Iron primes hepatic macrophages for NF-κB activation in alcoholic liver injury

Abstract

NF-kappaB activation induced by lipopolysaccharide (LPS) in cultured hepatic macrophages (HM) may be abrogated by pretreatment of cells with a lipophilic iron chelator, 1,2-dimethyl-3-hydroxypyrid-4-one (L1, deferiprone), suggesting a role for iron in this molecular event [M. Lin, M., R. A. Rippe, O. Niemelä, G. Brittenham, and H. Tsukamoto, Am. J. Physiol. 272 (Gastrointest. Liver Physiol. 35): G1355-G1364, 1997]. To ascertain the relevance in vivo of this hypothesis, HM from an experimental model of alcoholic liver injury were examined for the relationship between nuclear factor (NF)-kappaB activation and iron storage. HM showed a significant increase in nonheme iron concentration (+70%), accompanied by enhanced generation of electron paramagnetic resonance-detected radicals (+200%), NF-kappaB activation (+100%), and tumor necrosis factor-alpha (+150%) and macrophage inflammatory protein-1 (+280%) mRNA induction. Treatment of the cells ex vivo with L1 normalized all these parameters. HM content of ferritin protein, ferritin L chain mRNA, and hemeoxygenase-1 mRNA and splenic content of nonheme iron were increased, suggesting enhanced heme turnover as a cause of the increased iron storage and NF-kappaB activation. To test this possibility, increased iron content in HM was reproduced in vitro by phagocytosis of heat-treated red blood cells. Treatment caused a 40% increase in nonheme iron concentration and accentuated LPS-induced NF-kappaB activation twofold. Both effects could be abolished by pretreatment of cells with zinc protoporphyrin, a hemeoxygenase inhibitor. To extend this observation, animals were splenectomized before 9-wk alcohol feeding. Splenectomy resulted in further increments in HM nonheme iron storage (+60%) and NF-kappaB activation (+90%) and mononuclear cell infiltration (+450%), particularly around the iron-loaded HM in alcohol-fed animals. These results support the pivotal role of heme-derived iron in priming HM for NF-kappaB activation and expression of proinflammatory genes in alcoholic liver injury.

Signaling Role of Intracellular Iron in NF-κB Activation

“…Anti-human I B␣ and antibodies against p50 and p65 were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Iron 59 ( Cell Preparation-HMs were isolated from male Wistar rats by the Non-parenchymal Liver Cell Core of the Research Center for Alcoholic Liver and Pancreatic Diseases as previously published (24,25). Briefly, the liver was digested in situ by sequential perfusion with Pronase and collagenase, and non-parenchymal liver cells were fractionated by discontinuous gradient ultracentrifugation using arabinogalactan.…”

“…We have previously reported an opposite example of an enhanced chelatable pool of iron in HMs. In these HMs, which were isolated from a rat model of alcoholic liver disease, the expanded chelatable pool of iron was tightly associated with increased NF-B binding and TNF␣ expression, the changes of which could be entirely normalized by the treatment with the iron chelator, L1 (25). In fact, these cells exhibit a heightened rise in the labile pool of iron after LPS stimulation as compared with the cells from control animals.…”

Risk factors and mechanisms of hepatocarcinogenesis with special emphasis on alcohol and oxidative stress

“…protein iron complexes, exacerbates oxidative damage by alcohol and that it stimulates hepatic macrophages to produce ROS and pro-inflammatory cytokines (Tsukamoto et al, 1999;Caro and Cederbaum, 2004). Although the mechanism of ethanol-associated hepatic iron deposition is still unclear, it has been speculated that oxidative modification of cytosolic iron regulatory protein 1 (IRP1) causes repression of ferritin synthesis and stimulation of transferring receptor synthesis, with increased iron uptake (Rouault, 2003).…”

Iron homeostasis and H63D mutations in alcoholics with and without liver disease

“…Iron is also believed to be central in the pathogenesis of ALD, and some reports show iron overload as a predictive indicator of higher mortality [15] , and development of hepatocellular carcinoma [16] . In fact, iron overload and alcohol have a synergistic effect on the production of oxidative stress [17][18][19][20] .…”