Complement C5 and early oxygen kinetics during murine sepsis.

Sepsis and septic shock are among the most life-threatening conditions managed by emergency physicians. The incidence of this condition in the United States has been estimated to be 751,000 cases per year, roughly three cases per 1,000 population.1 Many with the illness will die. Although traditionally deemed a disease of intensive care units, a recent prospective study has underscored the fundamental importance of early recognition and aggressive therapy of this syndrome in the emergency department (ED).2 A fundamental component of therapy in that trial was the early use of mixed-venous oxygen saturation to gauge oxygen kinetics in critically ill patients. Oxygen-related metabolic abnormalities are common in life-threatening infection and occur at multiple scales throughout the host.3,4 Infection prompts the proliferation of hematopoietic cell lines renowned for their respiratory burst. Activated neutrophils and monocytes/macrophages produce superoxide, hydrogen peroxide, hypochlorous acid, nitric oxide, and peroxynitrite, each consuming oxygen.5 Widespread endothelial injury leads to loss of capillary integrity and tissue edema, lengthening the diffusion path from red blood cells to the mitochondrial inner membrane of peripheral cells. Mitochondria display a number of abnormalities that have been reviewed extensively.6 At the organ level, oxygen delivery (DO2) may be impaired by anemia, myocardial depression, maldistribution of splanchnic blood flow, and widened ventilation–perfusion inequality in the acutely injured lung. At the broadest scale, sepsis impacts central homeostatic mechanisms regulating physical activity and the wake–sleep cycle, both of which are major determinants of a host’s oxygen consumption during acute illness.7 To further complicate matters, in many septic patients, a so-called hypermetabolic state may eventually occur, although differences in oxygen consumption between septic patients with and without hypermetabolic features are subtle (about twofold) compared with differences between septic patients and normal humans performing daily activities such as exercise (about 12-fold).8 The net abnormality under these circumstances is usually decreased oxygen content in blood returning to the heart from distal vascular beds. As the utility of monitoring mixed or central venous oxygen content and devising therapies capable of improving it undergoes intense scrutiny in clinical trials, much remains unknown about the events leading to abnormal oxygen kinetics. It has long been recognized that lipopolysaccharide (LPS, an important structural constituent of the outer leaflet of the outer membrane of gram-negative bacteria) and, to a lesser extent, lipoteichoic acid (a similar part of the gram-positive cell wall) can precipitate many of the clinical features of sepsis in the absence of actual infection.9,10 These and other molecules set into motion a cascade of events that ultimately produce the clinical phenomenon of sepsis, including impaired oxygen kinetics. Yet the intermediate steps in this disease pathway remain obscure. In the current study we explored the relationship between impaired oxygen kinetics and life-threatening infection using a mouse model of polymicrobial abdominal sepsis, namely, cecal ligation and puncture (CLP). Specifically, we studied the contribution of the complement cascade as a link in the development of abnormal oxygen kinetics. The complement cascade and, in particular, the mannose-binding lectin and alternative pathways are constitutively expressed and activated within seconds of exposure to gram-negative or gram-positive bacteria. Central features of complement activation in this setting are rapid opsonization of bacterial surfaces by activated C3 (needed for optimal phagocytosis by neutrophils and macrophages), assembly within the bacterial cell wall of the membrane attack complex (MAC), which, in some instances, may be directly bactericidal, and release of small peptides (C5a and C3a) that serve as potent activators of cellular immunity. The role of complement in sepsis has been described in several recent reviews.11,12 Although many proteins contribute to complement activation, C5 occupies a pivotal position. Cleavage of C5 produces C5b, which initiates assembly of bactericidal MAC, and C5a. The role of C5a in life-threatening infection is complex. In the setting of gram-negative pneumonia, failure to generate C5a or the absence of the C5a receptor leads to an abnormal inflammatory response and increased mortality.13,14 But in models of abdominal sepsis, C5a production may be so robust as to harm the host.15 Widespread, dysregulated complement activation has been documented to produce deleterious effects on oxygen-dependent systems at the subcellular, cellular, and organ levels. Intravascular complement activation causes hypotension, metabolic acidosis, and loss of endothelial integrity in the lung and the gut.16 Strategies that block C5a’s effects have been shown to preserve neutrophil function, correct coagulation abnormalities, and improve survival in polymicrobial sepsis.15,17–19 The specific contribution of complement, or to the best of our knowledge, any innate immune pathway, to oxygen kinetics during sepsis has not been demonstrated, although it is reasonable to suspect a connection. A link between immunologic events and clinical phenomena such as altered oxygen demand may lead to new therapies or generate further enthusiasm for strategies under study (such as anti-C5a treatment). In this study we examined the relationship between C5 and abnormal oxygen kinetics following CLP in mice. Using an open-circuit calorimeter, we characterized oxygen consumption, carbon dioxide production, and the respiratory exchange ratio (VO2, VCO2, and RER, respectively) after CLP and correlated changes with seven-day mortality. Next, the role of C5 in these metabolic changes was considered. We argued that if C5 was involved, a distinct oxygen kinetics phenotype might be observed in animals genetically deficient in C5. Specifically, if C5 linked peritoneal contamination with the development of oxygenation abnormalities, C5-deficient animals might demonstrate less severe abnormalities. Furthermore, an argument in support of a role of C5 would be strengthened if, in the absence of fecal contamination of the abdomen, sterile intraperitoneal complement activation reproduced the oxygen kinetic abnormalities seen in sepsis. Lastly, to specifically address differential contributions of C5a and C5b, we also studied animals in which a function-blocking antibody directed against C5a had been administered at the time of injury.