Autocrine formation of hepcidin induces iron retention in human monocytes

Abstract

Hepcidin, a master regulator of iron homeostasis, is produced in small amounts by inflammatory monocytes/macrophages. Chronic immune activation leads to iron retention within monocytes/macrophages and the development of anemia of chronic disease (ACD). We questioned whether monocyte-derived hepcidin exerts autocrine regulation toward cellular iron metabolism. Monocyte hepcidin mRNA expression was significantly induced within 3 hours after stimulation with LPS or IL-6, and hepcidin mRNA expression was significantly higher in monocytes of ACD patients than in controls. In ACD patients, monocyte hepcidin mRNA levels were significantly correlated to serum IL-6 concentrations, and increased monocyte hepcidin mRNA levels were associated with decreased expression of the iron exporter ferroportin and iron retention in these cells. Transient transfection experiments using a ferroportin/EmGFP fusion protein construct demonstrated that LPS inducible hepcidin expression in THP-1 monocytes resulted in internalization and degradation of ferroportin. Transfection of monocytes with siRNA directed against hepcidin almost fully reversed this lipopolysaccharide-mediated effect. Using ferroportin mutation constructs, we found that ferroportin is mainly targeted by hepcidin when expressed on the cell surface. Our results suggest that ferroportin expression in inflammatory monocytes is negatively affected by autocrine formation of hepcidin, thus contributing to iron sequestration within monocytes as found in ACD. IntroductionA dysregulated iron homeostasis is a cornerstone of acute and chronic inflammatory processes involving cell-mediated immunity and frequently leads to the development of anemia, termed as anemia of chronic disease (ACD), or anemia of inflammation. 1,2 ACD is a multifactorial disease, where immune mechanisms play key pathogenetic roles. These include cytokine-mediated suppression of erythropoiesis, 3,4 a blunted erythropoietin response, [5][6][7] and an increased iron accumulation in and a defective iron recycling from the reticuloendothelial system. [8][9][10][11][12][13] The liver-derived acute phase protein hepcidin, which is induced by cytokines and iron, plays a key role in this concert. 14,15 It causes anemia when overexpressed, 16,17 whereas hepcidin mutations lead to hepatic iron overload, 18,19 which can be referred to its regulatory effects on cellular iron efflux. This is exerted after binding of hepcidin to the only known cellular iron exporter ferroportin, [20][21][22] leading to ferroportin internalization and blockade of duodenal iron absorption and macrophage iron recycling. 23 Because hepcidin expression is induced by inflammatory stimuli, including interleukin-6 (IL-6) or lipopolysaccharide (LPS), [24][25][26][27][28][29] an increased expression of this acute phase protein has been found to be associated with macrophage iron retention in ACD patients. 30,31 In addition, hepcidinindependent inhibition of ferroportin mRNA expression by inflammatory cytokines also contributes to macrophage iron rete...

Differential regulation of iron homeostasis during human macrophage polarized activation

“…The post-translational inhibition of Fpn mediated by liver-derived hepcidin is probably the key regulatory mechanism of Fpn expression under inflammatory conditions in vivo [10]. Furthermore, studies showing TLR4-dependent hepcidin expression in mice myeloid cells [39] and that Fpn expression at the cell membrane in inflammatory macrophages is negatively affected by the autocrine up-regulation of hepcidin [40,41] indicate that ''local'' mechanisms may contribute to iron sequestration within monocytes. As our microarray analysis showed a slight difference in hepcidin mRNA levels, the divergent Fpn expression in polarized macrophages could be hepcidin-mediated, at least in part.…”

Regulation of Iron Metabolism by Hepcidin under Conditions of Inflammation

“…induced a more robust hepcidin induction than in controls, and this leads to diminished ferroportin expression and a decrease in iron export in an autocrine manner (79). Taken together, this implies that hepcidin production at the site of infection may lead to localized iron sequestration and prevent iron theft by pathogens; however, further research is required to understand this response to microbial invasion.…”

Interferon‐γ limits the availability of iron for intramacrophage Salmonella typhimurium

“…The intracellular iron content of RAW264.7 macrophage-like cells was determined by means of Graphite furnace atomic absorption spectrometry (GFAAS), carried out as described [65] on a Unicam Model Solaar, 939 QZ, atomic-absorption spectrometer using extended lifetime graphite tubes (Thermo Electron Corporation). Cells were treated exactly as described above, washed three times in normal saline and centrifuged at 300 Â g for 5 min.…”