Regulation Of IL-8 Production By Complement Activated Product, C5a, During Inflammation Induced By Bacterial Infection

Infectious inflammation is comprehensive defense of non-specific response which is induced by bacterial invasion when the body is injured. The continued development of infectious inflammation can make immune system out of control. A large number of toxins from bacterium induce immune system to incompetent state and increase tissue damage following by sepsis. In western countries about 150 million people suffer sepsis, the mortality is 30%～50%.Complement system is a precise protein system which is composed of over 30 proteins and protein fragments including serum proteins, serosal proteins, and cell membrane receptors. Complement system is the first line of host defense to combat invading microbes and infectious agents, and excessive complement activation had been shown in septic patients.Among the complement-activated products, C5a is the most potent chemotactic factor with broad biological functions such as chemotactic response of neutrophils, release of granule enzymes from phagocytic cells, modulation of cytokines such as IL-6, vasodilators in the canine coronary vasculature, inhibition of neutrophil apoptosis during sepsis in vivo. Adhesion molecule expression and coagulation pathway are also affected by C5a. It has been demonstrated that overtly activated complement system, especially complement-activated product C5a, induced disorders in innate immune functions such as neutrophil dysfunction, platelet activation, and direct or indirect microvascular damage. Accumulating evidence showed beneficial effects for blockage of C5a or C5aR (CD88) in experimental sepsis.IL-8 is a cytokine possessing a strong chemoattractant activity and is able to activate and degranulate neutrophils. Due to its neutophil-activating properties, IL-8 appears to play an important role in the pathophysiology of sepsis. Hack et.al had demonstrated that IL-8 level is elevated in patients'serum in sepsis. It has been shown that IL-8 can be produced by a variety of cells, including blood monocytes, macrophages, and endothelial cells. In the disease setting, IL-8 may lead to tissue damage by chemoattracting a large number of neutrophils to inflammatory location. In a mouse model of sepsis, an early survival benefit was also found when the mouse IL-8 homologue, KC, was blocked, indicative of the essential role of IL-8 in the pathogenesis of sepsis development. It is of great significance to explore the regulation mechanism of IL-8.One of pathological effects is large mount of cytokines induced by C5a during infectious inflammation. More and more data showed that C5a played an important role in regulating inflammatory cytokines such as IL-6, IL-17, and TNFα. Regulatory role of C5a in IL-8 generation during infectious inflammation and the potential mechanism involved is what we are interested in.Herein, we employed in vitro and in vivo models of sepsis to investigate the functional relationship between overtly produced C5a and IL-8. Our research is divided into 4 parts: a) co-stimulatory effect and mechanism of exogenous C5a on IL-8 production in vitro; b) role of C5a in increasing IL-8 production induced by other TLR agonists; c) co-stimulatory effect and mechanism of endogenous C5a on IL-8 production in vitro; d) moderating effect of overexpressed C5a on mouse IL-8 production in mouse sepsis model.Our data revealed that C5a could strongly amplify IL-8 expression from human whole blood cells induced by LPS and other types of TLR agonists. ERK1/2 and p38, but not JNK, were mainly participated in signaling pathways for IL-8 production. In the whole blood stimulated by E.coli, C5a levels were quickly elevated and blockage of C5a significantly decreased E.coli-elicited IL-8 production. In the mouse model of sepsis induced by Cecal Ligation and Puncture (CLP), the markedly increased Keratinocyte-derived Cytokine (KC) could be strongly suppressed by blockage of C5a. These data suggest that excessive C5a functions as a critical inflammatory mediator to enhance IL-8 production mainly through MAPK signaling pathways.In the current study, we sought to determine the regulatory role of C5a in IL-8 generation in the setting of sepsis and the potential mechanism involved. In vitro we employed human whole blood cells to evaluate the contribution of C5a to IL-8 production induced by LPS and E.coli K12. In addition, an in vivo sepsis model induced by CLP was used to determine the functional relationship between C5a and IL-8 expression in vivoOur data indicate that C5a is a universal amplifier for IL-8 production which is an effector duing infectous inflammation. These findings would strengthen the understating of C5a biological functions and the mechanism by which blockage of C5a renders the therapeutic benefit in sepsis.