108 - The Impact of Inflammatory Cytokines on Liver Damage Caused by Elevated Generation of Mitochondrial Reactive Oxygen Species

Abstract

Systemic inflammatory response (SIR) is a defense reaction of the body against pathogens, but it can also cause damage to host tissues. Recently we have shown that this damage can be effectively reduced by inhibition of mitochondrial reactive oxygen species (mtROS) generation. Specific aim of this study was to clarify whether or not acute phase inflammatory cytokines, such as TNF-α, directly induce mtROS-mediated liver damage. Sprague-Dawley rats were challenged with lipopolysaccharide (LPS, E. coli, 2.5 mg/kg i.v.) to elicit SIR. To examine the impact of mtROS, we treated animals with MitoTEMPO (mT), a mitochondria targeted ROS scavenger. At 2, 4, 8 and 16 h after LPS injection we determined blood cytokine levels and organ damage parameters. In addition, immune cells isolated from different body compartments were examined on their capacity to generate ROS. In an in vitro study hepatocytes were incubated with different patterns of inflammatory mediators to further elucidate effects acute and later phase mediators on mtROS production. LPS challenge resulted in an increase of inflammatory cytokines levels only at early phase (2 and 4 hours), while severe liver damage occurred only in the late phase (16 h) of SIR and was successfully ameliorated by mT treatment. In contrast, early elevation of IL-1 and TNF-α, as well as ROS production of polymorphonuclear (PMN) leukocytes were not affected by mT. To clarify whether acute or late phase cytokines induce mtROS mediated liver damage we incubated blood cells with LPS for 6, 12 and 24 hours to create acute (6h) and late phase (12, 24h) patterns of inflammatory mediators. Incubation of hepatocytes with these cytokine patterns showed that although acute phase cytokines were completely released within first 6 h they did not elevate hepatocyte mtROS levels, while 12 and 24h cytokine patterns substantially elevated mtROS levels. Our data suggest that acute phase cytokines do not directly induce liver damage, but facilitate the release of late phase inflammatory mediator(s) causing liver damage mediated by mtROS. This damage occurs due to intracellular signaling within hepatocytes rather than due to overwhelmed activation of immune cells.