SUMMARYGenetically determined responsiveness to microbial stimuli such as lipopolysaccharide (LPS) may affect the pathophysiology of human sepsis. The D299G mutation in human Toll-like receptor-4 (TLR4) impairs LPS signalling in homozygous and heterozygous individuals. To investigate whether the presence of the TLR4(D299G) mutation may correlate with the development or outcome of sepsis following major visceral surgery the presence of TLR4(D299G) mutation was analysed in 307 Caucasian patients (154 without and 153 with sepsis). Sepsis was caused in 84% of patients by polymicrobial infection. The presence of the mutant TLR4 did not signi®cantly correlate with development or outcome of sepsis. Serum levels of tumour necrosis factor, interleukin (IL)-10, and IL-6 at sepsis onset did not signi®cantly differ between patients carrying wild-type and mutant TLR4. Moreover, studies in a murine model of polymicrobial septic peritonitis demonstrated that TLR4-de®ciency did neither in¯uence the systemic cytokine response nor the development of organ injury. The results suggest that the signalling capacity of TLR4 as affected by loss-offunction mutations does not in¯uence human or experimental sepsis caused by polymicrobial infection. Thus, in polymicrobial infection, other innate immune receptors may compensate for TLR4 defects.

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Intra-abdominal infection in patients following major visceral surgery is associated with high mortality. Using a macrophage depletion technique, we demonstrate that in murine septic peritonitis, Kupffer cells are a major source of systemic IL-10 levels. Kupffer cell-depleted mice were highly susceptible to the lethal effects of septic peritonitis and exhibited an increased bacterial load. Kupffer cell-depleted mice were protected by the administration of an IL-10-Fc fusion protein. Loss of Kupffer cell-derived IL-10 was associated with a weak increase in serum IL-12 levels, whereas TNF, IL-1α, and IL-18 levels were not significantly elevated, suggesting that the loss of Kupffer cell-derived IL-10 did not result in a toxic cytokine release syndrome. Instead, loss of Kupffer cell-derived IL-10 was associated with a reduced splenocyte production of IFN-γ that is required for immune protection in murine septic peritonitis. Therefore, the results suggest that the protective function of IL-10 in septic peritonitis may not be restricted to the anti-inflammatory activities of IL-10.

Sequestration of neutrophils and release of histotoxic mediators are considered important for the development of pathologic alterations of the lung defined as adult respiratory distress syndrome. Mechanisms of inflammatory lung injury caused by abdominal sepsis were investigated using the colon ascendens stent peritonitis (CASP) model that closely mimics the human disease. In the CASP model, a continuous leakage of intraluminal bacteria into the peritoneal cavity is induced by implantation of a stent in the ascending colon, generating a septic focus. In contrast to the cecal ligation and puncture model of peritonitis, survival of mice following CASP surgery is dependent on IFN-gamma, but independent of tumor necrosis factor (TNF). Here we show that the systemic inflammation induced by CASP surgery results in a rapid and profound increase of lung vascular permeability that was associated with the activation and recruitment of neutrophils to the lung. Activation of circulating granulocytes was characterized by increased production of serine proteinases and reactive oxygen metabolites, as well as elevated expression of cell surface Mac-1. Expression of MIP-2, KC, MIP-1alpha and E-selectin mRNA in lung was strongly increased within 3 h following CASP surgery, whereas up-regulation of IP-10, MCP-1 and P-selectin was delayed. In contrast, induction of RANTES, LIX, ICAM-1 and VCAM-1 mRNA was weak or not detectable after CASP surgery. Importantly, recruitment of leukocytes to the lung was normal in lipopolysaccharide-resistant mice, and was not affected by antibody neutralization of TNF or the chemokines MIP-2 and KC.

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Systemic levels of the key immune regulatory cytokines IL-12 and IL-18 were measured over time in 66 patients with postoperative sepsis (38 survivors and 28 nonsurvivors). Sepsis mortality was not significantly associated with any of the clinical parameters examined, including age, gender, underlying disease, and surgical procedure. Analysis of cytokine levels showed that during the entire observation period, IL-12 was significantly reduced in sepsis patients compared with control surgical patients without sepsis. IL-12 serum levels did not significantly differ between sepsis survivors and nonsurvivors. In contrast to IL-12, IL-18 serum levels were significantly higher in both surviving and nonsurviving sepsis patients than in controls. Importantly, we also observed that IL-18 levels were significantly increased in patients with lethal sepsis compared with sepsis survivors at all time points studied, including day 1 after sepsis diagnosis. IL-18 levels were significantly increased during the course of lethal sepsis, but remained at a comparable level in sepsis survivors. Logistic regression analysis of IL-18 values measured on days 1 or 2 of sepsis revealed that high serum IL-18 represents an early predictive factor for the lethal outcome of postoperative sepsis. Consistent with previous work in mouse models, our results suggest that IL-12 may contribute to protective immune reactions against a septic challenge, whereas IL-18 may preferentially promote organ injury and lethal shock.

High systemic CGRP levels were associated with lethal outcome already at the onset of sepsis, whereas high substance P levels were identified as late predictive indicators of lethal outcome. These results are consistent with the view that the neuropeptides, CGRP and substance P, may be involved in the pathogenesis of sepsis.

The innate immune system provides essential information about the presence of infectious danger and signals the activation and instruction of adaptive immunity. The present study addressed thequestion of whether prior exposure of the innate immune system to LPS may modulate host defense against acute septic peritonitis. We show that LPS priming 4 days, but not 2 days, prior to infectionenhances bacterial clearance and improves survival of septic peritonitis. Immune protection in day 4 LPS‐primed mice was specifically associated with a marked increase in the accumulation and activation of neutrophils at the site of infection. Accumulating neutrophils in day 4 LPS‐primed mice exhibited a normal production of reactive oxygen metabolites in response to in vivo exposureto intestinal bacteria. The local increase in neutrophil numbers was found to result from a reduced rate of apoptotic cell death. Inhibition of neutrophil apoptosis in LPS‐primed mice was mediated by soluble factor(s) distinct from G‐CSF and GM‐CSF. Thus, engagement of pattern recognition systems prior to infection may improve host defense by amplifying the effector cell response of innate immunity. The results also provide in vivo evidence that apoptosis of inflammatory cells represents an important process for the control of host defense to infection.