Hepatic ischemia and reperfusion (I/R) injury is a complication of liver transplantation and resectional surgery, hemorrhagic and endotoxin shock and thermal injury. A growing body of experimental data suggests that I/R-induced liver damage occurs in a biphasic manner involving an early reactive oxygen specie (ROS)-dependent and a late, PMN-dependent phase. Recent evidence suggest that ROS and NO as well as certain pro-inflammatory cytokines (e.g. TNF-alpha, IL-1alpha, IL-12) play important roles in modulating post-ischemic liver injury. However, neither the specific cell types nor the sources for ROS and NO production nor their relationship to cytokine expression and hepatocellular injury have been explored. The overall objective of this proposal is to better understand the mechanisms by which ROS and NO-derived metabolites modulate tissue injury in a murine model of liver I/R injury in vivo. Hypothesis: We propose that the oxidative redox imbalance created by the overproduction of ROS and/or by the decreased bioavailability of NO in the post-ischemic liver activates nuclear transcription factor kB (NF-kB) thereby promoting the expression of both injurious proinflammatory cytokines (e.g. TNF-alpha, IL-1alpha, IL-12) as well as protective proteins (e.g. MnSOD, A20, B-c12, c-IAPs). We propose that it is an early imbalance between the expression of these NF-kB-dependent cytokines and protective proteins in favor of the former that initiates I/R-induced liver injury. We suggest that post-ischemic liver injury is limited by the upregulation of the NF-kB-dependent protective genes. In order to test this hypothesis, we intend to: a) Characterize the temporal relationship among reactive oxygen species production, NF-kB-dependent expression of injurious cytokines and protective proteins and cell injury using Kupffer cells, sinusoidal endothelial cells and hepatocytes isolated from control and post-ischemic livers, b) Determine the role of NF-kB in modulating I/R-induced liver injury, c) Investigate the role that reactive oxygen species play in regulating post-ischemic liver injury and d) Define the role that NO plays in modulating the ROS-dependent activation of NF-kB and subsequent expression of injurious and protective mediators in the post-ischemic liver. Understanding the mechanisms by which these pro-inflammatory cytokines are regulated in vivo may provide new insight into the pathogenesis of liver I/R injury and may lead to more rational approaches to the design of new drug therapies for the treatment of post-ischemic liver damage.
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