis, little is known about the mediators controlling this process or their pathologic contribution. We therefore attempted to test the hypothesis that the hydrodynamic administration of small interfering RNA (siRNA) against the death receptor, Fas or caspase-8, should attenuate the onset of morbidity and mortality seen in sepsis, as produced by cecal ligation and puncture (CLP). We initially show that in vivo administration of green fluorescent protein (GFP) siRNA in GFP transgenic mice results in a decrease in GFP fluorescence in most tissues. Subsequently, we also found that treating septic nontransgenic mice with siRNA targeting Fas or caspase-8 but not GFP (used as a control here) decreased the mRNA, in a sustained fashion up to 10 days, and protein expression of Fas and caspase-8, respectively. In addition, transferasemediated dUTP (deoxyuridine triphosphate) nick end labeling (TUNEL) and active caspase-3 analyses revealed a decrease in apoptosis in the liver and spleen but not the thymus following siRNA treatment. Indices of liver damage were also decreased. Finally, the injection of Fas or caspase-8 given not only 30 minutes but up to 12 hours after CLP significantly improved the survival of septic mice. IntroductionSepsis affects approximately 750 000 people in the United States every year, and one third of the reported cases result in death. 1 Common causes include traumatic injury, severe bacterial infections, or severe burns. However, sepsis also frequently affects critically ill, elderly, pediatric, and postsurgical patients in the intensive care unit. With the exception of the recent application of activated protein C, most molecular-biologic-based therapies have failed clinically. Treatment using antibiotics mildly reduces risk of death; however, it is ineffective on mice with severe sepsis, as measured by high inflammatory cytokine levels. 2 Thus, there is an urgent need not only for better understanding of the pathology of sepsis and its resultant organ failure, but also for new therapeutic approaches and targets.We and other laboratories have previously reported that organ damage and mortality associated with sepsis in mouse models is at least in part due to the activation of the Fas-FasL (Fas ligand) signaling pathway, and not Toll-like receptor 4 (TLR4). 3 Studies using Fas fusion protein given 12 hours after cecal ligation and puncture (CLP) show a protective effect against apoptosis in Kupffer cells, which seems to benefit the liver in a way that reduces organ damage and, in turn, improves survival after sepsis. 4 This blockade of FasL also restores total hepatic, intestinal, and cardiac blood flow while attenuating the plasma levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), indicating reduced liver damage. 5 In addition, studies using FasL Ϫ/Ϫ mice provide further evidence for death r...

Recent research has yielded many interesting and potentially important therapeutic targets in sepsis. Specifically, the effects of antagonistic anti-cytokine therapies (tumor necrosis factor-alpha [TNF-α], interleukin-1 [IL-1]) and anti-endotoxin strategies utilizing antibodies against endotoxin or endotoxin receptor/carrier molecules (anti-CD14 or anti-LPS-binding protein) have been studied. Unfortunately, these approaches often failed clinically, and in many cases, the efficacy of these treatments was dependent on the severity of sepsis. Recently, clinical trials using insulin to lock blood glucose levels and activated protein C treatment have showed that while they provided some survival benefit, their efficacy does not appear to be predicated solely upon anti-inflammatory effects. Here, we will review work done in animal models of polymicrobial sepsis and clinical findings that support the hypothesis that apoptosis in the immune system is a pathologic event in sepsis that can be a therapeutic target. In this respect, experimental studies looking at the septic animal suggest that loss of lymphocytes during sepsis may be due to dysregulated apoptosis and that this appears to be brought on by a variety of mediators effecting 'intrinsic' as well as 'extrinsic' cell death pathways. From a therapeutic perspective this has provided a number of novel targets for clinically successful current, as well as future therapies, such as caspases (caspase inhibition/protease inhibition), pro-apoptotic protein-expression (via administration and/or over-expression of Bcl-2) and the death receptor family Fas-FasL (via. FasFP [fas fusion protein] and the application of siRNA against a number proapoptotic factors).

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