CYTOKINE REGULATION IN SEPSIS AND INFLAMMATION

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Our knowledge of the role that cytokines play in both the beneficial and pathologic sequelae to surgical injury and bacterial infection has greatly increased. In septic shock secondary to gram-negative bacteria, excessive tumor necrosis factor alpha (TNFalpha), and interleukin-1 (IL- 1) production contributes to the mortality and tissue damages that occur. A critical advance in the past two years has been the discovery and identification of at least two new classes of specific endogenous cytokine inhibitors, IL-1 receptor antagonist (IL-1ra) and the soluble TNF receptors (sTNFR's) which inhibit IL-1 and TNF bioactivity and circulate in the plasma of critically-ill patients. The fundamental premise of this application is that the successful host response to surgical injury and infection requires not only the early release of proinflammatory cytokines, but also the production of natural antagonists of proinflammatory cytokine action (IL-1ra), and the release of soluble TNF receptors (TNF receptors I [p55] and II [p75]), which serve to limit IL-1 and TNF's actions. Furthermore, simultaneous, production of cytokines with anti-inflammatory action (IL-10) restricts continued IL-1 and TNFalpha production in an autocrine manner, and up-regulates the production of IL-1ra The catastrophic host responses to septic shock, and the continued systemic inflammatory response syndrome, are the result of an imbalance among pro- and anti-inflammatory cytokine, and cytokine inhibitor production. The specific aims of this application are: 1) to describe the local production of pro- and anti-inflammatory cytokines and cytokine inhibitors in lethal and nonlethal infection models, and 2) to better understand the integrated mechanisms by which pro- and anti- inflammatory cytokines and cytokine inhibitors regulate tissue responses. Using murine models of lethal and nonlethal inflammation (E. coli bacteremia, cecal ligation and puncture or a sterile turpentine abscess), the regulation of tissue IL-1, TNFalpha, IL-4, IL-4, IL-10, IL-1ra and STNFR production will be determined. In mice, cytokine MRNA will be quantitated using the reverse transcriptase-polymerase chain reaction method, and cytokine protein will be assessed by both immunoassay (ELISA) and bioassay (IL-1, TNFalpha), where appropriate. In Papio (baboons), the systemic cytokine and cytokine inhibitor responses to lethal E. coli bacteremia, sublethal endotoxemia and pro-inflammatory cytokine infusions (IL-1alpha, TNFalpha and IL-8) will also be estimated by flow cytometry. To identify host responses which are dependent upon IL-1 and TNFalpha binding to their type I and type II receptors, specific monoclonal antibodies against the type I or type II IL-1 receptor will be administered to infected animals, or TNFalpha mutants that bind only to specific classes of TNF receptors will be administered to healthy baboons. In addition, the role that cytokine inhibitors (IL-1ra) and anti-inflammatory cytokines (IL-4, IL-10) play in the host response in these models will be delineated by the use of neutralizing monoclonal antibodies and recombinant protein. Such studies will strengthen our fundamental understanding of the metabolic response to injury and infection, and will provide a sound theoretical basis for future attempts to modulate the host response to injury and infection.